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Sample records for sensitizes androgen-independent human

  1. Microwave mediated radiosynthesis of [F-18] FLT and its in-vitro study with androgen independent human prostate cancer cell line (PC-3)

    International Nuclear Information System (INIS)

    Ponde, D.E.; Dence, C.S.; Oyama, N.; Welch, M.J.

    2003-01-01

    The aim of this work was to improve the radiosynthesis of [F-18] FLT and to study its usefulness in monitoring change of proliferative activity of prostate cancer cells in the early phase of therapy. Method: Starting with anhydrothymidine, [F-18] FLT was synthesized by microwave mediated nucleophilic displacement by fluoride ion followed by acid hydrolysis in a synthesis time of just 55 minutes, which included Oasis solid phase and HPLC purification. The total radiochemical yield was 10-15% (at EOS), and the radiochemical purity was >99%. An in vitro study was carried out with androgen-independent human prostate cancer cell line PC-3. Two X 10e5 cells were seeded in 6 well plates with Ham's F-12K medium with 2 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate supplemented with 10% heat activated FBS. One day later, PC-3 cells were at 50% confluent, the media was removed and the cells divided into two groups. In one group, cells were suspended in fresh media as above with 10% FBS, whereas in the other group cells were suspended in fresh media as above but without serum. Twenty-four hours later, [F-18] FLT was added to each flask (n=3). The cell-associated uptake of [F-18] FLT at 37 deg C was determined at 0, 1, 3, and 6 h after incubation. [F-18] FLT uptake in PC-3 cells decreased by 55% (from 9% to 4%) after 24h incubation with serum free media, indicating its potential usefulness to monitor cell proliferation in androgen-independent human prostate cancer. Studies to ascertain the uptake-mechanism are in the way. NIH grant HL13851

  2. Co-Targeting Prostate Cancer Epithelium and Bone Stroma by Human Osteonectin-Promoter-Mediated Suicide Gene Therapy Effectively Inhibits Androgen-Independent Prostate Cancer Growth.

    Directory of Open Access Journals (Sweden)

    Shian-Ying Sung

    Full Text Available Stromal-epithelial interaction has been shown to promote local tumor growth and distant metastasis. We sought to create a promising gene therapy approach that co-targets cancer and its supporting stromal cells for combating castration-resistant prostate tumors. Herein, we demonstrated that human osteonectin is overexpressed in the prostate cancer epithelium and tumor stroma in comparison with their normal counterpart. We designed a novel human osteonectin promoter (hON-522E containing positive transcriptional regulatory elements identified in both the promoter and exon 1 region of the human osteonectin gene. In vitro reporter assays revealed that the hON-522E promoter is highly active in androgen receptor negative and metastatic prostate cancer and bone stromal cells compared to androgen receptor-positive prostate cancer cells. Moreover, in vivo prostate-tumor-promoting activity of the hON-522E promoter was confirmed by intravenous administration of an adenoviral vector containing the hON-522E promoter-driven luciferase gene (Ad-522E-Luc into mice bearing orthotopic human prostate tumor xenografts. In addition, an adenoviral vector with the hON-522E-promoter-driven herpes simplex virus thymidine kinase gene (Ad-522E-TK was highly effective against the growth of androgen-independent human prostate cancer PC3M and bone stromal cell line in vitro and in pre-established PC3M tumors in vivo upon addition of the prodrug ganciclovir. Because of the heterogeneity of human prostate tumors, hON-522E promoter-mediated gene therapy has the potential for the treatment of hormone refractory and bone metastatic prostate cancers.

  3. The PPARγ ligand ciglitazone regulates androgen receptor activation differently in androgen-dependent versus androgen-independent human prostate cancer cells

    International Nuclear Information System (INIS)

    Moss, Patrice E.; Lyles, Besstina E.; Stewart, LaMonica V.

    2010-01-01

    The androgen receptor (AR) regulates growth and progression of androgen-dependent as well as androgen-independent prostate cancer cells. Peroxisome proliferator-activated receptor gamma (PPARγ) agonists have been reported to reduce AR activation in androgen-dependent LNCaP prostate cancer cells. To determine whether PPARγ ligands are equally effective at inhibiting AR activity in androgen-independent prostate cancer, we examined the effect of the PPARγ ligands ciglitazone and rosiglitazone on C4-2 cells, an androgen- independent derivative of the LNCaP cell line. Luciferase-based reporter assays and Western blot analysis demonstrated that PPARγ ligand reduced dihydrotestosterone (DHT)-induced increases in AR activity in LNCaP cells. However, in C4-2 cells, these compounds increased DHT-induced AR driven luciferase activity. In addition, ciglitazone did not significantly alter DHT-mediated increases in prostate specific antigen (PSA) protein or mRNA levels within C4-2 cells. siRNA-based experiments demonstrated that the ciglitazone-induced regulation of AR activity observed in C4-2 cells was dependent on the presence of PPARγ. Furthermore, overexpression of the AR corepressor cyclin D1 inhibited the ability of ciglitazone to induce AR luciferase activity in C4-2 cells. Thus, our data suggest that both PPARγ and cyclin D1 levels influence the ability of ciglitazone to differentially regulate AR signaling in androgen-independent C4-2 prostate cancer cells.

  4. The PPAR{gamma} ligand ciglitazone regulates androgen receptor activation differently in androgen-dependent versus androgen-independent human prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Moss, Patrice E.; Lyles, Besstina E.; Stewart, LaMonica V., E-mail: lstewart@mmc.edu

    2010-12-10

    The androgen receptor (AR) regulates growth and progression of androgen-dependent as well as androgen-independent prostate cancer cells. Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) agonists have been reported to reduce AR activation in androgen-dependent LNCaP prostate cancer cells. To determine whether PPAR{gamma} ligands are equally effective at inhibiting AR activity in androgen-independent prostate cancer, we examined the effect of the PPAR{gamma} ligands ciglitazone and rosiglitazone on C4-2 cells, an androgen- independent derivative of the LNCaP cell line. Luciferase-based reporter assays and Western blot analysis demonstrated that PPAR{gamma} ligand reduced dihydrotestosterone (DHT)-induced increases in AR activity in LNCaP cells. However, in C4-2 cells, these compounds increased DHT-induced AR driven luciferase activity. In addition, ciglitazone did not significantly alter DHT-mediated increases in prostate specific antigen (PSA) protein or mRNA levels within C4-2 cells. siRNA-based experiments demonstrated that the ciglitazone-induced regulation of AR activity observed in C4-2 cells was dependent on the presence of PPAR{gamma}. Furthermore, overexpression of the AR corepressor cyclin D1 inhibited the ability of ciglitazone to induce AR luciferase activity in C4-2 cells. Thus, our data suggest that both PPAR{gamma} and cyclin D1 levels influence the ability of ciglitazone to differentially regulate AR signaling in androgen-independent C4-2 prostate cancer cells.

  5. Epigenetic repression of regulator of G-protein signaling 2 promotes androgen-independent prostate cancer cell growth.

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    Wolff, Dennis W; Xie, Yan; Deng, Caishu; Gatalica, Zoran; Yang, Mingjie; Wang, Bo; Wang, Jincheng; Lin, Ming-Fong; Abel, Peter W; Tu, Yaping

    2012-04-01

    G-protein-coupled receptor (GPCR)-stimulated androgen-independent activation of androgen receptor (AR) contributes to acquisition of a hormone-refractory phenotype by prostate cancer. We previously reported that regulator of G-protein signaling (RGS) 2, an inhibitor of GPCRs, inhibits androgen-independent AR activation (Cao et al., Oncogene 2006;25:3719-34). Here, we show reduced RGS2 protein expression in human prostate cancer specimens compared to adjacent normal or hyperplastic tissue. Methylation-specific PCR analysis and bisulfite sequencing indicated that methylation of the CpG island in the RGS2 gene promoter correlated with RGS2 downregulation in prostate cancer. In vitro methylation of this promoter suppressed reporter gene expression in transient transfection studies, whereas reversal of this promoter methylation with 5-aza-2'-deoxycytidine (5-Aza-dC) induced RGS2 reexpression in androgen-independent prostate cancer cells and inhibited their growth under androgen-deficient conditions. Interestingly, the inhibitory effect of 5-Aza-dC was significantly reduced by an RGS2-targeted short hairpin RNA, indicating that reexpressed RGS2 contributed to this growth inhibition. Restoration of RGS2 levels by ectopic expression in androgen-independent prostate cancer cells suppressed growth of xenografts in castrated mice. Thus, RGS2 promoter hypermethylation represses its expression and unmasks a latent pathway for AR transactivation in prostate cancer cells. Targeting this reversible process may provide a new strategy for suppressing prostate cancer progression by reestablishing its androgen sensitivity. Copyright © 2011 UICC.

  6. The HDL receptor SR-BI is associated with human prostate cancer progression and plays a possible role in establishing androgen independence.

    Science.gov (United States)

    Schörghofer, David; Kinslechner, Katharina; Preitschopf, Andrea; Schütz, Birgit; Röhrl, Clemens; Hengstschläger, Markus; Stangl, Herbert; Mikula, Mario

    2015-08-07

    Human prostate cancer represents one of the most frequently diagnosed cancers in men worldwide. Currently, diagnostic methods are insufficient to identify patients at risk for aggressive prostate cancer, which is essential for early treatment. Recent data indicate that elevated cholesterol levels in the plasma are a prerequisite for the progression of prostate cancer. Here, we analyzed clinical prostate cancer samples for the expression of receptors involved in cellular cholesterol uptake. We screened mRNA microarray files of prostate cancer samples for alterations in the expression levels of cholesterol transporters. Furthermore, we performed immunohistochemistry analysis on human primary prostate cancer tissue sections derived from patients to investigate the correlation of SR-BI with clinicopathological parameters and the mTOR target pS6. In contrast to LDLR, we identified SR-BI mRNA and protein expression to be induced in high Gleason grade primary prostate cancers. Histologic analysis of prostate biopsies revealed that 53.6 % of all cancer samples and none of the non-cancer samples showed high SR-BI staining intensity. The disease-free survival time was reduced (P = 0.02) in patients expressing high intra-tumor levels of SR-BI. SR-BI mRNA correlated with HSD17B1 and HSD3B1 and SR-BI protein staining showed correlation with active ribosomal protein S6 (RS = 0.828, P prostate cancer formation, suggesting that increased levels of SR-BI may be involved in the generation of a castration-resistant phenotype.

  7. Regulation of expression of Na+,K+-ATPase in androgen-dependent and androgen-independent prostate cancer

    NARCIS (Netherlands)

    L.J. Blok (Leen); G.T.G. Chang; M. Steenbeek-Slotboom (M.); W.M. van Weerden (Wytske); H.G. Swarts; J.J.H.H.M. de Pont (J. J H H M); G.J. van Steenbrugge (Gert Jan); A.O. Brinkmann (Albert)

    1999-01-01

    textabstractThe β1-subunit of Na+,K+-ATPase was isolated and identified as an androgen down-regulated gene. Expression was observed at high levels in androgen-independent as compared to androgen-dependent (responsive) human prostate cancer cell lines and xenografts when grown in the presence of

  8. Increased expression of heparin binding EGF (HB-EGF), amphiregulin, TGF alpha and epiregulin in androgen-independent prostate cancer cell lines.

    DEFF Research Database (Denmark)

    Tørring, Niels; Sørensen, Boe Sandahl; Nexø, Ebba

    2000-01-01

    BACKGROUND: The proliferation of androgen-independent prostate cancer cell lines has previously been shown to be influenced by an autocrine loop of the epidermal growth factor (EGF) system. This observation has alerted us to study the expression of ligands and receptors from the EGF......-system in prostate cell lines. METHODS: The expression of the EGF system was determined by quantitative RT-PCR and ELISA in the normal prostate epithelial cell line (PNT1A), in the androgen sensitive-(LNCaP), and the androgen-independent (DU145 and PC3) prostate cancer cell lines. RESULTS: The expression of m...... which exhibit low expression of HER1. Similar results were obtained by ELISA. CONCLUSIONS: The data indicates a selective up-regulation of a subclass of ligands of the EGF-system in androgen-independent prostate cancer cell lines. We suggest this could be a mechanism to escape androgen dependence...

  9. A polymorphism in a transporter of testosterone is a determinant of androgen independence in prostate cancer.

    Science.gov (United States)

    Sharifi, Nima; Hamada, Akinobu; Sissung, Tristan; Danesi, Romano; Venzon, David; Baum, Caitlin; Gulley, James L; Price, Douglas K; Dahut, William L; Figg, William D

    2008-08-05

    To determine if patients with advanced prostate cancer carrying a polymorphism that codes for a more active testosterone transporter have less durable responses to androgen-deprivation therapy (ADT) than patients not carrying this polymorphism. We previously determined that a polymorphism in SLCO1B3 affects testosterone transport and that those men who have at least one wild-type T allele at the 334 T > G polymorphism in this gene have a shorter survival. We hypothesized that the T allele which increases testosterone transport would be associated with a shorter interval from ADT to androgen independence. We examined the association between this SLCO1B3 polymorphism and time from ADT to androgen independence, ADT to prostate-specific antigen (PSA) nadir and PSA nadir to androgen independence in 68 Caucasian patients with advanced prostate cancer who were treated with ADT with metastatic disease (D2) or biochemical failure with no metastatic disease (D0). When examined separately, patients in the individual stages tended to have a shorter time to androgen independence with the T allele in the D0 (P = 0.11) and D2 (P = 0.18) groups. Combining these groups and stratifying by stage yielded a statistically significant shorter time to androgen independence with the T allele (P = 0.048). A polymorphism in a transporter that increases testosterone import is associated with a shorter time to androgen independence in patients with prostate cancer who are treated with ADT.

  10. Inhibition of progression of androgen-dependent prostate LNCaP tumors to androgen independence in SCID mice by oral caffeine and voluntary exercise.

    Science.gov (United States)

    Zheng, Xi; Cui, Xiao-Xing; Huang, Mou-Tuan; Liu, Yue; Wagner, George C; Lin, Yong; Shih, Weichung Joe; Lee, Mao-Jung; Yang, Chung S; Conney, Allan H

    2012-01-01

    The effect of oral caffeine or voluntary running wheel exercise (RW) alone or in combination on the progression of human androgen-dependent LNCaP prostate tumors to androgen independence in male severe combined immunodeficiency mice was determined. The mice were injected subcutaneously with LNCaP cells, and when the tumors reached a moderate size, the mice were surgically castrated and treated with caffeine (0.40 mg/ml drinking water) or RW alone or in combination for 42 days. We found that caffeine administration or RW inhibited the progression and growth of androgen-dependent LNCaP tumors to androgen independence, and a combination of the 2 regimens was more effective than the individual regimens alone. The ratios of the percent mitotic cells/caspase-3 positive cells in tumors from the caffeine-treated, RW-treated, or combination-treated mice were decreased by 34%, 38%, and 52%, respectively. Caffeine treatment increased the percentage of mitotic tumor cells undergoing apoptosis (lethal mitosis) whereas RW inhibited the increase in interleukin-6 that occurred during the progression of LNCaP tumors from androgen dependence to androgen independence. Our results indicate that oral administration of caffeine in combination with voluntary exercise may be an effective strategy for the prevention of prostate cancer progression from androgen dependence to androgen independence.

  11. Expression of a hyperactive androgen receptor leads to androgen-independent growth of prostate cancer cells.

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    Hsieh, Chen-Lin; Cai, Changmeng; Giwa, Ahmed; Bivins, Aaronica; Chen, Shao-Yong; Sabry, Dina; Govardhan, Kumara; Shemshedini, Lirim

    2008-07-01

    Cellular changes that affect the androgen receptor (AR) can cause prostate cancer to transition from androgen dependent to androgen independent, which is usually lethal. One common change in prostate tumors is overexpression of the AR, which has been shown to lead to androgen-independent growth of prostate cancer cells. This led us to hypothesize that expression of a hyperactive AR would be sufficient for androgen-independent growth of prostate cancer cells. To test this hypothesis, stable lune cancer prostate (LNCaP) cell lines were generated, which express a virion phosphoprotein (VP)16-AR hybrid protein that contains full-length AR fused to the strong viral transcriptional activation domain VP16. This fusion protein elicited as much as a 20-fold stronger transcriptional activity than the natural AR. Stable expression of VP16-AR in LNCaP cells yielded androgen-independent cell proliferation, while under the same growth conditions the parental LNCaP cells exhibited only androgen-dependent growth. These results show that expression of a hyperactive AR is sufficient for androgen-independent growth of prostate cancer cells. To study the molecular basis of this enhanced growth, we measured the expression of soluble guanylyl cyclase-alpha1 (sGCalpha1), a subunit of the sGC, an androgen-regulated gene that has been shown to be involved in prostate cancer cell growth. Interestingly, the expression of sGCalpha1 is androgen independent in VP16-AR-expressing cells, in contrast to its androgen-induced expression in control LNCaP cells. RNA(I)-dependent inhibition of sGCalpha1 expression resulted in significantly reduced proliferation of VP16-AR cells, implicating an important role for sGCalpha1 in the androgen-independent growth of these cells.

  12. Phase I trial of yttrium-90-labeled anti-prostate-specific membrane antigen monoclonal antibody J591 for androgen-independent prostate cancer.

    Science.gov (United States)

    Milowsky, Matthew I; Nanus, David M; Kostakoglu, Lale; Vallabhajosula, Shankar; Goldsmith, Stanley J; Bander, Neil H

    2004-07-01

    To determine the maximum-tolerated dose (MTD), toxicity, human antihuman antibody (HAHA) response, pharmacokinetics, organ dosimetry, targeting, and preliminary efficacy of yttrium-90-labeled anti-prostate-specific membrane antigen monoclonal antibody J591 ((90)Y-J591) in patients with androgen-independent prostate cancer (PC). Patients with androgen-independent PC and evidence of disease progression received indium-111-J591 for pharmacokinetic and biodistribution determinations followed 1 week later by (90)Y-J591 at five dose levels: 5, 10, 15, 17.5, and 20 mCi/m(2). Patients were eligible for up to three re-treatments if platelet and neutrophil recovery was satisfactory. Twenty-nine patients with androgen-independent PC received (90)Y-J591, four of whom were re-treated. Dose limiting toxicity (DLT) was seen at 20 mCi/m(2), with two patients experiencing thrombocytopenia with non-life-threatening bleeding episodes requiring platelet transfusions. The 17.5-mCi/m(2) dose level was determined to be the MTD. No re-treated patients experienced DLT. Nonhematologic toxicity was not dose limiting. Targeting of known sites of bone and soft tissue metastases was seen in the majority of patients. No HAHA response was seen. Antitumor activity was seen, with two patients experiencing 85% and 70% declines in prostate-specific antigen (PSA) levels lasting 8 and 8.6 months, respectively, before returning to baseline. Both patients had objective measurable disease responses. An additional six patients (21%) experienced PSA stabilization. The recommended dose for (90)Y-J591 is 17.5 mCi/m(2). Acceptable toxicity, excellent targeting of known sites of PC metastases, and biologic activity in patients with androgen-independent PC warrant further investigation of (90)Y-J591 in the treatment of patients with PC.

  13. Evolving perspectives of the role of novel agents in androgen-independent prostate cancer

    Directory of Open Access Journals (Sweden)

    Sujith Kalmadi

    2008-01-01

    Full Text Available Metastatic androgen-independent prostate cancer presents an intriguing clinical challenge, with a subtle interaction between hormone-responsive and refractory tumor cell elements. The treatment of advanced prostate carcinoma, which had remained stagnant for several decades following the understanding of the link between androgenic stimulation and carcinogenesis, has now started to make steady headway with chemotherapy and targeted approaches. Metastatic prostate cancer is almost always treated with initial androgen deprivation, in various forms. However, despite such treatment androgen-independent prostate cancer cells eventually emerge and progress to threaten life. The therapeutic objectives for treatment of metastatic prostate cancer are to maintain the quality of life and prolong survival. The out-dated nihilistic dogma of deferring chemotherapy until the most advanced stages in advanced prostate cancer is now falling by the wayside with the development of newer effective, tolerable agents.

  14. Androgen-independent proliferation of LNCaP prostate cancer cells infected by xenotropic murine leukemia virus-related virus

    International Nuclear Information System (INIS)

    Kakoki, Katsura; Kamiyama, Haruka; Izumida, Mai; Yashima, Yuka; Hayashi, Hideki; Yamamoto, Naoki; Matsuyama, Toshifumi; Igawa, Tsukasa; Sakai, Hideki; Kubo, Yoshinao

    2014-01-01

    Highlights: • XMRV infection induces androgen-independent growth in LNCaP cells. • XMRV infection reduces expression of androgen receptor. • XMRV promotes appearance of androgen blocker-resistant prostate cancer cells. - Abstract: Xenotropic murine leukemia virus-related virus (XMRV) is a novel gammaretrovirus that was originally isolated from human prostate cancer. It is now believed that XMRV is not the etiologic agent of prostate cancer. An analysis of murine leukemia virus (MLV) infection in various human cell lines revealed that prostate cancer cell lines are preferentially infected by XMRV, and this suggested that XMRV infection may confer some sort of growth advantage to prostate cancer cell lines. To examine this hypothesis, androgen-dependent LNCaP cells were infected with XMRV and tested for changes in certain cell growth properties. We found that XMRV-infected LNCaP cells can proliferate in the absence of the androgen dihydrotestosterone. Moreover, androgen receptor expression is significantly reduced in XMRV-infected LNCaP cells. Such alterations were not observed in uninfected and amphotropic MLV-infected LNCaP cells. This finding explains why prostate cancer cell lines are preferentially infected with XMRV

  15. Activation of estrogen receptor beta (ERβ) regulates the expression of N-cadherin, E-cadherin and β-catenin in androgen-independent prostate cancer cells.

    Science.gov (United States)

    Silva, Rafael de Souza; Lombardi, Ana Paola G; de Souza, Deborah Simão; Vicente, Carolina M; Porto, Catarina S

    2018-03-01

    The aim of the present study was to investigate the impact of the activation of estrogen receptors on expression and localization of N-cadherin, E-cadherin and non-phosphorylated β-catenin in androgen-independent prostate cancer cells (PC-3 and DU-145) and in human post pubertal prostate epithelial cells (PNT1A). Expression of N-cadherin was detected in PNT1A and PC-3 cells, but not in DU-145 cells. E-cadherin was detected only in DU-145 cells and β-catenin was detected in all cells studied. N-cadherin and β-catenin were located preferentially in the cellular membrane of PNT1A cells and in the cytoplasm of PC-3 cells. E-cadherin and β-catenin were located preferentially in the cellular membrane of DU-145 cells. 17β-estradiol (E2) or the ERα-selective agonist PPT did not affect the content and localization of N-cadherin in PC-3 and PNT1A cells or E-cadherin in DU-145 cells. In PC-3 cells, ERβ-selective agonist DPN decreased the expression of N-cadherin. DPN-induced downregulation of N-cadherin was blocked by pretreatment with the ERβ-selective antagonist (PHTPP), indicating that ERβ1 is the upstream receptor regulating the expression of N-cadherin. In DU-145 cells, the activation of ERβ1 by DPN increased the expression of E-cadherin. Taken together, these results suggest that activation of ERβ1 is required to maintain an epithelial phenotype in PC-3 and DU-145 cells. The activation of ERβ1 also increased the expression of β-catenin in cytoplasm of PC-3 and in the cellular membrane of DU-145 cells. In conclusion, our results indicate differential expression and localization of N-cadherin, E-cadherin and β-catenin in androgen-independent prostate cancer cells. The reduction of N-cadherin content by activation of ERβ, exclusively observed in androgen-independent prostate cancer cells (PC-3), may be related to the activation of signaling pathways, such as the release of β-catenin into the cytoplasm, translocation of β-catenin to the nucleus and

  16. Quantitative PET Imaging with Novel HER3 Targeted Peptides Selected by Phage Display to Predict Androgen Independent Prostate Cancer Progression

    Science.gov (United States)

    2017-08-01

    Independent Prostate Cancer Progression PRINCIPAL INVESTIGATOR: Benjamin Larimer, PhD CONTRACTING ORGANIZATION: Massachusetts General Hospital Boston...3. DATES COVERED 1 Aug 2016 – 31 July 2017 4. TITLE AND SUBTITLE Cancer Progression 5a. CONTRACT NUMBER Quantitative PET Imaging with Novel HER3...Targeted Peptides Selected by Phage Display to Predict Androgen-Independent Prostate Cancer Progression 5b. GRANT NUMBER W81XWH-16-1-0447 5c

  17. Natural proteasome inhibitor celastrol suppresses androgen-independent prostate cancer progression by modulating apoptotic proteins and NF-kappaB.

    Directory of Open Access Journals (Sweden)

    Yao Dai

    Full Text Available Celastrol is a natural proteasome inhibitor that exhibits promising anti-tumor effects in human malignancies, especially the androgen-independent prostate cancer (AIPC with constitutive NF-κB activation. Celastrol induces apoptosis by means of proteasome inhibition and suppresses prostate tumor growth. However, the detailed mechanism of action remains elusive. In the current study, we aim to test the hypothesis that celastrol suppresses AIPC progression via inhibiting the constitutive NF-κB activity as well as modulating the Bcl-2 family proteins.We examined the efficacy of celastrol both in vitro and in vivo, and evaluated the role of NF-κB in celastrol-mediated AIPC regression. We found that celastrol inhibited cell proliferation in all three AIPC cell lines (PC-3, DU145 and CL1, with IC₅₀ in the range of 1-2 µM. Celastrol also suppressed cell migration and invasion. Celastrol significantly induced apoptosis as evidenced by increased sub-G1 population, caspase activation and PARP cleavage. Moreover, celastrol promoted cleavage of the anti-apoptotic protein Mcl-1 and activated the pro-apoptotic protein Noxa. In addition, celastrol rapidly blocked cytosolic IκBα degradation and nuclear translocation of RelA. Likewise, celastrol inhibited the expression of multiple NF-κB target genes that are involved in proliferation, invasion and anti-apoptosis. Celastrol suppressed AIPC tumor progression by inhibiting proliferation, increasing apoptosis and decreasing angiogenesis, in PC-3 xenograft model in nude mouse. Furthermore, increased cellular IκBα and inhibited expression of various NF-κB target genes were observed in tumor tissues.Our data suggest that, via targeting the proteasome, celastrol suppresses proliferation, invasion and angiogenesis by inducing the apoptotic machinery and attenuating constitutive NF-κB activity in AIPC both in vitro and in vivo. Celastrol as an active ingredient of traditional herbal medicine could thus be

  18. The Role of AKT in Androgen-Independent Progression of Human Prostate Cancer

    Science.gov (United States)

    2005-02-01

    myristoylated rapamycin-binding reversibly protecting tissue from apoptosis due to ischemic domain from FRAP/mTOR, called M-FRB, binds to lipid per- injury or...V[0__AAFX, BAD, GSK30 * Apoptosis M-FRB,2 - oT308 [ T3 NFP8B, FRAP, Rat - Apoptosls M-Akt v ,$473 M-APH.Akt F3-Akt L L ___ F3-APH.Akt :,,• , Akt-F3...vector. Twenty-four hours after transfection, cells were divided into aliquots 0 that were stimulated with sub-optimal levels (5 ng/ml) Vector M-Akt

  19. Ferruginol suppresses survival signaling pathways in androgen-independent human prostate cancer cells

    NARCIS (Netherlands)

    de Jesus, Marcelo Bispo; Zambuzzi, Willian Fernando; Ruela de Sousa, Roberta Regina; Areche, Carlos; Santos de Souza, Ana Carolina; Aoyama, Hiroshi; Schmeda-Hirschmann, Guillermo; Rodriguez, Jaime A.; Monteiro de Souza Brito, Alba Regina; Peppelenbosch, Maikel P.; den Hertog, Jeroen; de Paula, Eneida; Ferreira, Carmen Verissima

    Ferruginol, a bioactive compound isolated from a Chilean tree (Podocarpaceae), attracts attention as a consequence of its pharmacological properties, which include anti-fungal, anti-bacterial, cardioprotective, anti-oxidative, anti-plasmodial and anti-ulcerogenic actions. Nevertheless, the molecular

  20. Inhibition of Androgen-Independent Prostate Cancer by Estrogenic Compounds Is Associated with Increased Expression of Immune-Related Genes

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    Ilsa M. Coleman

    2006-10-01

    Full Text Available The clinical utility of estrogens for treating prostate cancer (CaP was established in the 1940s by Huggins. The classic model of the anti-CaP activity of estrogens postulates an indirect mechanism involving the suppression of androgen production. However, clinical, preclinical studies have shown that estrogens exert growth-inhibitory effects on CaP under low-androgen conditions, suggesting additional modes whereby estrogens affect CaP cells and/or the microenvironment. Here we have investigated the activity of 17β estradiol (E2 against androgen-independent CaP, identified molecular alterations in tumors exposed to E2. E2 treatment inhibited the growth of all four androgen-independent CaP xenografts studied (LuCaP 35V, LuCaP 23.1AI, LuCaP 49, LuCaP 58 in castrated male mice. The molecular basis of growth suppression was studied by cDNA microarray analysis, which indicated that multiple pathways are altered by E2 treatment. Of particular interest are changes in transcripts encoding proteins that mediate immune responses, regulate androgen receptor signaling. In conclusion, our data show that estrogens have powerful inhibitory effects on CaP in vivo in androgendepleted environments, suggest novel mechanisms of estrogen-mediated antitumor activity. These results indicate that incorporating estrogens into CaP treatment protocols could enhance therapeutic efficacy even in cases of advanced disease.

  1. Androgen receptor signaling is required for androgen-sensitive human prostate cancer cell proliferation and survival

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    Day Wanda V

    2005-04-01

    Full Text Available Abstract Background Androgens and androgen receptors (AR regulate normal prostate development and growth. They also are involved in pathological development of prostatic diseases, including benign prostatic hyperplasia (BPH and prostate cancer (PCa. Antiandrogen therapy for PCa, in conjunction with chemical or surgical castration, offers initial positive responses and leads to massive prostate cell death. However, cancer cells later appear as androgen-independent PCa. To investigate the role of AR in prostate cell proliferation and survival, we introduced a vector-based small interfering RNA (siRNA. This siRNA targeted 5'-untranslated region of AR mRNA for extended suppression of AR expression in androgen-sensitive human prostate LNCaP cells. Results The siRNA design successfully suppressed endogenous AR expression, as revealed by western blotting and immunofluorescence staining in LNCaP cells. LNCaP cells did not proliferate in the absence of AR and underwent apoptosis, based on elevated phospho-Histone H2B expression and higher number of apoptotic body as compared to control cells. Conclusion We demonstrated that AR is vital for prostate cell proliferation and survival in this androgen-sensitive prostate cell line. These results further strengthen the hypothesis that AR can be a therapeutic target for treating androgen-sensitive stages of PCa. Unlike antiandorgens, however, siRNA targeting AR provides a direct inactivation of AR function through the suppression of AR protein expression.

  2. Phenotypic Plasticity, Bet-Hedging, and Androgen Independence in Prostate Cancer: Role of Non-Genetic Heterogeneity

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    Mohit Kumar Jolly

    2018-03-01

    Full Text Available It is well known that genetic mutations can drive drug resistance and lead to tumor relapse. Here, we focus on alternate mechanisms—those without mutations, such as phenotypic plasticity and stochastic cell-to-cell variability that can also evade drug attacks by giving rise to drug-tolerant persisters. The phenomenon of persistence has been well-studied in bacteria and has also recently garnered attention in cancer. We draw a parallel between bacterial persistence and resistance against androgen deprivation therapy in prostate cancer (PCa, the primary standard care for metastatic disease. We illustrate how phenotypic plasticity and consequent mutation-independent or non-genetic heterogeneity possibly driven by protein conformational dynamics can stochastically give rise to androgen independence in PCa, and suggest that dynamic phenotypic plasticity should be considered in devising therapeutic dosing strategies designed to treat and manage PCa.

  3. Chemical Suppression of the Reactivated Androgen Signaling Pathway in Androgen-Independent Prostate Cancer

    Science.gov (United States)

    2011-07-01

    when it is ingested during pregnancy [20,21]. Aside from its role in development, Hh signaling also supports stem cells in adult tissues [22-24]. However...For the mo.~t commonly uti - lized human prostate cancer cell lines (LNCaP and derivatives, DUI45, PC3 or CWR22rvl) grown in culture, Shh, Glil/2 and

  4. Novel expressed sequences identified in a model of androgen independent prostate cancer

    Directory of Open Access Journals (Sweden)

    Jones Steven JM

    2007-01-01

    Full Text Available Abstract Background Prostate cancer is the most frequently diagnosed cancer in American men, and few effective treatment options are available to patients who develop hormone-refractory prostate cancer. The molecular changes that occur to allow prostate cells to proliferate in the absence of androgens are not fully understood. Results Subtractive hybridization experiments performed with samples from an in vivo model of hormonal progression identified 25 expressed sequences representing novel human transcripts. Intriguingly, these 25 sequences have small open-reading frames and are not highly conserved through evolution, suggesting many of these novel expressed sequences may be derived from untranslated regions of novel transcripts or from non-coding transcripts. Examination of a large metalibrary of human Serial Analysis of Gene Expression (SAGE tags demonstrated that only three of these novel sequences had been previously detected. RT-PCR experiments confirmed that the 6 sequences tested were expressed in specific human tissues, as well as in clinical samples of prostate cancer. Further RT-PCR experiments for five of these fragments indicated they originated from large untranslated regions of unannotated transcripts. Conclusion This study underlines the value of using complementary techniques in the annotation of the human genome. The tissue-specific expression of 4 of the 6 clones tested indicates the expression of these novel transcripts is tightly regulated, and future work will determine the possible role(s these novel transcripts may play in the progression of prostate cancer.

  5. Clinical evaluation of internal iliac artery anticancer drug infusion for the treatment of androgen-independent prostate cancer

    International Nuclear Information System (INIS)

    Cao Ye; Wang Jin; Nie Yong; Chen Hua; Huang Xinjie

    2008-01-01

    Objective: To evaluate the clinical efficacy of bilateral internal iliac artery chemotherapy infusion for the treatment of androgen-independent prostate cancer (ALPC). Methods: Thirty eight eases of confirmed AIPC were randomly divided into treatment group and control group. The patients in treatment group (23 cases) were treated with androgen deprivation therapy and regular internal iliac artery chemotherapy, while patients in control group (15 cases) were only received androgen deprivation therapy. The therapeutic efficacies of the two groups were compared and analyzed after completion of the treatment. Results: The clinical symptoms and maximum urine flow rates of' treatment group were improved rapidly 6 months later. After 2 years follow-up, the total efficacies of treatment group and control group were 65.2% and 26.7% respectively, showing a significant statistical difference (P<0.05). Conclusions: The treatment of AlPC with bilateral internal iliac artery chemotherapy is effective, providing melioration the quality of life and alleviation of the symptoms. (authors)

  6. Androgen and taxol cause cell type-specific alterations of centrosome and DNA organization in androgen-responsive LNCaP and androgen-independent DU145 prostate cancer cells

    Science.gov (United States)

    Schatten, H.; Ripple, M.; Balczon, R.; Weindruch, R.; Chakrabarti, A.; Taylor, M.; Hueser, C. N.

    2000-01-01

    We investigated the effects of androgen and taxol on the androgen-responsive LNCaP and androgen-independent DU145 prostate cancer cell lines. Cells were treated for 48 and 72 h with 0.05-1 nM of the synthetic androgen R1881 and with 100 nM taxol. Treatment of LNCaP cells with 0.05 nM R1881 led to increased cell proliferation, whereas treatment with 1 nM R1881 resulted in inhibited cell division, DNA cycle arrest, and altered centrosome organization. After treatment with 1 nM R1881, chromatin became clustered, nuclear envelopes convoluted, and mitochondria accumulated around the nucleus. Immunofluorescence microscopy with antibodies to centrosomes showed altered centrosome structure. Although centrosomes were closely associated with the nucleus in untreated cells, they dispersed into the cytoplasm after treatment with 1 nM R1881. Microtubules were only faintly detected in 1 nM R1881-treated LNCaP cells. The effects of taxol included microtubule bundling and altered mitochondria morphology, but not DNA organization. As expected, the androgen-independent prostate cancer cell line DU145 was not affected by R1881. Treatment with taxol resulted in bundling of microtubules in both cell lines. Additional taxol effects were seen in DU145 cells with micronucleation of DNA, an indication of apoptosis. Simultaneous treatment with R1881 and taxol had no additional effects on LNCaP or DU145 cells. These results suggest that LNCaP and DU145 prostate cancer cells show differences not only in androgen responsiveness but in sensitivity to taxol as well. Copyright 2000 Wiley-Liss, Inc.

  7. Quantitative PET Imaging with Novel HER3-Targeted Peptides Selected by Phage Display to Predict Androgen-Independent Prostate Cancer Progression

    Science.gov (United States)

    2017-12-01

    Independent Prostate Cancer Progression PRINCIPAL INVESTIGATOR: Benjamin Larimer, PhD CONTRACTING ORGANIZATION: Massachusetts General Hospital Boston...TYPE Final 3. DATES COVERED 1 Aug 2016 – 19 August 2017 Selected by Phage Display to Predict Androgen-Independent Prostate Cancer Progression 5a...highly specific peptide that targets HER3 for prostate cancer imaging. The peptide was labeled with a PET imaging radionuclide and injected into mice

  8. HOXB13 promotes androgen independent growth of LNCaP prostate cancer cells by the activation of E2F signaling

    Directory of Open Access Journals (Sweden)

    Choi Chan

    2010-05-01

    Full Text Available Abstract Background Androgen signaling plays a critical role in the development of prostate cancer and its progression. However, androgen-independent prostate cancer cells emerge after hormone ablation therapy, resulting in significant clinical problems. We have previously demonstrated that the HOXB13 homeodomain protein functions as a prostate cancer cell growth suppressor by inhibiting androgen-mediated signals. However, the role of the HOXB13 in androgen-independent growth of prostate cancer cells remains unexplained. Results In this report, we first demonstrated that HOXB13 was highly overexpressed in hormone-refractory tumors compared to tumors without prostate-specific antigen after initial treatment. Functionally, in an androgen-free environment minimal induction of HOXB13 in LNCaP prostate cancer cells, to the level of the normal prostate, markedly promoted cell proliferation while suppression inhibited cell proliferation. The HOXB13-mediated cell growth promotion in the absence of androgen, appears to be mainly accomplished through the activation of RB-E2F signaling by inhibiting the expression of the p21waf tumor suppressor. Indeed, forced expression of HOXB13 dramatically decreased expression of p21waf; this inhibition largely affected HOXB13-mediated promotion of E2F signaling. Conclusions Taken together, the results of this study demonstrated the presence of a novel pathway that helps understand androgen-independent survival of prostate cancer cells. These findings suggest that upregulation of HOXB13 is associated with an additive growth advantage of prostate cancer cells in the absence of or low androgen concentrations, by the regulation of p21-mediated E2F signaling.

  9. Withaferin A Induces Cell Death Selectively in Androgen-Independent Prostate Cancer Cells but Not in Normal Fibroblast Cells.

    Directory of Open Access Journals (Sweden)

    Yukihiro Nishikawa

    Full Text Available Withaferin A (WA, a major bioactive component of the Indian herb Withania somnifera, induces cell death (apoptosis/necrosis in multiple types of tumor cells, but the molecular mechanism underlying this cytotoxicity remains elusive. We report here that 2 μM WA induced cell death selectively in androgen-insensitive PC-3 and DU-145 prostate adenocarcinoma cells, whereas its toxicity was less severe in androgen-sensitive LNCaP prostate adenocarcinoma cells and normal human fibroblasts (TIG-1 and KD. WA also killed PC-3 cells in spheroid-forming medium. DNA microarray analysis revealed that WA significantly increased mRNA levels of c-Fos and 11 heat-shock proteins (HSPs in PC-3 and DU-145, but not in LNCaP and TIG-1. Western analysis revealed increased expression of c-Fos and reduced expression of the anti-apoptotic protein c-FLIP(L. Expression of HSPs such as HSPA6 and Hsp70 was conspicuously elevated; however, because siRNA-mediated depletion of HSF-1, an HSP-inducing transcription factor, reduced PC-3 cell viability, it is likely that these heat-shock genes were involved in protecting against cell death. Moreover, WA induced generation of reactive oxygen species (ROS in PC-3 and DU-145, but not in normal fibroblasts. Immunocytochemistry and immuno-electron microscopy revealed that WA disrupted the vimentin cytoskeleton, possibly inducing the ROS generation, c-Fos expression and c-FLIP(L suppression. These observations suggest that multiple events followed by disruption of the vimentin cytoskeleton play pivotal roles in WA-mediated cell death.

  10. Radiation sensitivity of human malignant lymphocytes

    International Nuclear Information System (INIS)

    Seshadri, R.; Matthews, C.; Morley, A.A.

    1985-01-01

    A simple and rapid in vitro technique to assess the sensitivity of human malignant lymphocytes to roentgen irradiation is described. A variety of established malignant lymphocyte cell lines were cloned in microwells and clone survival was used as the end-point. The survival of the clonogenic malignant lymphocyte down to a fraction of approximately 0.001 could be measured accurately. Except for a T-cell line, the radiation sensitivities of the cell lines were similar to that of normal T-lymphocytes. (orig.)

  11. Human sensitivity to vertical self-motion.

    Science.gov (United States)

    Nesti, Alessandro; Barnett-Cowan, Michael; Macneilage, Paul R; Bülthoff, Heinrich H

    2014-01-01

    Perceiving vertical self-motion is crucial for maintaining balance as well as for controlling an aircraft. Whereas heave absolute thresholds have been exhaustively studied, little work has been done in investigating how vertical sensitivity depends on motion intensity (i.e., differential thresholds). Here we measure human sensitivity for 1-Hz sinusoidal accelerations for 10 participants in darkness. Absolute and differential thresholds are measured for upward and downward translations independently at 5 different peak amplitudes ranging from 0 to 2 m/s(2). Overall vertical differential thresholds are higher than horizontal differential thresholds found in the literature. Psychometric functions are fit in linear and logarithmic space, with goodness of fit being similar in both cases. Differential thresholds are higher for upward as compared to downward motion and increase with stimulus intensity following a trend best described by two power laws. The power laws' exponents of 0.60 and 0.42 for upward and downward motion, respectively, deviate from Weber's Law in that thresholds increase less than expected at high stimulus intensity. We speculate that increased sensitivity at high accelerations and greater sensitivity to downward than upward self-motion may reflect adaptations to avoid falling.

  12. Transfected poly(I:C) activates different dsRNA receptors, leading to apoptosis or immunoadjuvant response in androgen-independent prostate cancer cells.

    Science.gov (United States)

    Palchetti, Sara; Starace, Donatella; De Cesaris, Paola; Filippini, Antonio; Ziparo, Elio; Riccioli, Anna

    2015-02-27

    Despite the effectiveness of surgery or radiation therapy for the treatment of early-stage prostate cancer (PCa), there is currently no effective strategy for late-stage disease. New therapeutic targets are emerging; in particular, dsRNA receptors Toll-like receptor 3 (TLR3) and cytosolic helicases expressed by cancer cells, once activated, exert a pro-apoptotic effect in different tumors. We previously demonstrated that the synthetic analog of dsRNA poly(I:C) induces apoptosis in the androgen-dependent PCa cell line LNCaP in a TLR3-dependent fashion, whereas only a weak apoptotic effect is observed in the more aggressive and androgen-independent PCa cells PC3 and DU145. In this paper, we characterize the receptors and the signaling pathways involved in the remarkable apoptosis induced by poly(I:C) transfected by Lipofectamine (in-poly(I:C)) compared with the 12-fold higher free poly(I:C) concentration in PC3 and DU145 cells. By using genetic inhibition of different poly(I:C) receptors, we demonstrate the crucial role of TLR3 and Src in in-poly(I:C)-induced apoptosis. Therefore, we show that the increased in-poly(I:C) apoptotic efficacy is due to a higher binding of endosomal TLR3. On the other hand, we show that in-poly(I:C) binding to cytosolic receptors MDA5 and RIG-I triggers IRF3-mediated signaling, leading uniquely to the up-regulation of IFN-β, which likely in turn induces increased TLR3, MDA5, and RIG-I proteins. In summary, in-poly(I:C) activates two distinct antitumor pathways in PC3 and DU145 cells: one mediated by the TLR3/Src/STAT1 axis, leading to apoptosis, and the other one mediated by MDA5/RIG-I/IRF3, leading to immunoadjuvant IFN-β expression. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Genetics of human sensitivity to ultraviolet radiation

    Science.gov (United States)

    Cleaver, James E.

    1994-07-01

    the major human health effects of solar and artificial UV light occur from the UVB and UVC wavelength ranges and involve a variety of short-term and long-term deleterious changes to the skin and eyes. the more important initial damage to cellular macromolecules involves dimerization of adjacent pyrimidines in DNA to produce cyclobutane pyrimidine dimes, (6-4) pyrimidine- pyrimidone, and (6-4) dewar photoproducts. these photoproducts can be repaired by a genetically regulated enzyme system (nucleotide excision repair) which removes oligonucleotides 29-30 nucleotides long that contain the photoproducts, and synthesizes replacement patches. At least a dozen gene products are involved in the process of recognizing photoproducts in DNA, altering local DNA helicity and cleaving the polynucleotide chain at defined positions either side of a photoproduct. Hereditary mutations in many of these genes are recognized in the human genetic disorders xeroderma pigmentosum (XP), Cockayne syndrome (CS), and trichothiodystrophy (TTD). Several of the gene products have other functions involving the regulation of gene transcription which accounts for the complex clinical presentation of repair deficient diseases that involve sensitivity of the skin and eyes to UV light, increased solar carcinogenesis (in XP), demyelination, and ganglial calcification (in CS), hair abnormalities (in TTD), and developmental and neurological abnormalities

  14. Cyproterone acetate enhances TRAIL-induced androgen-independent prostate cancer cell apoptosis via up-regulation of death receptor 5.

    Science.gov (United States)

    Chen, Linjie; Wolff, Dennis W; Xie, Yan; Lin, Ming-Fong; Tu, Yaping

    2017-03-07

    Virtually all prostate cancer deaths occur due to obtaining the castration-resistant phenotype after prostate cancer cells escaped from apoptosis and/or growth suppression initially induced by androgen receptor blockade. TNF-related apoptosis-inducing ligand (TRAIL) was an attractive cancer therapeutic agent due to its minimal toxicity to normal cells and remarkable apoptotic activity in tumor cells. However, most localized cancers including prostate cancer are resistant to TRAIL-induced apoptosis, thereby creating a therapeutic challenge of inducing TRAIL sensitivity in cancer cells. Herein the effects of cyproterone acetate, an antiandrogen steroid, on the TRAIL-induced apoptosis of androgen receptor-negative prostate cancer cells are reported. Cell apoptosis was assessed by both annexin V/propidium iodide labeling and poly (ADP-ribose) polymerase cleavage assays. Gene and protein expression changes were determined by quantitative real-time PCR and western blot assays. The effect of cyproterone acetate on gene promoter activity was determined by luciferase reporter assay. Cyproterone acetate but not AR antagonist bicalutamide dramatically increased the susceptibility of androgen receptor-negative human prostate cancer PC-3 and DU145 cells to TRAIL-induced apoptosis but no effects on immortalized human prostate stromal PS30 cells and human embryonic kidney HEK293 cells. Further investigation of the TRAIL-induced apoptosis pathway revealed that cyproterone acetate exerted its effect by selectively increasing death receptor 5 (DR5) mRNA and protein expression. Cyproterone acetate treatment also increased DR5 gene promoter activity, which could be abolished by mutation of a consensus binding domain of transcription factor CCAAT-enhancer-binding protein homologous protein (CHOP) in the DR5 gene promoter. Cyproterone acetate increases CHOP expression in a concentration and time-dependent manner and endoplasmic reticulum stress reducer 4-phenylbutyrate could block

  15. Proteasomal degradation of sphingosine kinase 1 and inhibition of dihydroceramide desaturase by the sphingosine kinase inhibitors, SKi or ABC294640, induces growth arrest in androgen-independent LNCaP-AI prostate cancer cells.

    Science.gov (United States)

    McNaughton, Melissa; Pitman, Melissa; Pitson, Stuart M; Pyne, Nigel J; Pyne, Susan

    2016-03-29

    Sphingosine kinases (two isoforms termed SK1 and SK2) catalyse the formation of the bioactive lipid sphingosine 1-phosphate. We demonstrate here that the SK2 inhibitor, ABC294640 (3-(4-chlorophenyl)-adamantane-1-carboxylic acid (pyridin-4-ylmethyl)amide) or the SK1/SK2 inhibitor, SKi (2-(p-hydroxyanilino)-4-(p-chlorophenyl)thiazole)) induce the proteasomal degradation of SK1a (Mr = 42 kDa) and inhibit DNA synthesis in androgen-independent LNCaP-AI prostate cancer cells. These effects are recapitulated by the dihydroceramide desaturase (Des1) inhibitor, fenretinide. Moreover, SKi or ABC294640 reduce Des1 activity in Jurkat cells and ABC294640 induces the proteasomal degradation of Des1 (Mr = 38 kDa) in LNCaP-AI prostate cancer cells. Furthermore, SKi or ABC294640 or fenretinide increase the expression of the senescence markers, p53 and p21 in LNCaP-AI prostate cancer cells. The siRNA knockdown of SK1 or SK2 failed to increase p53 and p21 expression, but the former did reduce DNA synthesis in LNCaP-AI prostate cancer cells. Moreover, N-acetylcysteine (reactive oxygen species scavenger) blocked the SK inhibitor-induced increase in p21 and p53 expression but had no effect on the proteasomal degradation of SK1a. In addition, siRNA knockdown of Des1 increased p53 expression while a combination of Des1/SK1 siRNA increased the expression of p21. Therefore, Des1 and SK1 participate in regulating LNCaP-AI prostate cancer cell growth and this involves p53/p21-dependent and -independent pathways. Therefore, we propose targeting androgen-independent prostate cancer cells with compounds that affect Des1/SK1 to modulate both de novo and sphingolipid rheostat pathways in order to induce growth arrest.

  16. An in vitro human skin test for assessing sensitization potential.

    Science.gov (United States)

    Ahmed, S S; Wang, X N; Fielding, M; Kerry, A; Dickinson, I; Munuswamy, R; Kimber, I; Dickinson, A M

    2016-05-01

    Sensitization to chemicals resulting in an allergy is an important health issue. The current gold-standard method for identification and characterization of skin-sensitizing chemicals was the mouse local lymph node assay (LLNA). However, for a number of reasons there has been an increasing imperative to develop alternative approaches to hazard identification that do not require the use of animals. Here we describe a human in-vitro skin explant test for identification of sensitization hazards and the assessment of relative skin sensitizing potency. This method measures histological damage in human skin as a readout of the immune response induced by the test material. Using this approach we have measured responses to 44 chemicals including skin sensitizers, pre/pro-haptens, respiratory sensitizers, non-sensitizing chemicals (including skin-irritants) and previously misclassified compounds. Based on comparisons with the LLNA, the skin explant test gave 95% specificity, 95% sensitivity, 95% concordance with a correlation coefficient of 0.9. The same specificity and sensitivity were achieved for comparison of results with published human sensitization data with a correlation coefficient of 0.91. The test also successfully identified nickel sulphate as a human skin sensitizer, which was misclassified as negative in the LLNA. In addition, sensitizers and non-sensitizers identified as positive or negative by the skin explant test have induced high/low T cell proliferation and IFNγ production, respectively. Collectively, the data suggests the human in-vitro skin explant test could provide the basis for a novel approach for characterization of the sensitizing activity as a first step in the risk assessment process. Copyright © 2015 John Wiley & Sons, Ltd.

  17. Oncolytic targeting of androgen-sensitive prostate tumor by the respiratory syncytial virus (RSV): consequences of deficient interferon-dependent antiviral defense

    International Nuclear Information System (INIS)

    Echchgadda, Ibtissam; Chang, Te-Hung; Sabbah, Ahmed; Bakri, Imad; Ikeno, Yuji; Hubbard, Gene B; Chatterjee, Bandana; Bose, Santanu

    2011-01-01

    Oncolytic virotherapy for cancer treatment utilizes viruses for selective infection and death of cancer cells without any adverse effect on normal cells. We previously reported that the human respiratory syncytial virus (RSV) is a novel oncolytic virus against androgen-independent PC-3 human prostate cancer cells. The present study extends the result to androgen-dependent prostate cancer, and explores the underlying mechanism that triggers RSV-induced oncolysis of prostate cancer cells. The oncolytic effect of RSV on androgen-sensitive LNCaP human prostate cancer cells and on androgen-independent RM1 murine prostate cancer cells was studied in vitro in culture and in vivo in a xenograft or allograft tumor model. In vitro, cell viability, infectivity and apoptosis were monitored by MTT assay, viral plaque assay and annexin V staining, respectively. In vivo studies involved virus administration to prostate tumors grown in immune compromised nude mice and in syngeneic immune competent C57BL/6J mice. Anti-tumorogenic oncolytic activity was monitored by measuring tumor volume, imaging bioluminescent tumors in live animals and performing histopathological analysis and TUNEL assay with tumors We show that RSV imposes a potent oncolytic effect on LNCaP prostate cancer cells. RSV infectivity was markedly higher in LNCaP cells compared to the non-tumorigenic RWPE-1 human prostate cells. The enhanced viral burden led to LNCaP cell apoptosis and growth inhibition of LNCaP xenograft tumors in nude mice. A functional host immune response did not interfere with RSV-induced oncolysis, since growth of xenograft tumors in syngeneic C57BL/6J mice from murine RM1 cells was inhibited upon RSV administration. LNCaP cells failed to activate the type-I interferon (IFNα/β)-induced transcription factor STAT-1, which is required for antiviral gene expression, although these cells could produce IFN in response to RSV infection. The essential role of IFN in restricting infection was further

  18. Effect of aberrations in human eye on contrast sensitivity function

    Science.gov (United States)

    Quan, Wei; Wang, Feng-lin; Wang, Zhao-qi

    2011-06-01

    The quantitative analysis of the effect of aberrations in human eye on vision has important clinical value in the correction of aberrations. The wave-front aberrations of human eyes were measured with the Hartmann-Shack wave-front sensor and modulation transfer function (MTF) was computed from the wave-front aberrations. Contrast sensitivity function (CSF) was obtained from MTF and the retinal aerial image modulation (AIM). It is shown that the 2nd, 3rd, 4th, 5th, 6th Zernike aberrations deteriorate contrast sensitivity function. When the 2nd, 3rd, 4th, 5th, 6th Zernike aberrations are corrected high contrast sensitivity function can be obtained.

  19. Multivariate Models for Prediction of Human Skin Sensitization ...

    Science.gov (United States)

    One of the lnteragency Coordinating Committee on the Validation of Alternative Method's (ICCVAM) top priorities is the development and evaluation of non-animal approaches to identify potential skin sensitizers. The complexity of biological events necessary to produce skin sensitization suggests that no single alternative method will replace the currently accepted animal tests. ICCVAM is evaluating an integrated approach to testing and assessment based on the adverse outcome pathway for skin sensitization that uses machine learning approaches to predict human skin sensitization hazard. We combined data from three in chemico or in vitro assays - the direct peptide reactivity assay (DPRA), human cell line activation test (h-CLAT) and KeratinoSens TM assay - six physicochemical properties and an in silico read-across prediction of skin sensitization hazard into 12 variable groups. The variable groups were evaluated using two machine learning approaches , logistic regression and support vector machine, to predict human skin sensitization hazard. Models were trained on 72 substances and tested on an external set of 24 substances. The six models (three logistic regression and three support vector machine) with the highest accuracy (92%) used: (1) DPRA, h-CLAT and read-across; (2) DPRA, h-CLAT, read-across and KeratinoSens; or (3) DPRA, h-CLAT, read-across, KeratinoSens and log P. The models performed better at predicting human skin sensitization hazard than the murine

  20. Sensitive chain specific radioimmunoassay for human immunoglobulins using monoclonal antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Sikora, K; Alderson, T St.J.; Ellis, J [Ludwig Institute for Cancer Research, Cambridge (UK)

    1983-02-25

    A sensitive radioimmunoassay is described for human immunoglobulins. This solid-phase assay uses commercially available monoclonal antibodies and is specific for different Ig chain types. Levels of less than 20 ng/ml Ig are detectable. The assay is suitable for the analysis of human hybridoma supernatants.

  1. Sensitivity of risk parameters to human errors for a PWR

    International Nuclear Information System (INIS)

    Samanta, P.; Hall, R.E.; Kerr, W.

    1980-01-01

    Sensitivities of the risk parameters, emergency safety system unavailabilities, accident sequence probabilities, release category probabilities and core melt probability were investigated for changes in the human error rates within the general methodological framework of the Reactor Safety Study for a Pressurized Water Reactor (PWR). Impact of individual human errors were assessed both in terms of their structural importance to core melt and reliability importance on core melt probability. The Human Error Sensitivity Assessment of a PWR (HESAP) computer code was written for the purpose of this study

  2. Multivariate Models for Prediction of Human Skin Sensitization Hazard

    Science.gov (United States)

    Strickland, Judy; Zang, Qingda; Paris, Michael; Lehmann, David M.; Allen, David; Choksi, Neepa; Matheson, Joanna; Jacobs, Abigail; Casey, Warren; Kleinstreuer, Nicole

    2016-01-01

    One of ICCVAM’s top priorities is the development and evaluation of non-animal approaches to identify potential skin sensitizers. The complexity of biological events necessary to produce skin sensitization suggests that no single alternative method will replace the currently accepted animal tests. ICCVAM is evaluating an integrated approach to testing and assessment based on the adverse outcome pathway for skin sensitization that uses machine learning approaches to predict human skin sensitization hazard. We combined data from three in chemico or in vitro assays—the direct peptide reactivity assay (DPRA), human cell line activation test (h-CLAT), and KeratinoSens™ assay—six physicochemical properties, and an in silico read-across prediction of skin sensitization hazard into 12 variable groups. The variable groups were evaluated using two machine learning approaches, logistic regression (LR) and support vector machine (SVM), to predict human skin sensitization hazard. Models were trained on 72 substances and tested on an external set of 24 substances. The six models (three LR and three SVM) with the highest accuracy (92%) used: (1) DPRA, h-CLAT, and read-across; (2) DPRA, h-CLAT, read-across, and KeratinoSens; or (3) DPRA, h-CLAT, read-across, KeratinoSens, and log P. The models performed better at predicting human skin sensitization hazard than the murine local lymph node assay (accuracy = 88%), any of the alternative methods alone (accuracy = 63–79%), or test batteries combining data from the individual methods (accuracy = 75%). These results suggest that computational methods are promising tools to effectively identify potential human skin sensitizers without animal testing. PMID:27480324

  3. A non-human primate model for gluten sensitivity.

    Directory of Open Access Journals (Sweden)

    Michael T Bethune

    2008-02-01

    Full Text Available Gluten sensitivity is widespread among humans. For example, in celiac disease patients, an inflammatory response to dietary gluten leads to enteropathy, malabsorption, circulating antibodies against gluten and transglutaminase 2, and clinical symptoms such as diarrhea. There is a growing need in fundamental and translational research for animal models that exhibit aspects of human gluten sensitivity.Using ELISA-based antibody assays, we screened a population of captive rhesus macaques with chronic diarrhea of non-infectious origin to estimate the incidence of gluten sensitivity. A selected animal with elevated anti-gliadin antibodies and a matched control were extensively studied through alternating periods of gluten-free diet and gluten challenge. Blinded clinical and histological evaluations were conducted to seek evidence for gluten sensitivity.When fed with a gluten-containing diet, gluten-sensitive macaques showed signs and symptoms of celiac disease including chronic diarrhea, malabsorptive steatorrhea, intestinal lesions and anti-gliadin antibodies. A gluten-free diet reversed these clinical, histological and serological features, while reintroduction of dietary gluten caused rapid relapse.Gluten-sensitive rhesus macaques may be an attractive resource for investigating both the pathogenesis and the treatment of celiac disease.

  4. QSAR models of human data can enrich or replace LLNA testing for human skin sensitization

    OpenAIRE

    Alves, Vinicius M.; Capuzzi, Stephen J.; Muratov, Eugene; Braga, Rodolpho C.; Thornton, Thomas; Fourches, Denis; Strickland, Judy; Kleinstreuer, Nicole; Andrade, Carolina H.; Tropsha, Alexander

    2016-01-01

    Skin sensitization is a major environmental and occupational health hazard. Although many chemicals have been evaluated in humans, there have been no efforts to model these data to date. We have compiled, curated, analyzed, and compared the available human and LLNA data. Using these data, we have developed reliable computational models and applied them for virtual screening of chemical libraries to identify putative skin sensitizers. The overall concordance between murine LLNA and human skin ...

  5. Radiation sensitivity of human lung cancer cell lines

    International Nuclear Information System (INIS)

    Carmichael, J.; Degraff, W.G.; Gamson, J.; Russo, G.; Mitchell, J.B.; Gazdar, A.F.; Minna, J.D.; Levitt, M.L.

    1989-01-01

    X-Ray survival curves were determined using a panel of 17 human lung cancer cell lines, with emphasis on non-small cell lung cancer (NSCLC). In contrast to classic small cell lung cancer (SCLC) cell lines, NSCLC cell lines were generally less sensitive to radiation as evidenced by higher radiation survival curve extrapolation numbers, surviving fraction values following a 2Gy dose (SF2) and the mean inactivation dose values (D) values. The spectrum of in vitro radiation responses observed was similar to that expected in clinical practice, although mesothelioma was unexpectedly sensitive in vitro. Differences in radiosensitivity were best distinguished by comparison of SF2 values. Some NSCLC lines were relatively sensitive, and in view of this demonstrable variability in radiation sensitivity, the SF2 value may be useful for in vitro predictive assay testing of clinical specimens. (author)

  6. Differential sensitivity of Chironomus and human hemoglobin to gamma radiation

    International Nuclear Information System (INIS)

    Gaikwad, Pallavi S.; Panicker, Lata; Mohole, Madhura; Sawant, Sangeeta; Mukhopadhyaya, Rita; Nath, Bimalendu B.

    2016-01-01

    Chironomus ramosus is known to tolerate high doses of gamma radiation exposure. Larvae of this insect possess more than 95% of hemoglobin (Hb) in its circulatory hemolymph. This is a comparative study to see effect of gamma radiation on Hb of Chironomus and humans, two evolutionarily diverse organisms one having extracellular and the other intracellular Hb respectively. Stability and integrity of Chironomus and human Hb to gamma radiation was compared using biophysical techniques like Dynamic Light Scattering (DLS), UV-visible spectroscopy, fluorescence spectrometry and CD spectroscopy after exposure of whole larvae, larval hemolymph, human peripheral blood, purified Chironomus and human Hb. Sequence- and structure-based bioinformatics methods were used to analyze the sequence and structural similarities or differences in the heme pockets of respective Hbs. Resistivity of Chironomus Hb to gamma radiation is remarkably higher than human Hb. Human Hb exhibited loss of heme iron at a relatively low dose of gamma radiation exposure as compared to Chironomus Hb. Unlike human Hb, the heme pocket of Chironomus Hb is rich in aromatic amino acids. Higher hydophobicity around heme pocket confers stability of Chironomus Hb compared to human Hb. Previously reported gamma radiation tolerance of Chironomus can be largely attributed to its evolutionarily ancient form of extracellular Hb as evident from the present study. -- Highlights: •Comparison of radiation tolerant Chironomus Hb and radiation sensitive Human Hb. •Amino acid composition of midge and human heme confer differential hydrophobicity. •Heme pocket of evolutionarily ancient midge Hb provide gamma radiation resistivity.

  7. Differential sensitivity of Chironomus and human hemoglobin to gamma radiation

    Energy Technology Data Exchange (ETDEWEB)

    Gaikwad, Pallavi S. [Stress Biology Research Laboratory, Department of Zoology, Savitribai Phule University, Pune, 411007 (India); Molecular Biology Division, Bhabha Atomic Research Centre, Trombay, Mumbai, 400085 (India); Panicker, Lata [Solid State Physics Division, Bhabha Atomic Research Centre, Trombay, Mumbai, 400085 (India); Mohole, Madhura; Sawant, Sangeeta [Bioinformatics Center, Savitribai Phule Pune University, Pune, 411007 (India); Mukhopadhyaya, Rita [Molecular Biology Division, Bhabha Atomic Research Centre, Trombay, Mumbai, 400085 (India); Nath, Bimalendu B., E-mail: bbnath@gmail.com [Stress Biology Research Laboratory, Department of Zoology, Savitribai Phule University, Pune, 411007 (India)

    2016-08-05

    Chironomus ramosus is known to tolerate high doses of gamma radiation exposure. Larvae of this insect possess more than 95% of hemoglobin (Hb) in its circulatory hemolymph. This is a comparative study to see effect of gamma radiation on Hb of Chironomus and humans, two evolutionarily diverse organisms one having extracellular and the other intracellular Hb respectively. Stability and integrity of Chironomus and human Hb to gamma radiation was compared using biophysical techniques like Dynamic Light Scattering (DLS), UV-visible spectroscopy, fluorescence spectrometry and CD spectroscopy after exposure of whole larvae, larval hemolymph, human peripheral blood, purified Chironomus and human Hb. Sequence- and structure-based bioinformatics methods were used to analyze the sequence and structural similarities or differences in the heme pockets of respective Hbs. Resistivity of Chironomus Hb to gamma radiation is remarkably higher than human Hb. Human Hb exhibited loss of heme iron at a relatively low dose of gamma radiation exposure as compared to Chironomus Hb. Unlike human Hb, the heme pocket of Chironomus Hb is rich in aromatic amino acids. Higher hydophobicity around heme pocket confers stability of Chironomus Hb compared to human Hb. Previously reported gamma radiation tolerance of Chironomus can be largely attributed to its evolutionarily ancient form of extracellular Hb as evident from the present study. -- Highlights: •Comparison of radiation tolerant Chironomus Hb and radiation sensitive Human Hb. •Amino acid composition of midge and human heme confer differential hydrophobicity. •Heme pocket of evolutionarily ancient midge Hb provide gamma radiation resistivity.

  8. Identification of failure sequences sensitive to human error

    International Nuclear Information System (INIS)

    1987-06-01

    This report prepared by the participants of the technical committee meeting on ''Identification of Failure Sequences Sensitive to Human Error'' addresses the subjects discussed during the meeting and the conclusions reached by the committee. Chapter 1 reviews the INSAG recommendations and the main elements of the IAEA Programme in the area of human element. In Chapter 2 the role of human actions in nuclear power plants safety from insights of operational experience is reviewed. Chapter 3 is concerned with the relationship between probabilistic safety assessment and human performance associated with severe accident sequences. Chapter 4 addresses the role of simulators in view of training for accident conditions. Chapter 5 presents the conclusions and future trends. The seven papers presented by members of this technical committee are also included in this technical document. A separate abstract was prepared for each of these papers

  9. Sensitive detection of viral transcripts in human tumor transcriptomes.

    Directory of Open Access Journals (Sweden)

    Sven-Eric Schelhorn

    Full Text Available In excess of 12% of human cancer incidents have a viral cofactor. Epidemiological studies of idiopathic human cancers indicate that additional tumor viruses remain to be discovered. Recent advances in sequencing technology have enabled systematic screenings of human tumor transcriptomes for viral transcripts. However, technical problems such as low abundances of viral transcripts in large volumes of sequencing data, viral sequence divergence, and homology between viral and human factors significantly confound identification of tumor viruses. We have developed a novel computational approach for detecting viral transcripts in human cancers that takes the aforementioned confounding factors into account and is applicable to a wide variety of viruses and tumors. We apply the approach to conducting the first systematic search for viruses in neuroblastoma, the most common cancer in infancy. The diverse clinical progression of this disease as well as related epidemiological and virological findings are highly suggestive of a pathogenic cofactor. However, a viral etiology of neuroblastoma is currently contested. We mapped 14 transcriptomes of neuroblastoma as well as positive and negative controls to the human and all known viral genomes in order to detect both known and unknown viruses. Analysis of controls, comparisons with related methods, and statistical estimates demonstrate the high sensitivity of our approach. Detailed investigation of putative viral transcripts within neuroblastoma samples did not provide evidence for the existence of any known human viruses. Likewise, de-novo assembly and analysis of chimeric transcripts did not result in expression signatures associated with novel human pathogens. While confounding factors such as sample dilution or viral clearance in progressed tumors may mask viral cofactors in the data, in principle, this is rendered less likely by the high sensitivity of our approach and the number of biological replicates

  10. A new sensitive method for detecting human endogenous (leukocyte) pyrogen.

    Science.gov (United States)

    Bodel, P; Miller, H

    1978-03-01

    Endogenous, or leukocyte pyrogen (EP), the mediator of fever, is currently detected by injection of pyrogen-containing supernatants into rabbits. This assay has been of little value in the study of human fever because it required injection of relatively large amounts of pyrogen. We now report that injection of medium containing human EP produces fever in mice. Supernatant from 1 c 10(5) granulocytes, stimulated by phagocytosis of staphylococci and incubated overnight, or 1 x 10(4) monocytes similarly treated, produce clear pyrogenic responses. This method for detecting EP is about 100-fold more sensitive than the rabbit assay, and it appears to be specific for EP. Preliminary studies of EP released by small samples of needle liver biopsies from febrile and afebrile patients suggests that this sensitive assay may be useful for investigations into the mechanisms of clinical fever.

  11. Zebra finches are sensitive to prosodic features of human speech.

    Science.gov (United States)

    Spierings, Michelle J; ten Cate, Carel

    2014-07-22

    Variation in pitch, amplitude and rhythm adds crucial paralinguistic information to human speech. Such prosodic cues can reveal information about the meaning or emphasis of a sentence or the emotional state of the speaker. To examine the hypothesis that sensitivity to prosodic cues is language independent and not human specific, we tested prosody perception in a controlled experiment with zebra finches. Using a go/no-go procedure, subjects were trained to discriminate between speech syllables arranged in XYXY patterns with prosodic stress on the first syllable and XXYY patterns with prosodic stress on the final syllable. To systematically determine the salience of the various prosodic cues (pitch, duration and amplitude) to the zebra finches, they were subjected to five tests with different combinations of these cues. The zebra finches generalized the prosodic pattern to sequences that consisted of new syllables and used prosodic features over structural ones to discriminate between stimuli. This strong sensitivity to the prosodic pattern was maintained when only a single prosodic cue was available. The change in pitch was treated as more salient than changes in the other prosodic features. These results show that zebra finches are sensitive to the same prosodic cues known to affect human speech perception. © 2014 The Author(s) Published by the Royal Society. All rights reserved.

  12. QSAR models of human data can enrich or replace LLNA testing for human skin sensitization

    Science.gov (United States)

    Alves, Vinicius M.; Capuzzi, Stephen J.; Muratov, Eugene; Braga, Rodolpho C.; Thornton, Thomas; Fourches, Denis; Strickland, Judy; Kleinstreuer, Nicole; Andrade, Carolina H.; Tropsha, Alexander

    2016-01-01

    Skin sensitization is a major environmental and occupational health hazard. Although many chemicals have been evaluated in humans, there have been no efforts to model these data to date. We have compiled, curated, analyzed, and compared the available human and LLNA data. Using these data, we have developed reliable computational models and applied them for virtual screening of chemical libraries to identify putative skin sensitizers. The overall concordance between murine LLNA and human skin sensitization responses for a set of 135 unique chemicals was low (R = 28-43%), although several chemical classes had high concordance. We have succeeded to develop predictive QSAR models of all available human data with the external correct classification rate of 71%. A consensus model integrating concordant QSAR predictions and LLNA results afforded a higher CCR of 82% but at the expense of the reduced external dataset coverage (52%). We used the developed QSAR models for virtual screening of CosIng database and identified 1061 putative skin sensitizers; for seventeen of these compounds, we found published evidence of their skin sensitization effects. Models reported herein provide more accurate alternative to LLNA testing for human skin sensitization assessment across diverse chemical data. In addition, they can also be used to guide the structural optimization of toxic compounds to reduce their skin sensitization potential. PMID:28630595

  13. Salivary Proteome Patterns Affecting Human Salt Taste Sensitivity.

    Science.gov (United States)

    Stolle, Theresa; Grondinger, Freya; Dunkel, Andreas; Meng, Chen; Médard, Guillaume; Kuster, Bernhard; Hofmann, Thomas

    2017-10-25

    To investigate the role of perireceptor events in inter-individual variability in salt taste sensitivity, 31 volunteers were monitored in their detection functions for sodium chloride (NaCl) and classified into sensitive (0.6-1.7 mmol/L), medium-sensitive (1.8-6.9 mmol/L), and nonsensitive (7.0-11.2 mmol/L) subjects. Chemosensory intervention of NaCl-sensitive (S + ) and nonsensitive (S - ) panellists with potassium chloride, ammonium chloride, and sodium gluconate showed the salt taste sensitivity to be specific for NaCl. As no significant differences were found between S + and S - subjects in salivary sodium and protein content, salivary proteome differences and their stimulus-induced dynamic changes were analyzed by tryptic digestion, iTRAQ labeling, and liquid chromatography-tandem mass spectrometry analysis. Differences in the salivary proteome between S + and S - subjects were found primarily in resting saliva and were largely independent of the dynamic alterations observed upon salt stimulation. Gene ontology enrichment analysis of key proteins, i.e., immunoglobulin heavy constant y1, myeloblastin, cathepsin G, and kallikrein, revealed significantly increased serine-type endopeptidase activity for the S + group, while the S - group exhibited augmented cysteine-type endopeptidase inhibitor activity by increased abundances in lipocalin-1 and cystatin-D, -S, and -SN, respectively. As proteases have been suggested to facilitate transepithelial sodium transport by cleaving the y-subunit of the epithelial sodium channel (ENaC) and protease inhibitors have been shown to reduce ENaC-mediated sodium transport, the differentially modulated proteolytic activity patterns observed in vivo for S + and S - subjects show evidence of them playing a crucial role in affecting human NaCl sensitivity.

  14. Sensitive periods in human development: evidence from musical training.

    Science.gov (United States)

    Penhune, Virginia B

    2011-10-01

    One of the primary goals of cognitive neuroscience is to understand the interaction between genes, development and specific experience. A particularly fascinating example of this interaction is a sensitive period - a time during development when experience has a differential effect on behavior and the brain. Behavioral and brain imaging studies in musicians have provided suggestive evidence for a possible sensitive period for musical training; showing that musicians who began training early show better task performance and greater changes in auditory and motor regions of the brain. However, these studies have not controlled for likely differences between early- (ET) and late-trained (LT) musicians in the number of years of musical experience. This review presents behavioral work from our laboratory comparing the performance of ET (before age seven) and LT musicians who were matched for years of experience on the ability to tap in synchrony with auditory and visual rhythms. The results demonstrate the existence of a possible sensitive period for musical training that has its greatest impact on measures of sensorimotor integration. Work on motor learning in children and how this might relate to the observed sensitive period effect is also reviewed. These studies are described in the context of what is currently known about sensitive periods in animals and humans; drawing on evidence from anatomy and physiology, studies of deafness, as well as structural and functional neuroimaging studies in trained musicians. The possible mechanisms underlying sensitive periods for musical training are discussed based on current theories describing the influence of both low-level features of sensory experience and higher-level cognitive processing. Copyright © 2011 Elsevier Srl. All rights reserved.

  15. The sensitivity of human mesenchymal stem cells to ionizing radiation

    International Nuclear Information System (INIS)

    Chen, M.-F.; Lin, C.-T.; Chen, W.-C.; Yang, C.-T.; Chen, C.-C.; Liao, S.-K.; Liu, J.M.; Lu, C.-H.; Lee, K.-D.

    2006-01-01

    Purpose: Recent studies have shown that mesenchymal stem cells (MSCs) obtained from bone marrow transplantation patients originate from the host. This clinical observation suggests that MSCs in their niches could be resistant to irradiation. However, the biologic responses of bone marrow MSCs to irradiation have rarely been described in the literature. Methods and Materials: In this study, human bone marrow-derived, clonally expanded MSCs were used to investigate their sensitivity to irradiation in vitro, and the cellular mechanisms that may facilitate resistance to irradiation. The human lung cancer cell line A549 and the breast cancer cell line HCC1937 were used as controls for radiosensitivity; the former line has been shown to be radioresistant and the latter radiosensitive. We then examined their in vitro biologic changes and sensitivities to radiation therapy. Results: Our results suggest that MSCs are characterized as resistant to irradiation. Several cellular mechanisms were demonstrated that may facilitate resistance to irradiation: ATM protein phosphorylation, activation of cell-cycle checkpoints, double-strand break repair by homologous recombination and nonhomologous end joining (NHEJ), and the antioxidant capacity for scavenging reactive oxygen species. Conclusions: As demonstrated, MSCs possess a better antioxidant reactive oxygen species-scavenging capacity and active double-strand break repair to facilitate their radioresistance. These findings provide a better understanding of radiation-induced biologic responses in MSCs and may lead to the development of better strategies for stem cell treatment and cancer therapy

  16. Discrimination of skin sensitizers from non-sensitizers by interleukin-1α and interleukin-6 production on cultured human keratinocytes.

    Science.gov (United States)

    Jung, Daun; Che, Jeong-Hwan; Lim, Kyung-Min; Chun, Young-Jin; Heo, Yong; Seok, Seung Hyeok

    2016-09-01

    In vitro testing methods for classifying sensitizers could be valuable alternatives to in vivo sensitization testing using animal models, such as the murine local lymph node assay (LLNA) and the guinea pig maximization test (GMT), but there remains a need for in vitro methods that are more accurate and simpler to distinguish skin sensitizers from non-sensitizers. Thus, the aim of our study was to establish an in vitro assay as a screening tool for detecting skin sensitizers using the human keratinocyte cell line, HaCaT. HaCaT cells were exposed to 16 relevant skin sensitizers and 6 skin non-sensitizers. The highest dose used was the dose causing 75% cell viability (CV75) that we determined by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. The levels of extracellular production of interleukin-1α (IL-1α) and IL-6 were measured. The sensitivity of IL-1α was 63%, specificity was 83% and accuracy was 68%. In the case of IL-6, sensitivity: 69%, specificity: 83% and accuracy: 73%. Thus, this study suggests that measuring extracellular production of pro-inflammatory cytokines IL-1α and IL-6 by human HaCaT cells may potentially classify skin sensitizers from non-sensitizers. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  17. Resveratrol sensitization of DU145 prostate cancer cells to ionizing radiation is associated to ceramide increase.

    Science.gov (United States)

    Scarlatti, Francesca; Sala, Giusy; Ricci, Clara; Maioli, Claudio; Milani, Franco; Minella, Marco; Botturi, Marco; Ghidoni, Riccardo

    2007-08-08

    Radiotherapy is an established therapeutic modality for prostate cancer. Since it is well known that radiotherapy is limited due to its severe toxicity towards normal cells at high dose and minimal effect at low dose, the search for biological compounds that increase the sensitivity of tumors cells to radiation may improve the efficacy of therapy. Resveratrol, a natural antioxidant, was shown to inhibit carcinogenesis in animal models, and to block the process of tumor initiation and progression. The purpose of this study was to examine whether or not resveratrol can sensitize DU145, an androgen-independent human prostate cancer cell line, to ionizing radiation. We report here that DU145 cells are resistant to ionizing radiation-induced cell death, but pretreatment with resveratrol significantly enhances cell death. Resveratrol acts synergistically with ionizing radiation to inhibit cell survival in vitro. Resveratrol also potentiates ionizing radiation-induced ceramide accumulation, by promoting its de novo biosynthesis. This confirms ceramide as an effective mediator of the anticancer potential induced by resveratrol.

  18. Comparison of risk sensitivity to human errors in the Oconee and LaSalle PRAs

    International Nuclear Information System (INIS)

    Wong, S.; Higgins, J.

    1991-01-01

    This paper describes the comparative analyses of plant risk sensitivity to human errors in the Oconee and La Salle Probabilistic Risk Assessment (PRAs). These analyses were performed to determine the reasons for the observed differences in the sensitivity of core melt frequency (CMF) to changes in human error probabilities (HEPs). Plant-specific design features, PRA methods, and the level of detail and assumptions in the human error modeling were evaluated to assess their influence risk estimates and sensitivities

  19. Machine Learning Approaches for Predicting Human Skin Sensitization Hazard

    Science.gov (United States)

    One of ICCVAM’s top priorities is the development and evaluation of non-animal approaches to identify potential skin sensitizers. The complexity of biological events necessary for a substance to elicit a skin sensitization reaction suggests that no single in chemico, in vit...

  20. Multivariate Models for Prediction of Skin Sensitization Hazard in Humans

    Science.gov (United States)

    One of ICCVAM’s highest priorities is the development and evaluation of non-animal approaches to identify potential skin sensitizers. The complexity of biological events necessary for a substance to elicit a skin sensitization reaction suggests that no single alternative me...

  1. Differential sensitivity to human communication in dogs, wolves, and human infants.

    Science.gov (United States)

    Topál, József; Gergely, György; Erdohegyi, Agnes; Csibra, Gergely; Miklósi, Adám

    2009-09-04

    Ten-month-old infants persistently search for a hidden object at its initial hiding place even after observing it being hidden at another location. Recent evidence suggests that communicative cues from the experimenter contribute to the emergence of this perseverative search error. We replicated these results with dogs (Canis familiaris), who also commit more search errors in ostensive-communicative (in 75% of the total trials) than in noncommunicative (39%) or nonsocial (17%) hiding contexts. However, comparative investigations suggest that communicative signals serve different functions for dogs and infants, whereas human-reared wolves (Canis lupus) do not show doglike context-dependent differences of search errors. We propose that shared sensitivity to human communicative signals stems from convergent social evolution of the Homo and the Canis genera.

  2. Multivariate Models for Prediction of Human Skin Sensitization Hazard.

    Science.gov (United States)

    One of the lnteragency Coordinating Committee on the Validation of Alternative Method's (ICCVAM) top priorities is the development and evaluation of non-animal approaches to identify potential skin sensitizers. The complexity of biological events necessary to produce skin sensiti...

  3. Quantitative relationship between the local lymph node assay and human skin sensitization assays.

    Science.gov (United States)

    Schneider, K; Akkan, Z

    2004-06-01

    The local lymph node assay (LLNA) is a new test method which allows for the quantitative assessment of sensitizing potency in the mouse. Here, we investigate the quantitative correlation between results from the LLNA and two human sensitization tests--specifically, human repeat insult patch tests (HRIPTs) and human maximization tests (HMTs). Data for 57 substances were evaluated, of which 46 showed skin sensitizing properties in human tests, whereas 11 yielded negative results in humans. For better comparability data from mouse and human tests were transformed to applied doses per skin area, which ranged over four orders of magnitude for the substances considered. Regression analysis for the 46 human sensitizing substances revealed a significant positive correlation between the LLNA and human tests. The correlation was better between LLNA and HRIPT data (n=23; r=0.77) than between LLNA and HMT data (n=38; r=0.65). The observed scattering of data points is related to various uncertainties, in part associated with insufficiencies of data from older HMT studies. Predominantly negative results in the LLNA for another 11 substances which showed no skin sensitizing activity in human maximization tests further corroborate the correspondence between LLNA and human tests. Based on this analysis, the LLNA can be considered a reliable basis for relative potency assessments for skin sensitizers. Proposals are made for the regulatory exploitation of the LLNA: four potency groups can be established, and assignment of substances to these groups according to the outcome of the LLNA can be used to characterize skin sensitizing potency in substance-specific assessments. Moreover, based on these potency groups, a more adequate consideration of sensitizing substances in preparations becomes possible. It is proposed to replace the current single concentration limit for skin sensitizers in preparations, which leads to an all or nothing classification of a preparation as sensitizing to

  4. Caffeine markedly sensitizes human mesothelioma cell lines to pemetrexed

    Science.gov (United States)

    Min, Sang Hee; Goldman, I. David; Zhao, Rongbao

    2013-01-01

    Pemetrexed is a new generation antifolate approved for the treatment of mesothelioma and non-small cell lung cancer. Caffeine is known to augment radiation or chemotherapeutic drug-induced cell killing. The current study addresses the impact of caffeine on the activity of pemetrexed in mesothelioma cell lines. Caffeine enhanced pemetrexed activity in all four mesothelioma cell lines tested (H2052, H2373, H28 and MSTO-211H). Caffeine sensitized H2052 cells in a dose- and schedule-dependent manner, and was associated with a markedly decreased clonogenic survival. Caffeine sensitization occurred only in cells subjected to pulse, but not continuous, exposure to pemetrexed. Similar pemetrexed sensitization was also observed with the clinically better tolerated caffeine analog, theobromine. Pemetrexed sensitization by caffeine was associated with an increase in pemetrexed-induced phosphorylation of ataxia-telangiectasia-mutated (ATM) and Chk1. These data indicate that caffeine and its analog, theobromine, may be a useful approach to enhance pemetrexed-based chemotherapy. PMID:17594092

  5. Is There a Sensitive Period in Human Incest Avoidance?

    Directory of Open Access Journals (Sweden)

    Liqun Luo

    2011-04-01

    Full Text Available Many studies support the proposition that early cosocialization with opposite-sex children has the effect of inhibiting later mutual sexual attraction, but the existence of a period in the life cycle in which individuals are sensitive to the effect of early cosocialization has been a matter of controversy. Drawing on earlier traditional psychological research, and on more recent work guided by parental investment theory, we hypothesized that only for maternal perinatal association (MPA-absent males a less-than-around-three-years age difference with the sister can predict stronger aversion to sibling incest. The results corroborated the hypothesis. The results can be interpreted as support for the existence of a sensitive period as well as for the potent role of MPA. Cross-cultural comparative studies were called on to further test the hypothesis.

  6. Human sensitization to Prosopis juliflora antigen in Saudi Arabia.

    Science.gov (United States)

    Al-Frayh, A; Hasnain, S M; Gad-El-Rab, M O; Al-Turki, T; Al-Mobeireek, K; Al-Sedairy, S T

    1999-01-01

    Allergenicity to Prosopis juliflora pollen antigen has been reported from only a few countries, including the US, South Africa, India and Kuwait. In some parts of Saudi Arabia, species of Prosopis have been introduced by the millions as roadside ornamentation. There appear to be four flowering seasons during which pollen grains float in all directions. However, the role of Prosopis pollen as the sensitizing and/or triggering agent of allergic asthma and/or rhinitis in the Kingdom has never been evaluated. A total of 473 allergic patients suffering from bronchial asthma in four different geographical regions (Abha, Qassim, Hofuf and Gizan), and attending allergy clinics and chest disease centers of university and Ministry of Health hospitals in the region were tested for immediate hypersensitivity reaction to Prosopis juliflora allergens. Airborne pollen grains at one center were also studied for one full year, using volumetric sampling techniques. A total of 76.1% patients in Qassim, 37.5% in Gizan, 29% in Abha and 11% in Hofuf reacted positively to Prosopis antigen. Multiple sensitivities to other pollen antigens were detected in all patients. The level of airborne Prosopis pollen detected in Gizan exceeded 90 grains m -3 of air. In view of the documented evidence of Prosopis-involved allergenicity, the role of Prosopis pollen as a sensitizing factor in Saudi Arabia has been confirmed. However, the cause of elicitation of symptoms in many multiple sensitive patients, together with the question of cross-reactivities, needs thorough and detailed investigation. In vitro confirmation of all positive results is also required to incriminate Prosopis as one of the major allergens in parts of Saudi Arabia.

  7. Optimizing human activity patterns using global sensitivity analysis.

    Science.gov (United States)

    Fairchild, Geoffrey; Hickmann, Kyle S; Mniszewski, Susan M; Del Valle, Sara Y; Hyman, James M

    2014-12-01

    Implementing realistic activity patterns for a population is crucial for modeling, for example, disease spread, supply and demand, and disaster response. Using the dynamic activity simulation engine, DASim, we generate schedules for a population that capture regular (e.g., working, eating, and sleeping) and irregular activities (e.g., shopping or going to the doctor). We use the sample entropy (SampEn) statistic to quantify a schedule's regularity for a population. We show how to tune an activity's regularity by adjusting SampEn, thereby making it possible to realistically design activities when creating a schedule. The tuning process sets up a computationally intractable high-dimensional optimization problem. To reduce the computational demand, we use Bayesian Gaussian process regression to compute global sensitivity indices and identify the parameters that have the greatest effect on the variance of SampEn. We use the harmony search (HS) global optimization algorithm to locate global optima. Our results show that HS combined with global sensitivity analysis can efficiently tune the SampEn statistic with few search iterations. We demonstrate how global sensitivity analysis can guide statistical emulation and global optimization algorithms to efficiently tune activities and generate realistic activity patterns. Though our tuning methods are applied to dynamic activity schedule generation, they are general and represent a significant step in the direction of automated tuning and optimization of high-dimensional computer simulations.

  8. Migration of human antigen-presenting cells in a human skin graft onto nude mice model after contact sensitization

    NARCIS (Netherlands)

    Hoefakker, S.; Balk, H.P.; Boersma, W.J.A.; Joost, T. van; Notten, W.R.F.; Claassen, E.

    1995-01-01

    Fluorescent contact chemical allergens provoke sensitization after application on both syngeneic and allogeneic skin grafts in mice. We attempted to determine whether the functional activity in a contact sensitization response of human skin graft was affected at the level of antigen uptake and

  9. What did domestication do to dogs? A new account of dogs' sensitivity to human actions.

    Science.gov (United States)

    Udell, Monique A R; Dorey, Nicole R; Wynne, Clive D L

    2010-05-01

    Over the last two decades increasing evidence for an acute sensitivity to human gestures and attentional states in domestic dogs has led to a burgeoning of research into the social cognition of this highly familiar yet previously under-studied animal. Dogs (Canis lupus familiaris) have been shown to be more successful than their closest relative (and wild progenitor) the wolf, and than man's closest relative, the chimpanzee, on tests of sensitivity to human social cues, such as following points to a container holding hidden food. The "Domestication Hypothesis" asserts that during domestication dogs evolved an inherent sensitivity to human gestures that their non-domesticated counterparts do not share. According to this view, sensitivity to human cues is present in dogs at an early age and shows little evidence of acquisition during ontogeny. A closer look at the findings of research on canine domestication, socialization, and conditioning, brings the assumptions of this hypothesis into question. We propose the Two Stage Hypothesis, according to which the sensitivity of an individual animal to human actions depends on acceptance of humans as social companions, and conditioning to follow human limbs. This offers a more parsimonious explanation for the domestic dog's sensitivity to human gestures, without requiring the use of additional mechanisms. We outline how tests of this new hypothesis open directions for future study that offer promise of a deeper understanding of mankind's oldest companion.

  10. X-ray sensitivity of human tumor cells in vitro

    International Nuclear Information System (INIS)

    Weichselbaum, R.R.; Nove, J.; Little, J.B.

    1980-01-01

    Clonally-derived cells from ten human malignant tumors considered radiocurable (breast, neuroblastoma, medulloblastoma) or non-radiocurable (osteosarcoma, hypernephroma, glioblastoma, melanoma) were studied in cell culture and their in vitro x-ray survival curve parameters determined (anti n, D 0 ). There were no significant differences among the tumor cell lines suggesting that survival parameters in vitro do not explain differences in clinical radiocurability. Preliminary investigation with density inhibited human tumor cells indicate that such an approach may yield information regarding inherent cellular differences in radiocurability

  11. Oxygen-Sensitive K+ Channels Modulate Human Chorionic Gonadotropin Secretion from Human Placental Trophoblast

    Science.gov (United States)

    Díaz, Paula; Sibley, Colin P.; Greenwood, Susan L.

    2016-01-01

    Human chorionic gonadotropin (hCG) is a key autocrine/paracrine regulator of placental syncytiotrophoblast, the transport epithelium of the human placenta. Syncytiotrophoblast hCG secretion is modulated by the partial pressure of oxygen (pO2), reactive oxygen species (ROS) and potassium (K+) channels. Here we test the hypothesis that K+ channels mediate the effects of pO2 and ROS on hCG secretion. Placental villous explants from normal term pregnancies were cultured for 6 days at 6% (normoxia), 21% (hyperoxia) or 1% (hypoxia) pO2. On days 3–5, explants were treated with 5mM 4-aminopyridine (4-AP) or tetraethylammonium (TEA), blockers of pO2-sensitive voltage-gated K+ (KV) channels, or ROS (10–1000μM H2O2). hCG secretion and lactate dehydrogenase (LDH) release, a marker of necrosis, were determined daily. At day 6, hCG and LDH were measured in tissue lysate and 86Rb (K+) efflux assessed to estimate syncytiotrophoblast K+ permeability. hCG secretion and 86Rb efflux were significantly greater in explants maintained in 21% pO2 than normoxia. 4-AP/TEA inhibited hCG secretion to a greater extent at 21% than 6% and 1% pO2, and reduced 86Rb efflux at 21% but not 6% pO2. LDH release and tissue LDH/hCG were similar in 6%, 21% and 1% pO2 and unaffected by 4-AP/TEA. H2O2 stimulated 86Rb efflux and hCG secretion at normoxia but decreased 86Rb efflux, without affecting hCG secretion, at 21% pO2. 4-AP/TEA-sensitive K+ channels participate in pO2-sensitive hCG secretion from syncytiotrophoblast. ROS effects on both hCG secretion and 86Rb efflux are pO2-dependent but causal links between the two remain to be established. PMID:26863525

  12. Human error in strabismus surgery: Quantification with a sensitivity analysis

    NARCIS (Netherlands)

    S. Schutte (Sander); J.R. Polling (Jan Roelof); F.C.T. van der Helm (Frans); H.J. Simonsz (Huib)

    2009-01-01

    textabstractBackground: Reoperations are frequently necessary in strabismus surgery. The goal of this study was to analyze human-error related factors that introduce variability in the results of strabismus surgery in a systematic fashion. Methods: We identified the primary factors that influence

  13. Human error in strabismus surgery : Quantification with a sensitivity analysis

    NARCIS (Netherlands)

    Schutte, S.; Polling, J.R.; Van der Helm, F.C.T.; Simonsz, H.J.

    2008-01-01

    Background- Reoperations are frequently necessary in strabismus surgery. The goal of this study was to analyze human-error related factors that introduce variability in the results of strabismus surgery in a systematic fashion. Methods- We identified the primary factors that influence the outcome of

  14. Variable high pressure processing sensitivities for GII human noroviruses

    Science.gov (United States)

    Human norovirus (HuNoV) is the leading cause of foodborne diseases worldwide. High pressure processing (HPP) is one of the most promising non-thermal technologies for decontamination of viral pathogens in foods. However, the survival of HuNoVs by HPP is poorly understood because these viruses cann...

  15. Evidence supporting oral sensitivity to complex carbohydrates independent of sweet taste sensitivity in humans.

    Directory of Open Access Journals (Sweden)

    Julia Y Q Low

    Full Text Available Compared to simple sugars, complex carbohydrates have been assumed invisible to taste. However, two recent studies proposed that there may be a perceivable taste quality elicited by complex carbohydrates independent of sweet taste. There is precedent with behavioural studies demonstrating that rats are very attracted to complex carbohydrates, and that complex carbohydrates are preferred to simple sugars at low concentrations. This suggests that rats may have independent taste sensors for simple sugars and complex carbohydrates. The aim of this paper is to investigate oral sensitivities of two different classes of complex carbohydrates (a soluble digestible and a soluble non-digestible complex carbohydrate, and to compare these to other caloric and non-nutritive sweeteners in addition to the prototypical tastes using two commonly used psychophysical measures. There were strong correlations between the detection thresholds and mean intensity ratings for complex carbohydrates (maltodextrin, oligofructose (r = 0.94, P 0.05. However, moderate correlations were observed between perceived intensities of complex carbohydrates and sweeteners (r = 0.48-0.61, P < 0.05. These data provide evidence that complex carbohydrates can be sensed in the oral cavity over a range of concentrations independent of sweet taste sensitivity at low concentrations, but with partial overlap with sweet taste intensity at higher concentrations.

  16. Effect of ethanol of the radiation sensitivity of human hemoglobin

    International Nuclear Information System (INIS)

    Szweda-Lewandowska, Z.; Puchala, M.

    1981-01-01

    Radiation sensitivity of oxy-, deoxy-, and methemoglobin (HbOs, Hbbj, and MetHb) in water solutions containing 0.2 M ethanol and in ethanol-free solutions was compared. Radiation sensitivity was estimated on the basis of changes in absorbance at the Soret band (a = 430 nm for deoxyhemoglobin), changes in the absorbance ration Avqv/Avwt determined after conversion of irradiated preparations to methemoglobin, and changes in the value of parameters describing the reaction of hemoglobin oxygenation. The protection coefficient p of hemoglobin by ethanol (ratio of a change in the absence of ethanol to that in its presence) calculated from changes in absorbance at the Soret band equaled about 1.5 at a 4-Mrad dose in all bases except MetHb irradiated in air for which p was much higher (about 3.2). The protection coefficient p' calculated from Dtx values for changes in Avchemically bondv/Avwt equaled 2.2 for HbOs, and 2.8 for MetHb for preparations irradiated in air; p' = 1.7 for Hbbj and 1.8 for MetHb for preparations irradiated under argon. On the basis of these results, the role of /sup ./OH radicals and oxygen in the radiation damage of hemoglobin is discussed

  17. Human sensitization to Prosopis Juliflora antigen in Saudi Arabia

    International Nuclear Information System (INIS)

    Al-Frayh, A.; Gad-el-Rab, Mohammed O.; Al-Mobeireek, K.; Al-Turki, T.; Hasnain, Syed M.; Al-Sedairy, Sultan T.

    1999-01-01

    Allergenicity Prosopis juliflora pollen antigen has been reported fromonly a few countries, including the US, South Africa, India and Kuwait. Insome parts of Saudi Arabia, species of Prosopis have been introduced by themillions as roadside ornamentation. There appear to be four flowering seasonsduring which pollen grains float in all directions. However, the role ofProsopis pollen as the sensitizing and/or rhinitis in the Kingdom has neverbeen evaluated. A total of 473 allergic patients suffering from the bronchialasthma in four different geographical regions (Abha, Qassim, Hofuf, Gizan),and attending allergy clinics and chest disease centers of university andMinistry of Health hospitals in the region were tested for immediatehypersensitivity reaction to Prosopis Juliflora allergens. Airborne pollengrains at one center were also studied for one full year, using volumetricsampling techniques. A total of 76.1% patients in Qassim, 37.5% in Gizan, 29%in Abha and 11% in Hofuf reacted positively to Prosopis antigen. Multiplesensitivities to other pollen antigens were detected in all patients. Thelevel of airborne Prosopis pollen detected in Gizan exceeded 90 grains m ofair. In view of documented evidence of Prosopis pollen as a sensitizingfactor in Saudi Arabia has been confirmed. However the cause of elicitationof symptoms in many multiple sensitive patients, together with the questionof cross-reactivities, needs thorough and detailed investigation. In vitroconfirmation of all positive results is also required to incriminate Prosopisas one of the major allergens in parts of Saudi Arabia. (author)

  18. Human gut microbes impact host serum metabolome and insulin sensitivity

    DEFF Research Database (Denmark)

    Pedersen, Helle Krogh; Gudmundsdottir, Valborg; Nielsen, Henrik Bjørn

    2016-01-01

    Insulin resistance is a forerunner state of ischaemic cardiovascular disease and type 2 diabetes. Here we show how the human gut microbiome impacts the serum metabolome and associates with insulin resistance in 277 non-diabetic Danish individuals. The serum metabolome of insulin-resistant individ......Insulin resistance is a forerunner state of ischaemic cardiovascular disease and type 2 diabetes. Here we show how the human gut microbiome impacts the serum metabolome and associates with insulin resistance in 277 non-diabetic Danish individuals. The serum metabolome of insulin......-resistant individuals is characterized by increased levels of branched-chain amino acids (BCAAs), which correlate with a gut microbiome that has an enriched biosynthetic potential for BCAAs and is deprived of genes encoding bacterial inward transporters for these amino acids. Prevotella copri and Bacteroides vulgatus...

  19. Targeting Rad50 sensitizes human nasopharyngeal carcinoma cells to radiotherapy

    International Nuclear Information System (INIS)

    Chang, Lihong; Huang, Jiancong; Wang, Kai; Li, Jingjia; Yan, Ruicheng; Zhu, Ling; Ye, Jin; Wu, Xifu; Zhuang, Shimin; Li, Daqing; Zhang, Gehua

    2016-01-01

    The Mre11-Rad50-Nbs1 (MRN) complex is well known for its crucial role in initiating DNA double strand breaks (DSBs) repair pathways to resistant irradiation (IR) injury and thus facilitating radioresistance which severely reduces radiocurability of nasopharyngeal cancer (NPC). Targeting native cellular MRN function would sensitize NPC cells to IR. A recombinant adenovirus containing a mutant Rad50 gene (Ad-RAD50) expressing Rad50 zinc hook domain but lacking the ATPase domain and the Mre11 interaction domain was constructed to disrupt native cellular MRN functions. The effects of Ad-RAD50 on the MRN functions were assessed in NPC cells lines using western blot, co-immunoprecipitation and confocal microscopy analyses. The increased radiosensitivity of transient Ad-RAD50 to IR was examined in NPC cells, including MTT assay, colony formation. The molecular mechanisms of radiosensitization were confirmed by neutral comet assay and western bolts. Nude mice subcutaneous injection, tumor growth curve and TUNEL assay were used to evaluate tumor regression and apoptosis in vivo. Rad50 is remarkably upregulated in NPC cells after IR, implying the critical role of Rad50 in MRN functions. The transient expression of this mutant Rad50 decreased the levels of native cellular Rad50, Mre11 and Nbs1, weakened the interactions among these proteins, abrogated the G2/M arrest induced by DSBs and reduced the DNA repair ability in NPC cells. A combination of IR and mutant RAD50 therapy produced significant tumor cytotoxicity in vitro, with a corresponding increase in DNA damage, prevented proliferation and cell viability. Furthermore, Ad-RAD50 sensitized NPC cells to IR by causing dramatic tumor regression and inducing apoptosis in vivo. Our findings define a novel therapeutic approach to NPC radiosensitization via targeted native cellular Rad50 disruption. The online version of this article (doi:10.1186/s12885-016-2190-8) contains supplementary material, which is available to

  20. Sensitivity analysis of human brain structural network construction

    Directory of Open Access Journals (Sweden)

    Kuang Wei

    2017-12-01

    Full Text Available Network neuroscience leverages diffusion-weighted magnetic resonance imaging and tractography to quantify structural connectivity of the human brain. However, scientists and practitioners lack a clear understanding of the effects of varying tractography parameters on the constructed structural networks. With diffusion images from the Human Connectome Project (HCP, we characterize how structural networks are impacted by the spatial resolution of brain atlases, total number of tractography streamlines, and grey matter dilation with various graph metrics. We demonstrate how injudicious combinations of highly refined brain parcellations and low numbers of streamlines may inadvertently lead to disconnected network models with isolated nodes. Furthermore, we provide solutions to significantly reduce the likelihood of generating disconnected networks. In addition, for different tractography parameters, we investigate the distributions of values taken by various graph metrics across the population of HCP subjects. Analyzing the ranks of individual subjects within the graph metric distributions, we find that the ranks of individuals are affected differently by atlas scale changes. Our work serves as a guideline for researchers to optimize the selection of tractography parameters and illustrates how biological characteristics of the brain derived in network neuroscience studies can be affected by the choice of atlas parcellation schemes. Diffusion tractography has been proven to be a promising noninvasive technique to study the network properties of the human brain. However, how various tractography and network construction parameters affect network properties has not been studied using a large cohort of high-quality data. We utilize data provided by the Human Connectome Project to characterize the changes to network properties induced by varying the brain parcellation atlas scales, the number of reconstructed tractography tracks, and the degree of grey

  1. Evaluation of human skin tests for potential dermal irritant and contact sensitizing products: a position paper

    NARCIS (Netherlands)

    Loveren H van; Jong WH de; Garssen J; LPI

    1998-01-01

    Prediction of human cutaneous irritation and sensitization in view of hazard identification has primarily relied on the use of laboratory animals. Such studies in laboratory animals have been very instrumental in the detection of potential contact sensitizing agents. There are however many

  2. Sensitivity-enhanced 13C MR spectroscopy of the human brain at 3 Tesla.

    NARCIS (Netherlands)

    Klomp, D.W.J.; Renema, W.K.J.; Graaf, M. de; Galan, B.E. de; Kentgens, A.P.M.; Heerschap, A.

    2006-01-01

    A new coil design for sensitivity-enhanced 13C MR spectroscopy (MRS) of the human brain is presented. The design includes a quadrature transmit/receive head coil optimized for 13C MR sensitivity. Loss-less blocking circuits inside the coil conductors allow this coil to be used inside a homogeneous

  3. Sensitivity-enhanced C-13 MR spectroscopy of the human brain at 3 Tesla

    NARCIS (Netherlands)

    Klomp, D.W.J.; Renema, W.K.J.; Graaf, M. van der; Galan, B.E. de; Kentgens, A.P.M.; Heerschap, A.

    2006-01-01

    A new coil design for sensitivity-enhanced C-13 MR spectroscopy (MRS) of the human brain is presented. The design includes a quadrature transmit/receive head coil optimized for C-13 MR sensitivity. Loss-less blocking circuits inside the coil conductors allow this coil to be used inside a homogeneous

  4. d -Limonene sensitizes docetaxel-induced cytotoxicity in human prostate cancer cells: Generation of reactive oxygen species and induction of apoptosis

    Directory of Open Access Journals (Sweden)

    Rabi Thangaiyan

    2009-01-01

    Full Text Available Background: Clinical trials have shown that docetaxel combined with other novel agents can improve the survival of androgen-independent prostate cancer patients. d -Limonene, a non-nutrient dietary component, has been found to inhibit various cancer cell growths without toxicity. We sought to characterize whether a non-toxic dose of d -limonene may enhance tumor response to docetaxel in an in vitro model of metastatic prostate cancer. Materials and Methods: Human prostate carcinoma DU-145 and normal prostate epithelial PZ-HPV-7 cells were treated with various concentrations of d -limonene, docetaxel or a combination of both, and cell viability was determined by MTT assay. Intracellular reactive oxygen species (ROS, reduced glutathione (GSH and caspase activity were measured. Apoptosis and apoptosis-related proteins were studied by enzyme-linked immunosorbent assay and Western blotting, respectively. Results: d -Limonene and docetaxel in combination significantly enhanced the cytotoxicity to DU-145 cells than PZ-HPV-7 cells. Exposure of DU-145 cells to a combined d -limonene and docetaxel resulted in higher ROS generation, depletion of GSH, accompanied by increased caspase activity than docetaxel alone. It also triggered a series of effects involving cytochrome c , cleavages of caspase-9, 3 and poly (ADP-ribose polymerase, and a shift in Bad:Bcl-xL ratio in favor of apoptosis. Apoptotic effect was significantly blocked on pretreatment with N -acetylcystein, indicating that antitumor effect is initiated by ROS generation, and caspase cascades contribute to the cell death. Conclusion: Our results show, for the first time, that d -limonene enhanced the antitumor effect of docetaxel against prostate cancer cells without being toxic to normal prostate epithelial cells. The combined beneficial effect could be through the modulation of proteins involved in mitochondrial pathway of apoptosis. d -Limonene could be used as a potent non-toxic agent to

  5. Hunted woolly monkeys (Lagothrix poeppigii show threat-sensitive responses to human presence.

    Directory of Open Access Journals (Sweden)

    Sarah Papworth

    Full Text Available Responding only to individuals of a predator species which display threatening behaviour allows prey species to minimise energy expenditure and other costs of predator avoidance, such as disruption of feeding. The threat sensitivity hypothesis predicts such behaviour in prey species. If hunted animals are unable to distinguish dangerous humans from non-dangerous humans, human hunting is likely to have a greater effect on prey populations as all human encounters should lead to predator avoidance, increasing stress and creating opportunity costs for exploited populations. We test the threat sensitivity hypothesis in wild Poeppigi's woolly monkeys (Lagothrix poeppigii in Yasuní National Park, Ecuador, by presenting human models engaging in one of three behaviours "hunting", "gathering" or "researching". These experiments were conducted at two sites with differing hunting pressures. Visibility, movement and vocalisations were recorded and results from two sites showed that groups changed their behaviours after being exposed to humans, and did so in different ways depending on the behaviour of the human model. Results at the site with higher hunting pressure were consistent with predictions based on the threat sensitivity hypothesis. Although results at the site with lower hunting pressure were not consistent with the results at the site with higher hunting pressure, groups at this site also showed differential responses to different human behaviours. These results provide evidence of threat-sensitive predator avoidance in hunted primates, which may allow them to conserve both time and energy when encountering humans which pose no threat.

  6. Vibration sensitivity of human muscle spindles and Golgi tendon organs.

    Science.gov (United States)

    Fallon, James B; Macefield, Vaughan G

    2007-07-01

    The responses of the various muscle receptors to vibration are more complicated than a naïve categorization into stretch (muscle spindle primary ending), length (muscle spindle secondary endings), and tension (Golgi tendon organs) receptors. To emphasize the similarity of responses to small length changes, we recorded from 58 individual muscle afferents subserving receptors in the ankle or toe dorsiflexors of awake human subjects (32 primary endings, 20 secondary endings, and six Golgi tendon organs). Transverse sinusoidal vibration was applied to the distal tendon of the receptor-bearing muscle, while subjects either remained completely relaxed or maintained a weak isometric contraction of the appropriate muscle. In relaxed muscle, few units responded in a 1:1 manner to vibration, and there was no evidence of a preferred frequency of activation. In active muscle the response profiles of all three receptor types overlapped, with no significant difference in threshold between receptor types. These results emphasize that when intramuscular tension increases during a voluntary contraction, Golgi tendon organs and muscle spindle secondary endings, not just muscle spindle primary endings, can effectively encode small imposed length changes.

  7. Sensitivity of risk parameters to human errors in reactor safety study for a PWR

    International Nuclear Information System (INIS)

    Samanta, P.K.; Hall, R.E.; Swoboda, A.L.

    1981-01-01

    Sensitivities of the risk parameters, emergency safety system unavailabilities, accident sequence probabilities, release category probabilities and core melt probability were investigated for changes in the human error rates within the general methodological framework of the Reactor Safety Study (RSS) for a Pressurized Water Reactor (PWR). Impact of individual human errors were assessed both in terms of their structural importance to core melt and reliability importance on core melt probability. The Human Error Sensitivity Assessment of a PWR (HESAP) computer code was written for the purpose of this study. The code employed point estimate approach and ignored the smoothing technique applied in RSS. It computed the point estimates for the system unavailabilities from the median values of the component failure rates and proceeded in terms of point values to obtain the point estimates for the accident sequence probabilities, core melt probability, and release category probabilities. The sensitivity measure used was the ratio of the top event probability before and after the perturbation of the constituent events. Core melt probability per reactor year showed significant increase with the increase in the human error rates, but did not show similar decrease with the decrease in the human error rates due to the dominance of the hardware failures. When the Minimum Human Error Rate (M.H.E.R.) used is increased to 10 -3 , the base case human error rates start sensitivity to human errors. This effort now allows the evaluation of new error rate data along with proposed changes in the man machine interface

  8. Human skeletal muscle perilipin 2 and 3 expression varies with insulin sensitivity

    DEFF Research Database (Denmark)

    Vigelsø Hansen, Andreas; Prats Gavalda, Clara; Ploug, Thorkil

    2013-01-01

    Background: Impaired insulin sensitivity may partly arise from a dysregulated lipid metabolism in human skeletal muscle. This study investigates the expression levels of perilipin 2, 3, and 5, and four key lipases in human skeletal muscle from the subjects that exhibit a range from normal to very...

  9. A sensitive chain specific radioimmunoassay for human immunoglobulins using monoclonal antibodies

    International Nuclear Information System (INIS)

    Sikora, K.; Alderson, T.St.J.; Ellis, J.

    1983-01-01

    A sensitive radioimmunoassay is described for human immunoglobulins. This solid-phase assay uses commercially available monoclonal antibodies and is specific for different Ig chain types. Levels of less than 20 ng/ml Ig are detectable. The assay is suitable for the analysis of human hybridoma supernatants. (Auth.)

  10. Sensitivity to ionizing radiation and chemotherapeutic agents in gemcitabine-resistant human tumor cell lines

    NARCIS (Netherlands)

    van Bree, Chris; Castro Kreder, Natasja; Loves, Willem J. P.; Franken, Nicolaas A. P.; Peters, Godefridus J.; Haveman, Jaap

    2002-01-01

    Purpose: To determine cross-resistance to anti-tumor treatments in 2',2'difluorodeoxycytidine (dFdC, gemcitabine)-resistant human tumor cells. Methods and Materials: Human lung carcinoma cells SW-1573 (SWp) were made resistant to dFdC (SWg). Sensitivity to cisplatin (cDDP), paclitaxel,

  11. Dependencies of the radiation sensitivity of human tooth enamel in EPR dosimetry

    International Nuclear Information System (INIS)

    Wieser, A.; El-Faramawy, N.; Meckbach, R.

    2001-01-01

    The EPR dose response of tooth enamel was determined for human molars collected in Egypt. The influence of age, gender and residence of the tooth donors as well as tooth position and sample preparation on EPR sensitivity and its variability over the enamel samples was investigated. The EPR sensitivity and its variability were found to depend only on the sample preparation procedure. The variability in EPR sensitivity of enamel from Egyptian teeth was maximally 10% and the mean sensitivity was in good agreement with that of German teeth

  12. Irradiation sensitivity of human and porcine mesenchymal stem cells

    International Nuclear Information System (INIS)

    Singh, S.

    2009-01-01

    Surgical resection, chemotherapy, radiotherapy, and combinations thereof are a plethora of possible treatment modalities of head and neck malignancies. Treatment regimens including radiotherapy however put jaws at risk of subsequent osteoradionecrosis. Besides cancer cells, irradiation impacts on all tissue-inherent cells, including mesenchymal stem cells (MSCs). Since it is the bone and bone marrow MSC, which contributes to bone regeneration through proliferation and osteogenic differentiation of its progeny, the influence of irradiation on MSC viability and the respective differentiation capacity appears to be critical. However to date, only a few reports picked MSCs role out as a pivotal topic. As a first attempt, we irradiated human bone derived MSC in vitro. With increasing doses the cells self-renewal capabilities were greatly reduced. Notably however, the mitotically stalled cells were still capable of differentiating into osteoblasts and preadipocytes. Next, the mandibles of Sus scrofa domestica were irradiated with a total dose of 18 Gy. At different time points post radiatio, MSCs were isolated from bone autopsies. In comparison between irradiated and non- irradiated samples, no significant differences regarding the proliferation and osteogenic differentiation potential of tissue specific MSC became apparent Therefore, pig mandibles were irradiated with doses of 9 and 18 Gy, and MSCs were isolated immediately afterwards. No significant differences between the untreated and bone irradiated with 9 Gy with respect of proliferation and osteogenic differentiation were observed. Cells isolated from 18 Gy irradiated specimens exhibited a greatly reduced osteogenic differentiation capacity, and during the first two weeks proliferation rates of explanted cells were greatly diminished. Thereafter, cells recovered and showed proliferation behaviour comparable to control samples. These results imply that MSCs can cope with irradiation up to relatively high doses

  13. Tuning and sensitivity of the human vestibular system to low-frequency vibration.

    Science.gov (United States)

    Todd, Neil P McAngus; Rosengren, Sally M; Colebatch, James G

    2008-10-17

    Mechanoreceptive hair-cells of the vertebrate inner ear have a remarkable sensitivity to displacement, whether excited by sound, whole-body acceleration or substrate-borne vibration. In response to seismic or substrate-borne vibration, thresholds for vestibular afferent fibre activation have been reported in anamniotes (fish and frogs) in the range -120 to -90 dB re 1g. In this article, we demonstrate for the first time that the human vestibular system is also extremely sensitive to low-frequency and infrasound vibrations by making use of a new technique for measuring vestibular activation, via the vestibulo-ocular reflex (VOR). We found a highly tuned response to whole-head vibration in the transmastoid plane with a best frequency of about 100 Hz. At the best frequency we obtained VOR responses at intensities of less than -70 dB re 1g, which was 15 dB lower than the threshold of hearing for bone-conducted sound in humans at this frequency. Given the likely synaptic attenuation of the VOR pathway, human receptor sensitivity is probably an order of magnitude lower, thus approaching the seismic sensitivity of the frog ear. These results extend our knowledge of vibration-sensitivity of vestibular afferents but also are remarkable as they indicate that the seismic sensitivity of the human vestibular system exceeds that of the cochlea for low-frequencies.

  14. Hapten-specific lymphocyte transformation in humans sensitized with NDMA or DNCB.

    Science.gov (United States)

    SoebergB; Andersen, V

    1976-01-01

    The primary immune response to a contact sensitizing dose of para-N-dimethylnitrosaniline (NDMA) and dinitrochlorobenzene (DNCB) was obtained in humans and measured in vitro by increased thymidine incorporation into sensitized lymphocytes. No cross-reaction was found between these two haptens, and it is thus possible on two separate occasions to quantify and follow the primary cellular immune response in man. PMID:963911

  15. Lack of influence of GTP cyclohydrolase gene (GCH1 variations on pain sensitivity in humans

    Directory of Open Access Journals (Sweden)

    Dionne Raymond A

    2007-03-01

    Full Text Available Abstract Objectives To assess the effect of variations in GTP cyclohydrolase gene (GCH1 on pain sensitivity in humans. Methods Thermal and cold pain sensitivity were evaluated in a cohort of 735 healthy volunteers. Among this cohort, the clinical pain responses of 221 subjects after the surgical removal of impacted third molars were evaluated. Genotyping was done for 38 single nucleotide polymorphisms (SNPs whose heterozygosity > 0.2 in GCH1. Influence of the genetic variations including SNPs and haplotypes on pain sensitivity were analyzed. Results Minor allele frequencies and linkage disequilibrium show significant differences in European Americans, African Americans, Hispanic Americans and Asian Americans. Association analyses in European Americans do not replicate the previously reported important influence of GCH1 variations on pain sensitivity. Conclusion Considering population stratification, previously reported associations between GCH1 genetic variations and pain sensitivity appear weak or negligible in this well characterized model of pain.

  16. Hyperbaric oxygen therapy attenuates central sensitization induced by a thermal injury in humans

    DEFF Research Database (Denmark)

    Rasmussen, V M; Borgen, A E; Jansen, E C

    2015-01-01

    BACKGROUND: Hyperbaric oxygen (HBO2 ) treatment has in animal experiments demonstrated antinociceptive effects. It was hypothesized that these effects would attenuate secondary hyperalgesia areas (SHAs), an expression of central sensitization, after a first-degree thermal injury in humans. METHODS...... was demonstrated. However, in the nine volunteers starting with the control session, a statistical significant attenuation of SHAs was demonstrated in the HBO2 session (P = 0.004). CONCLUSIONS: The results indicate that HBO2 therapy in humans attenuates central sensitization induced by a thermal skin injury......, compared with control. These new and original findings in humans corroborate animal experimental data. The thermal injury model may give impetus to future human neurophysiological studies exploring the central effects of hyperbaric oxygen treatment....

  17. Sensitivity of C6 Glioma Cells Carrying the Human Poliovirus Receptor to Oncolytic Polioviruses.

    Science.gov (United States)

    Sosnovtseva, A O; Lipatova, A V; Grinenko, N F; Baklaushev, V P; Chumakov, P M; Chekhonin, V P

    2016-10-01

    A humanized line of rat C6 glioma cells expressing human poliovirus receptor was obtained and tested for the sensitivity to oncolytic effects of vaccine strains of type 1, 2, and 3 polioviruses. Presentation of the poliovirus receptor on the surface of C6 glioma cells was shown to be a necessary condition for the interaction of cells with polioviruses, but insufficient for complete poliovirus oncolysis.

  18. Muscular heat and mechanical pain sensitivity after lengthening contractions in humans and animals.

    Science.gov (United States)

    Queme, Fernando; Taguchi, Toru; Mizumura, Kazue; Graven-Nielsen, Thomas

    2013-11-01

    Mechanical sensitivity of muscle nociceptors was previously shown to increase 2 days after lengthening contractions (LC), but heat sensitivity was not different despite nerve growth factor (NGF) being upregulated in the muscle during delayed-onset muscle soreness (DOMS). The discrepancy of these results and lack of other reports drove us to assess heat sensitivity during DOMS in humans and to evaluate the effect of NGF on the heat response of muscle C-fibers. Pressure pain thresholds and pain intensity scores to intramuscular injection of isotonic saline at 48°C and capsaicin were recorded in humans after inducing DOMS. The response of single unmyelinated afferents to mechanical and heat stimulations applied to their receptive field was recorded from muscle-nerve preparations in vitro. In humans, pressure pain thresholds were reduced but heat and capsaicin pain responses were not increased during DOMS. In rats, the mechanical but not the heat sensitivity of muscle C-fibers was increased in the LC group. NGF applied to the receptive field facilitated the heat sensitivity relative to the control. The absence of facilitated heat sensitivity after LC, despite the NGF sensitization, may be explained if the NGF concentration produced after LC is not sufficient to sensitize nociceptor response to heat. This article presents new findings on the basic mechanisms underlying hyperalgesia during DOMS, which is a useful model to study myofascial pain syndrome, and the role of NGF on muscular nociception. This might be useful in the search for new pharmacologic targets and therapeutic approaches. Copyright © 2013 American Pain Society. Published by Elsevier Inc. All rights reserved.

  19. Radioimmunoelectrophoresis, a sensitive method for detecting cleavage of the fifth component of human complement (C5)

    International Nuclear Information System (INIS)

    Perez, H.D.; Ong, R.; Banda, D.; Goldstein, I.M.

    1983-01-01

    A method has been developed for detecting cleavage of human C5 in serum and whole blood as a consequence of complement activation. Standard, single-dimension immunoelectrophoresis was performed using as antibody a radioiodinated IgG fraction prepared from a commercially available antiserum to human C5. Autoradiographs developed after radioimmunoelectrophoresis of either normal human serum or functionally pure human C5 revealed only one precipitin band. In contrast, when either zymosan-treated serum or trypsin-treated human C5 were examined with this technique, two additional precipitin bands were detected. One migrated more anodally than native C5 while the other remained at the origin (cathode). Radioimmunoelectrophoresis was significantly more sensitive as an indicator of complement activation in human serum than either measurements of total hemolytic complement or a standard assay for complement (C5)-derived chemotactic activity. (Auth.)

  20. Inhibition of Notch1 increases paclitaxel sensitivity to human breast cancer

    Institute of Scientific and Technical Information of China (English)

    Zhao Li; Ma Yongjie; Gu Feng; Fu Li

    2014-01-01

    Background Paclitaxel (PAC) is the first-line chemotherapy drug for most breast cancer patients,but clinical studies showed that some breast cancer patients were insensitive to PAC,which led to chemotherapy failure.It was reported that Notch1 signaling participated in drug resistance of breast cancer.Here,we show whether Notch1 expression is related to PAC sensitivity of breast cancer.Methods We employed Notch1 siRNA and Notch1 inhibitor,N-[N-(3,5-difluorophenacetyl)-1-alanyl]-S-phenylglycine t-butylester (DAPT),to down regulate Notch1 expression in human breast cancer cells MDA-MB-231,and detected the inhibition effect by Western blotting and reverse trans cription-polymerase chain reaction,respectively.After 24 hours exposure to different concentration of PAC (0,1,5,10,15,20,and 25 μg/ml),the viability of the control group and experimental group cells was tested by MTT.We also examined the expression of Notch1 in PAC sensitive and nonsensitive breast cancer patients,respectively by immunohistochemistry (IHC).The PAC sensitivity of breast cancer patients were identified by collagen gel droplet embedded culture-drug sensitivity test (CD-DST).Results Down regulation of Notch1 expression by Notch1siRNA interference or Notch1 inhibitor increased the PAC sensitivity in MDA-MB-231 cells (P <0.05).Also,the expression of Notch1 in PAC sensitive patients was much lower than that of PAC non-sensitive patients (P <0.01).Conclusion Notch1 expression has an effect on PAC sensitivity in breast cancer patients,and the inhibition of Notch1 increases paclitaxel sensitivity to human breast cancer.

  1. Human sensitivity to eye contact in 2D and 3D videoconferencing

    NARCIS (Netherlands)

    Eijk, van R.L.J.; Kuijsters, A.; Dijkstra, K.I.; IJsselsteijn, W.A.

    2010-01-01

    Gaze awareness and eye contact serve important functions in social interaction. In order to maintain those functions in 2D and 3D videoconferencing systems, human sensitivity to eye contact and gaze direction needs to be taken into account in the design of such systems. Here we experimentally

  2. Tumor necrosis factor alpha selectively sensitizes human immunodeficiency virus-infected cells to heat and radiation

    International Nuclear Information System (INIS)

    Wong, G.H.; McHugh, T.; Weber, R.; Goeddel, D.V.

    1991-01-01

    We report here that infection of the human T-cell line HUT-78 with human immunodeficiency virus (HIV) increases its sensitivity to heat and radiation toxicity. A possible explanation for this result may be the reduced expression of manganous superoxide dismutase (MnSOD) in HIV-infected cells compared to uninfected cells. Tumor necrosis factor alpha (TNF-alpha) further sensitizes HIV-infected cells but not uninfected cells to heat and radiation. This is consistent with the ability of TNF-alpha to induce the expression of MnSOD in uninfected but not in HIV-infected cells. HIV-infected HUT-78 cell lines engineered to overexpress MnSOD are more resistant to heat and radiation than HIV-infected cells that do not overexpress MnSOD. However, treatment with TNF-alpha still sensitizes these cells to heat and radiation

  3. Calcineurin inhibitors acutely improve insulin sensitivity without affecting insulin secretion in healthy human volunteers

    DEFF Research Database (Denmark)

    Øzbay, Aygen; Møller, Niels; Juhl, Claus

    2012-01-01

    and tacrolimus has been attributed to both beta cell dysfunction and impaired insulin sensitivity. WHAT THIS STUDY ADDS: This is the first trial to investigate beta cell function and insulin sensitivity using gold standard methodology in healthy human volunteers treated with clinically relevant doses...... of ciclosporin and tacrolimus. We document that both drugs acutely increase insulin sensitivity, while first phase and pulsatile insulin secretion remain unaffected. This study demonstrates that ciclosporin and tacrolimus have similar acute effects on glucose metabolism in healthy humans. AIM The introduction...... of calcineurin inhibitors (CNIs) ciclosporin (CsA) and tacrolimus (Tac) has improved the outcome of organ transplants, but complications such as new onset diabetes mellitus after transplantation (NODAT) cause impairment of survival rates. The relative contribution of each CNI to the pathogenesis and development...

  4. Culture-sensitive neural substrates of human cognition: a transcultural neuroimaging approach.

    Science.gov (United States)

    Han, Shihui; Northoff, Georg

    2008-08-01

    Our brains and minds are shaped by our experiences, which mainly occur in the context of the culture in which we develop and live. Although psychologists have provided abundant evidence for diversity of human cognition and behaviour across cultures, the question of whether the neural correlates of human cognition are also culture-dependent is often not considered by neuroscientists. However, recent transcultural neuroimaging studies have demonstrated that one's cultural background can influence the neural activity that underlies both high- and low-level cognitive functions. The findings provide a novel approach by which to distinguish culture-sensitive from culture-invariant neural mechanisms of human cognition.

  5. Effect of training on insulin sensitivity of glucose uptake and lipolysis in human adipose tissue

    DEFF Research Database (Denmark)

    Stallknecht, B; Larsen, J J; Mikines, K J

    2000-01-01

    Training increases insulin sensitivity of both whole body and muscle in humans. To investigate whether training also increases insulin sensitivity of adipose tissue, we performed a three-step hyperinsulinemic, euglycemic clamp in eight endurance-trained (T) and eight sedentary (S) young men...... (glucose only). Adipose tissue blood flow was measured by (133)Xe washout. In the basal state, adipose tissue blood flow tended to be higher in T compared with S subjects, and in both groups blood flow was constant during the clamp. The change from basal in arterial-interstitial glucose concentration......-time: T, 44 +/- 9 min (n = 7); S, 102 +/- 23 min (n = 5); P training enhances insulin sensitivity of glucose uptake in subcutaneous adipose tissue and in skeletal muscle. Furthermore, interstitial glycerol data suggest that training also increases insulin sensitivity of lipolysis...

  6. Human chromosome 9 can complement UV sensitivity of xeroderma pigmentosum group A cells

    International Nuclear Information System (INIS)

    Ishizaki, Kanji; Sasaki, Masao S.; Ikenaga, Mituo; Nakamura, Yusuke

    1990-01-01

    A single human chromosome derived from normal human fibroblasts and tagged with the G418 resistance gene was transferred into SV40-transformed xeroderma pigmentosum group A (XP-A) cells via microcell fusion. When chromosome 1 or 12 was transferred, UV sensitivity of microcell hybrid cells was not changed. By contrast, after transferring chromosome 9,7 of 11 reipient clones were as UV-resistant as normal human cells. Four other clones were still as UV-sensitive as the parental XP-A cells. Southern hybridization analysis using a polymorphic probe, pEKZ19.3, which is homologous to a sequence of the D9S17 locus on chromosome 9, has confirmed that at least a part of normal human chromosome 9 was transferred into the recipient clones. However, amounts iof UV-induced unscheduled DNA synthesis in the UV-resistant clones were only one-third of those in normal human cells. These results indicate that a gene on chromosome 9 can confer complementation of high UV sensitivity of XP-A cells although it is still possible that 2 or more genes might be involved in the defective-repair phenotypes of XP-A. (author). 20 refs.; 3 figs.; 1 tab

  7. In vitro sensitization of human lymphocytes to a myeloma cell-related antigen

    International Nuclear Information System (INIS)

    Whitson, M.E.; Griffin, G.D.; Novelli, G.D.; Solomon, A.

    1981-01-01

    Peripheral blood lymphocytes from normal human donors were cocultivated with cells from two established human multiple myeloma cell lines, RPMI 8226 and K-737, and with lymphoblastoid cells from a third B cell line, RAMM. After a comparison of three methods of lymphocyte sensitization, a 6-day incubation protocol with equal numbers of normal lymphocytes and mitomycin C-treated tumor cells was selected. Cells fom the RPMI 8226 myeloma line stimulated the differentiation of lymphocytes into cytotoxic effector cells as measured by 51 Cr release from labeled target cells. The RPMI 8226-sensitized lymphocytes were cytotoxic for myeloma cells (RPMI 8226 and K-737) and for lymphoblastoid cells (RAMM) but not for cells from human lung tumor lines (A549, A427, MB9812), a breast carcinoma line (ALAB), a normal diploid fibroblast line (HSBP), or normal lymphocytes

  8. In vitro sensitization of human lymphocytes to a myeloma cell-related antigen

    Energy Technology Data Exchange (ETDEWEB)

    Whitson, M.E. (Univ. of South Carolina, Columbia); Griffin, G.D.; Novelli, G.D.; Solomon, A.

    1981-01-01

    Peripheral blood lymphocytes from normal human donors were cocultivated with cells from two established human multiple myeloma cell lines, RPMI 8226 and K-737, and with lymphoblastoid cells from a third B cell line, RAMM. After a comparison of three methods of lymphocyte sensitization, a 6-day incubation protocol with equal numbers of normal lymphocytes and mitomycin C-treated tumor cells was selected. Cells fom the RPMI 8226 myeloma line stimulated the differentiation of lymphocytes into cytotoxic effector cells as measured by /sup 51/Cr release from labeled target cells. The RPMI 8226-sensitized lymphocytes were cytotoxic for myeloma cells (RPMI 8226 and K-737) and for lymphoblastoid cells (RAMM) but not for cells from human lung tumor lines (A549, A427, MB9812), a breast carcinoma line (ALAB), a normal diploid fibroblast line (HSBP), or normal lymphocytes.

  9. Distinct fermentation and antibiotic sensitivity profiles exist in salmonellae of canine and human origin.

    Science.gov (United States)

    Wallis, Corrin V; Lowden, Preena; Marshall-Jones, Zoe V; Hilton, Anthony C

    2018-02-26

    Salmonella enterica is a recognised cause of diarrhoea in dogs and humans, yet the potential for transfer of salmonellosis between dogs and their owners is unclear, with reported evidence both for and against Salmonella as a zoonotic pathogen. A collection of 174 S. enterica isolates from clinical infections in humans and dogs were analysed for serotype distribution, carbon source utilisation, chemical and antimicrobial sensitivity profiles. The aim of the study was to understand the degree of conservation in phenotypic characteristics of isolates across host species. Serovar distribution across human and canine isolates demonstrated nine serovars common to both host species, 24 serovars present in only the canine collection and 39 solely represented within the human collection. Significant differences in carbon source utilisation profiles and ampicillin, amoxicillin and chloramphenicol sensitivity profiles were detected in isolates of human and canine origin. Differences between the human and canine Salmonella collections were suggestive of evolutionary separation, with canine isolates better able to utilise several simple sugars than their human counterparts. Generally higher minimum inhibitory concentrations of three broad-spectrum antimicrobials, commonly used in veterinary medicine, were also observed in canine S. enterica isolates. Differential carbon source utilisation and antimicrobial sensitivity profiles in pathogenic Salmonella isolated from humans and dogs are suggestive of distinct reservoirs of infection for these hosts. Although these findings do not preclude zoonotic or anthroponotic potential in salmonellae, the separation of carbon utilisation and antibiotic profiles with isolate source is indicative that infectious isolates are not part of a common reservoir shared frequently between these host species.

  10. Comparative sensitivity of human and rat neural cultures to chemical-induced inhibition of neurite outgrowth

    Energy Technology Data Exchange (ETDEWEB)

    Harrill, Joshua A.; Freudenrich, Theresa M.; Robinette, Brian L.; Mundy, William R., E-mail: mundy.william@epa.gov

    2011-11-15

    There is a need for rapid, efficient and cost-effective alternatives to traditional in vivo developmental neurotoxicity testing. In vitro cell culture models can recapitulate many of the key cellular processes of nervous system development, including neurite outgrowth, and may be used as screening tools to identify potential developmental neurotoxicants. The present study compared primary rat cortical cultures and human embryonic stem cell-derived neural cultures in terms of: 1) reproducibility of high content image analysis based neurite outgrowth measurements, 2) dynamic range of neurite outgrowth measurements and 3) sensitivity to chemicals which have been shown to inhibit neurite outgrowth. There was a large increase in neurite outgrowth between 2 and 24 h in both rat and human cultures. Image analysis data collected across multiple cultures demonstrated that neurite outgrowth measurements in rat cortical cultures were more reproducible and had higher dynamic range as compared to human neural cultures. Human neural cultures were more sensitive than rat cortical cultures to chemicals previously shown to inhibit neurite outgrowth. Parallel analysis of morphological (neurite count, neurite length) and cytotoxicity (neurons per field) measurements were used to detect selective effects on neurite outgrowth. All chemicals which inhibited neurite outgrowth in rat cortical cultures did so at concentrations which did not concurrently affect the number of neurons per field, indicating selective effects on neurite outgrowth. In contrast, more than half the chemicals which inhibited neurite outgrowth in human neural cultures did so at concentrations which concurrently decreased the number of neurons per field, indicating that effects on neurite outgrowth were secondary to cytotoxicity. Overall, these data demonstrate that the culture models performed differently in terms of reproducibility, dynamic range and sensitivity to neurite outgrowth inhibitors. While human neural

  11. Comparative sensitivity of human and rat neural cultures to chemical-induced inhibition of neurite outgrowth

    International Nuclear Information System (INIS)

    Harrill, Joshua A.; Freudenrich, Theresa M.; Robinette, Brian L.; Mundy, William R.

    2011-01-01

    There is a need for rapid, efficient and cost-effective alternatives to traditional in vivo developmental neurotoxicity testing. In vitro cell culture models can recapitulate many of the key cellular processes of nervous system development, including neurite outgrowth, and may be used as screening tools to identify potential developmental neurotoxicants. The present study compared primary rat cortical cultures and human embryonic stem cell-derived neural cultures in terms of: 1) reproducibility of high content image analysis based neurite outgrowth measurements, 2) dynamic range of neurite outgrowth measurements and 3) sensitivity to chemicals which have been shown to inhibit neurite outgrowth. There was a large increase in neurite outgrowth between 2 and 24 h in both rat and human cultures. Image analysis data collected across multiple cultures demonstrated that neurite outgrowth measurements in rat cortical cultures were more reproducible and had higher dynamic range as compared to human neural cultures. Human neural cultures were more sensitive than rat cortical cultures to chemicals previously shown to inhibit neurite outgrowth. Parallel analysis of morphological (neurite count, neurite length) and cytotoxicity (neurons per field) measurements were used to detect selective effects on neurite outgrowth. All chemicals which inhibited neurite outgrowth in rat cortical cultures did so at concentrations which did not concurrently affect the number of neurons per field, indicating selective effects on neurite outgrowth. In contrast, more than half the chemicals which inhibited neurite outgrowth in human neural cultures did so at concentrations which concurrently decreased the number of neurons per field, indicating that effects on neurite outgrowth were secondary to cytotoxicity. Overall, these data demonstrate that the culture models performed differently in terms of reproducibility, dynamic range and sensitivity to neurite outgrowth inhibitors. While human neural

  12. Androgen Metabolism in Progression to Androgen-Independent Prostate Cancer

    Science.gov (United States)

    2011-06-01

    described CRPC bone marrow metastases (8), we detected TMPRSS2:ERG transcripts (TMPRSS2 exon 2-ERG exon 4) in 11 of 29 cases. Affymetrix oligonucleotide...and immunoblotted. B, RT-PCR for ERG ( exon 9/10), TMPRSS2 ( exon 5/6), and PSA mRNA after DHT stimulation. C, cells in CSS medium treated with DHT and...therapeutic index CYP3A4 sub- strates were excluded. The treatment was ketoconazole 400 mg orally thrice daily, hydro- cortisone (30 mg/AM and 10 mg/PM

  13. The variation field of the radiosensitivity in the human population: hypersensitivity and hypo sensitivity

    International Nuclear Information System (INIS)

    Bouffler, S.

    2009-01-01

    It is generally admitted in the radiotherapy practice that 5% of patients will present serious reactions of normal tissues. Studies made on human population confirm it exists variations of cell radiosensitivity and genetic factors contribute in a significant manner to the observed variations. The researches of markers able to predict the reactions of normal tissues to the therapy focussed on the identification of sensitive sub-group. It is however obvious that exists also a part of the population relatively hypo sensitive. It would be interesting to make studies on the genome to find genes associated to serious reactions of normal tissues to radiotherapy. (N.C.)

  14. Is Cytox 3522 (10% methylene-bis-thiocyanate) a human skin sensitizer?

    DEFF Research Database (Denmark)

    Andersen, Klaus Ejner; Hamann, K

    1983-01-01

    Methylene-bis-thiocyanate is an antimicrobial agent in Cytox 3522 (American Cyanamid Corporation) and Nalco 206 (Nalco Chemical Company). Both are wide-spectrum industrial biocides. Cytox 3522 showed a strong sensitization potential in guineau pigs using the Guinea Pig Maximization Test and the O...... and the Open Epicutaneous Test procedures. In spite of this, human sensitization has not been reported. This may be explained by several factors such as limited exposure, low-use concentration and a lack of diagnostic patch tests in selected patients....

  15. UVA photolysis using the protein-bound sensitizers present in human lens

    International Nuclear Information System (INIS)

    Ortwerth, B.J.; Olesen, P.R.

    1994-01-01

    This research was undertaken to demonstrate that the protein-bound chromophores in aged human lens can act as sensitizers for protein damage by UVA light. The water-insoluble (WI) proteins from pooled human and bovine lenses were solubilized by sonication in water and illuminated with UV light similar in output to that transmitted by the cornea. Analysis of the irradiated proteins showed a linear decrease in sulfhydryl groups with a 30% loss after 2 h. No loss was seen when native α-crystallin was irradiated under the same conditions. A 25% loss of histidine residues was also observed with the human lens WI fraction, and sodium dodecyl sulfate polyacrylamide gels indicated considerable protein cross-linking. Similar photodamage was seen with a WI fraction from old bovine lenses. While the data show the presence of UVA sensitizers, some histidine destruction and protein cross-linking were also obtained with α-crystallin and with lysozyme which argue that part of the histidine loss in the human WISS was likely due to tryptophan acting as a sensitizer. (Author)

  16. Isolation of uv-sensitive variants of human FL cells by a viral suicide method

    International Nuclear Information System (INIS)

    Shiomi, T.; Sato, K.

    1979-01-01

    A new method (viral suicide method) for the isolation of uv-sensitive mutants is described. Colonies of mutagenized human FL cells were infected with uv-irradiated Herpes simplex viruses and surviving ones which seemed to be deficient in host cell reactivation (HCR) were examined for their uv sensitivity. Nineteen of 238 clones examined were sensitive to uv irradiation at the time of the isolation. After recloning, four of these clones have been studied and two (UVS-1 and UVS-2) of them are stable in their uv sensitivity for 4 months in culture. uv sensitivity of UVS-1, UVS-2, and the parental FL cells are as follows: the extrapolation numbers (n) are 2.2, 2.1, and 1.8 and mean lethal doses (DO) are 2.9, 3.7, and 7.8 J/m 2 for UVS-1, UVS-2, and the parental FL cells, respectively. They are no more sensitive than FL cells to x-irradiation. The ability of HCR in UVS-2 cells is apparently lower than that in FL cells, whereas UVS-1 cells are the same as FL cells in the ability

  17. Advances in radiation biology: Relative radiation sensitivities of human organ systems. Volume 12

    International Nuclear Information System (INIS)

    Lett, J.T.; Altman, K.I.; Ehmann, U.K.; Cox, A.B.

    1987-01-01

    This volume is a thematically focused issue of Advances in Radiation Biology. The topic surveyed is relative radiosensitivity of human organ systems. Topics considered include relative radiosensitivities of the thymus, spleen, and lymphohemopoietic systems; relative radiosensitivities of the small and large intestine; relative rediosensitivities of the oral cavity, larynx, pharynx, and esophagus; relative radiation sensitivity of the integumentary system; dose response of the epidermal; microvascular, and dermal populations; relative radiosensitivity of the human lung; relative radiosensitivity of fetal tissues; and tolerance of the central and peripheral nervous system to therapeutic irradiation

  18. Advances in radiation biology: Relative radiation sensitivities of human organ systems. Volume 12

    Energy Technology Data Exchange (ETDEWEB)

    Lett, J.T.; Altman, K.I.; Ehmann, U.K.; Cox, A.B.

    1987-01-01

    This volume is a thematically focused issue of Advances in Radiation Biology. The topic surveyed is relative radiosensitivity of human organ systems. Topics considered include relative radiosensitivities of the thymus, spleen, and lymphohemopoietic systems; relative radiosensitivities of the small and large intestine; relative rediosensitivities of the oral cavity, larynx, pharynx, and esophagus; relative radiation sensitivity of the integumentary system; dose response of the epidermal; microvascular, and dermal populations; relative radiosensitivity of the human lung; relative radiosensitivity of fetal tissues; and tolerance of the central and peripheral nervous system to therapeutic irradiation.

  19. Growth inhibitory effects of the dual ErbB1/ErbB2 tyrosine kinase inhibitor PKI-166 on human prostate cancer xenografts.

    Science.gov (United States)

    Mellinghoff, Ingo K; Tran, Chris; Sawyers, Charles L

    2002-09-15

    Experiments with human prostate cancer cell lines have shown that forced overexpression of the ErbB2-receptor tyrosine kinase (RTK) promotes androgen-independent growth and increases androgen receptor-transcriptional activity in a ligand-independent fashion. To investigate the relationship between ErbB-RTK signaling and androgen in genetically unmanipulated human prostate cancer, we performed biochemical and biological studies with the dual ErbB1/ErbB2 RTK inhibitor PKI-166 using human prostate cancer xenograft models with isogenic sublines reflecting the transition from androgen-dependent to androgen-independent growth. In the presence of low androgen concentrations, PKI-166 showed profound growth-inhibitory effects on tumor growth, which could be partially reversed by androgen add-back. At physiological androgen concentrations, androgen withdrawal greatly enhanced the ability of PKI-166 to retard tumor growth. The level of extracellular signal-regulated kinase activation correlated with the response to PKI-166 treatment, whereas the expression levels of ErbB1 and ErbB2 did not. These results suggest that ErbB1/ErbB2 RTKs play an important role in the biology of androgen-independent prostate cancer and provide a rationale for clinical evaluation of inhibitors targeted to this pathway.

  20. Cryptolepine, isolated from Sida acuta, sensitizes human gastric adenocarcinoma cells to TRAIL-induced apoptosis.

    Science.gov (United States)

    Ahmed, Firoj; Toume, Kazufumi; Ohtsuki, Takashi; Rahman, Mahmudur; Sadhu, Samir Kumar; Ishibashi, Masami

    2011-01-01

    Bioassay guided separation of Sida acuta whole plants led to the isolation of an alkaloid, cryptolepine (1), along with two kaempferol glycosides (2-3). Compound 1 showed strong activity in overcoming TRAIL-resistance in human gastric adenocarcinoma (AGS) cells at 1.25, 2.5 and 5 μm. Combined treatment of 1 and TRAIL sensitized AGS cells to TRAIL-induced apoptosis at the aforementioned concentrations. Copyright © 2010 John Wiley & Sons, Ltd.

  1. Sensitivity field distributions for segmental bioelectrical impedance analysis based on real human anatomy

    International Nuclear Information System (INIS)

    Danilov, A A; Rudnev, S G; V Vassilevski, Yu; Kramarenko, V K; Nikolaev, D V; Smirnov, A V; Salamatova, V Yu

    2013-01-01

    In this work, an adaptive unstructured tetrahedral mesh generation technology is applied for simulation of segmental bioimpedance measurements using high-resolution whole-body model of the Visible Human Project man. Sensitivity field distributions for a conventional tetrapolar, as well as eight- and ten-electrode measurement configurations are obtained. Based on the ten-electrode configuration, we suggest an algorithm for monitoring changes in the upper lung area.

  2. Sensitivity field distributions for segmental bioelectrical impedance analysis based on real human anatomy

    Science.gov (United States)

    Danilov, A. A.; Kramarenko, V. K.; Nikolaev, D. V.; Rudnev, S. G.; Salamatova, V. Yu; Smirnov, A. V.; Vassilevski, Yu V.

    2013-04-01

    In this work, an adaptive unstructured tetrahedral mesh generation technology is applied for simulation of segmental bioimpedance measurements using high-resolution whole-body model of the Visible Human Project man. Sensitivity field distributions for a conventional tetrapolar, as well as eight- and ten-electrode measurement configurations are obtained. Based on the ten-electrode configuration, we suggest an algorithm for monitoring changes in the upper lung area.

  3. Skeletal muscle phosphatidylcholine fatty acids and insulin sensitivity in normal humans.

    Science.gov (United States)

    Clore, J N; Li, J; Gill, R; Gupta, S; Spencer, R; Azzam, A; Zuelzer, W; Rizzo, W B; Blackard, W G

    1998-10-01

    The fatty acid composition of skeletal muscle membrane phospholipids (PL) is known to influence insulin responsiveness in humans. However, the contribution of the major PL of the outer (phosphatidylcholine, PC) and inner (phosphatidylethanolamine, PE) layers of the sarcolemma to insulin sensitivity is not known. Fatty acid composition of PC and PE from biopsies of vastus lateralis from 27 normal men and women were correlated with insulin sensitivity determined by the hyperinsulinemic euglycemic clamp technique at insulin infusion rates of 0.4, 1.0, and 10.0 mU . kg-1 . min-1. Significant variation in the half-maximal insulin concentration (ED50) was observed in the normal volunteers (range 24.0-146.0 microU/ml), which correlated directly with fasting plasma insulin (r = 0.75, P insulin sensitivity was observed in PE (NS). These studies suggest that the fatty acid composition of PC may be of particular importance in the relationship between fatty acids and insulin sensitivity in normal humans.

  4. Perceiving polarization with the naked eye: characterization of human polarization sensitivity

    Science.gov (United States)

    Temple, Shelby E.; McGregor, Juliette E.; Miles, Camilla; Graham, Laura; Miller, Josie; Buck, Jordan; Scott-Samuel, Nicholas E.; Roberts, Nicholas W.

    2015-01-01

    Like many animals, humans are sensitive to the polarization of light. We can detect the angle of polarization using an entoptic phenomenon called Haidinger's brushes, which is mediated by dichroic carotenoids in the macula lutea. While previous studies have characterized the spectral sensitivity of Haidinger's brushes, other aspects remain unexplored. We developed a novel methodology for presenting gratings in polarization-only contrast at varying degrees of polarization in order to measure the lower limits of human polarized light detection. Participants were, on average, able to perform the task down to a threshold of 56%, with some able to go as low as 23%. This makes humans the most sensitive vertebrate tested to date. Additionally, we quantified a nonlinear relationship between presented and perceived polarization angle when an observer is presented with a rotatable polarized light field. This result confirms a previous theoretical prediction of how uniaxial corneal birefringence impacts the perception of Haidinger's brushes. The rotational dynamics of Haidinger's brushes were then used to calculate corneal retardance. We suggest that psychophysical experiments, based upon the perception of polarized light, are amenable to the production of affordable technologies for self-assessment and longitudinal monitoring of visual dysfunctions such as age-related macular degeneration. PMID:26136441

  5. Neural prediction errors reveal a risk-sensitive reinforcement-learning process in the human brain.

    Science.gov (United States)

    Niv, Yael; Edlund, Jeffrey A; Dayan, Peter; O'Doherty, John P

    2012-01-11

    Humans and animals are exquisitely, though idiosyncratically, sensitive to risk or variance in the outcomes of their actions. Economic, psychological, and neural aspects of this are well studied when information about risk is provided explicitly. However, we must normally learn about outcomes from experience, through trial and error. Traditional models of such reinforcement learning focus on learning about the mean reward value of cues and ignore higher order moments such as variance. We used fMRI to test whether the neural correlates of human reinforcement learning are sensitive to experienced risk. Our analysis focused on anatomically delineated regions of a priori interest in the nucleus accumbens, where blood oxygenation level-dependent (BOLD) signals have been suggested as correlating with quantities derived from reinforcement learning. We first provide unbiased evidence that the raw BOLD signal in these regions corresponds closely to a reward prediction error. We then derive from this signal the learned values of cues that predict rewards of equal mean but different variance and show that these values are indeed modulated by experienced risk. Moreover, a close neurometric-psychometric coupling exists between the fluctuations of the experience-based evaluations of risky options that we measured neurally and the fluctuations in behavioral risk aversion. This suggests that risk sensitivity is integral to human learning, illuminating economic models of choice, neuroscientific models of affective learning, and the workings of the underlying neural mechanisms.

  6. Sensitive radioimmunoassay for detection of antibodies to recombinant human interferon-alpha A

    International Nuclear Information System (INIS)

    Palleroni, A.V.; Trown, P.W.

    1986-01-01

    A radioimmunoassay (RIA) for the detection of antibodies to recombinant human leukocyte interferon A (rHuIFN-alpha A) in human serum has been developed and validated against the standard antiviral neutralization bioassay (ANB). The assay measures the binding of 125 I-labeled rHuIFN-alpha A to immunoglobulins in serum. Aliquots of patients' sera are incubated with 125 I-rHuIFN-alpha A and the complexes formed between antibodies in the sera and the 125 I-rHuIFN-alpha A are precipitated with goat anti-human IgG serum. The radioactivity in the immune precipitate is a measure of the quantity of antibody (if present) in the serum. The sensitivity of this RIA is 5 ng of IgG/ml of serum

  7. The study on the effect of artesunate on the radio-sensitivity of human cervical cancer

    International Nuclear Information System (INIS)

    Geng Chong; Cao Jianping; Ni Qianying

    2011-01-01

    To investigate the effect of artesunate on radio-sensitivity of human cervical cancer cells in vitro. The human cervical cancer cells HeLa and Siha were used as the experimental cells. MTT assay was used to determine the most appropriate drug concentration in the subsequent experiment, and the effect of human cervical cancer cells treated with artesunate and irradiation of 60 Co γ-rays was studied by using conventional chromosomal aberration analysis and cytokinesis block method (CB method). The results show that when the concentration of artesunate in this experiment was 2.0 μmol/L for HeLa cell and 4.0 μmol/L for Siha cell respectively, the chromosome aberration, micronuclei cell and micronuclei rates of HeLa cells treated with artesunate were more serious than that of the only irradiation, but there is almost no change with Siha cells. (authors)

  8. γ-radiation induces cellular sensitivity and aberrant methylation in human tumor cell lines.

    Science.gov (United States)

    Kumar, Ashok; Rai, Padmalatha S; Upadhya, Raghavendra; Vishwanatha; Prasada, K Shama; Rao, B S Satish; Satyamoorthy, Kapettu

    2011-11-01

    Ionizing radiation induces cellular damage through both direct and indirect mechanisms, which may include effects from epigenetic changes. The purpose of this study was to determine the effect of ionizing radiation on DNA methylation patterns that may be associated with altered gene expression. Sixteen human tumor cell lines originating from various cancers were initially tested for radiation sensitivity by irradiating them with γ-radiation in vitro and subsequently, radiation sensitive and resistant cell lines were treated with different doses of a demethylating agent, 5-Aza-2'-Deoxycytidine (5-aza-dC) and a chromatin modifier, Trichostatin-A (TSA). Survival of these cell lines was measured using 3-(4, 5-Dimethylthiazol- 2-yl)-2, 5-diphenyltetrazolium (MTT) and clonogenic assays. The effect of radiation on global DNA methylation was measured using reverse phase high performance liquid chromatography (RP-HPLC). The transcription response of methylated gene promoters, from cyclin-dependent kinase inhibitor 2A (p16(INK4a)) and ataxia telangiectasia mutated (ATM) genes, to radiation was measured using a luciferase reporter assay. γ-radiation resistant (SiHa and MDAMB453) and sensitive (SaOS2 and WM115) tumor cell lines were examined for the relationship between radiation sensitivity and DNA methylation. Treatment of cells with 5-aza-dC and TSA prior to irradiation enhanced DNA strand breaks, G2/M phase arrest, apoptosis and cell death. Exposure to γ-radiation led to global demethylation in a time-dependent manner in tumor cells in relation to resistance and sensitivity to radiation with concomitant activation of p16(INK4a) and ATM gene promoters. These results provide important information on alterations in DNA methylation as one of the determinants of radiation effects, which may be associated with altered gene expression. Our results may help in delineating the mechanisms of radiation resistance in tumor cells, which can influence diagnosis, prognosis and

  9. S100A10 protein expression is associated with oxaliplatin sensitivity in human colorectal cancer cells

    Directory of Open Access Journals (Sweden)

    Suzuki Sayo

    2011-12-01

    Full Text Available Abstract Background Individual responses to oxaliplatin (L-OHP-based chemotherapy remain unpredictable. The objective of our study was to find candidate protein markers for tumor sensitivity to L-OHP from intracellular proteins of human colorectal cancer (CRC cell lines. We performed expression difference mapping (EDM analysis of whole cell lysates from 11 human CRC cell lines with different sensitivities to L-OHP by using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS, and identified a candidate protein by liquid chromatography/mass spectrometry ion trap time-of-flight (LCMS-IT-TOF. Results Of the qualified mass peaks obtained by EDM analysis, 41 proteins were differentially expressed in 11 human colorectal cancer cell lines. Among these proteins, the peak intensity of 11.1 kDa protein was strongly correlated with the L-OHP sensitivity (50% inhibitory concentrations (P R2 = 0.80. We identified this protein as Protein S100-A10 (S100A10 by MS/MS ion search using LCMS-IT-TOF. We verified its differential expression and the correlation between S100A10 protein expression levels in drug-untreated CRC cells and their L-OHP sensitivities by Western blot analyses. In addition, S100A10 protein expression levels were not correlated with sensitivity to 5-fluorouracil, suggesting that S100A10 is more specific to L-OHP than to 5-fluorouracil in CRC cells. S100A10 was detected in cell culture supernatant, suggesting secretion out of cells. Conclusions By proteomic approaches including SELDI technology, we have demonstrated that intracellular S100A10 protein expression levels in drug-untreated CRC cells differ according to cell lines and are significantly correlated with sensitivity of CRC cells to L-OHP exposure. Our findings provide a new clue to searching predictive markers of the response to L-OHP, suggesting that S100A10 is expected to be one of the candidate protein markers.

  10. Assessment of metal sensitizer potency with the reconstructed human epidermis IL-18 assay.

    Science.gov (United States)

    Gibbs, Susan; Kosten, Ilona; Veldhuizen, Rosalien; Spiekstra, Sander; Corsini, Emanuela; Roggen, Erwin; Rustemeyer, Thomas; Feilzer, Albert J; Kleverlaan, Cees J

    2018-01-15

    According to the new EU Medical Devices (MDR) legislation coming into effect in 2017, manufactures will have to comply with higher standards of quality and safety for medical devices in order to meet common safety concerns regarding such products. Metal alloys are extensively used in dentistry and medicine (e.g. orthopedic surgery and cardiology) even though clinical experience suggests that many metals are sensitizers. The aim of this study was to further test the applicability domain of the in vitro reconstructed human epidermis (RhE) IL-18 assay developed to identify contact allergens and in doing so: i) determine whether different metal salts, representing leachables from metal alloys used in medical devices, could be correctly labelled and classified; and ii) assess the ability of different salts for the same metal to penetrate the skin stratum corneum. Twenty eight chemicals including 15 metal salts were topically exposed to RhE. Nickel, chrome, gold, palladium were each tested in two different salt forms, and titanium in 4 different salt forms. Metal salts were labelled (YES/NO) as sensitizer if a threshold of more than 5 fold IL18 release was reached. The in vitro estimation of expected sensitization induction level (potency) was assessed by interpolating in vitro EC50 and IL-18 SI2 with LLNA EC3 and human NOEL values from standard reference curves generated using DNCB (extreme) and benzocaine (weak). Metal salts, in contrast to other chemical sensitizers and with the exception of potassium dichromate (VI) and cobalt (II) chloride, were not identified as contact allergens since they only induced a small or no increase in IL-18 production. This finding was not related to a lack of stratum corneum skin penetration since EC50 values (decrease in metabolic activity; MTT assay) were obtained after topical RhE exposure to 8 of the 15 metal salts. For nickel, gold and palladium salts, differences in EC50 values between two salts for the same metal could not be

  11. Factors influencing the drug sensitization of human tumor cells for in situ lipofection.

    Science.gov (United States)

    Son, K; Huang, L

    1996-07-01

    The cisplatin induced enhancement of in situ lipofection was optimized by considering the factors that can increase the degree of sensitization. Two other anticancer drugs, mechlorethamine (nitrogen mustard) and taxol, enhanced CAT gene expression but the degree of sensitization was not as great as cisplatin. Besides human 2008 ovarian cancer cells we also found that human lung (A549) and head and neck cancer cells (SCC 25) were transiently sensitized by cisplatin. The transfectability of the two commercially available cationic liposomes, Lipofectin and LipofectAmine, was either weak or not consistent among tumors tested. In vivo transfection efficiency of 2008 cells was the highest at 1 microgram DNA per nmol or microgram liposome with all three cationic liposomes. In vitro transfection efficiency of 2008 cells at 1:1 (microgram of DNA:nmole of DC-chol/DOPE liposome) increased in a dose-dependent manner while at 1:10, an optimal ratio for in vitro lipofection, rapidly decreased with an increase in dose. This result indicated that there was a correlation between in vivo and in vitro lipofection at 1:1 ratio for delivering liposomal DNA. Most of the DNA injected into the tumor was concentrated in the tumor and in the skin above the tumor whether cisplatin was preinjected or liposomes were used as carriers.

  12. Females are sensitive to unpleasant human emotions regardless of the emotional context of photographs.

    Science.gov (United States)

    Kato, Ryousuke; Takeda, Yuji

    2017-06-09

    Previous studies have demonstrated that females exhibit higher sensitivity than males to the emotional state of a person in a photograph. The present study examined whether such females' sensitivity to human emotions could be observed even when the background emotional contexts were incongruent with facial expressions. The late positive potential (LPP) was measured while 19-female and 15-male participants viewed a photograph of a face with varied emotional expressions (pleasant, neutral, or unpleasant) superimposed on a background photograph with varied valences (pleasant, neutral, or unpleasant). The results showed that unpleasant background photographs elicited a larger LPP compared to pleasant and neutral background photographs in both female and male participants. In contrast, a larger LPP for the unpleasant face photographs was observed only in female participants. Furthermore, the effect of face photographs did not interact with the effect of background photographs. These results suggest that females are sensitive to human emotions regardless of the emotional context. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. DNA-mediated strand displacement facilitates sensitive electronic detection of antibodies in human serums.

    Science.gov (United States)

    Dou, Baoting; Yang, Jianmei; Shi, Kai; Yuan, Ruo; Xiang, Yun

    2016-09-15

    We describe here the development of a sensitive and convenient electronic sensor for the detection of antibodies in human serums. The sensor is constructed by self-assembly formation of a mixed monolayer containing the small molecule epitope conjugated double stranded DNA probes on gold electrode. The target antibody binds the epitope on the dsDNA probe and lowers the melting temperature of the duplex, which facilitates the displacement of the antibody-linked strand of the duplex probe by an invading methylene blue-tagged single stranded DNA (MB-ssDNA) through the strand displacement reaction and leads to the capture of many MB-ssDNA on the sensor surface. Subsequent electrochemical oxidation of the methylene blue labels results in amplified current response for sensitive monitoring of the antibodies. The antibody assay conditions are optimized and the sensor exhibits a linear range between 1.0 and 25.0nM with a detection limit of 0.67nM for the target antibody. The sensor is also selective and can be employed to detect the target antibodies in human serum samples. With the advantages of using small molecule epitope as the antibody recognition element over traditional antigen, the versatile manipulability of the DNA probes and the unique properties of the electrochemical transduction technique, the developed sensor thus hold great potential for simple and sensitive detection of different antibodies and other proteins in real samples. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Non-human biota dose assessment. Sensitivity analysis and knowledge quality assessment

    International Nuclear Information System (INIS)

    Smith, K.; Robinson, C.; Jackson, D.; La Cruz, I. de; Zinger, I.; Avila, R.

    2010-10-01

    This report provides a summary of a programme of work, commissioned within the BIOPROTA collaborative forum, to assess the quantitative and qualitative elements of uncertainty associated with biota dose assessment of potential impacts of long-term releases from geological disposal facilities (GDF). Quantitative and qualitative aspects of uncertainty were determined through sensitivity and knowledge quality assessments, respectively. Both assessments focused on default assessment parameters within the ERICA assessment approach. The sensitivity analysis was conducted within the EIKOS sensitivity analysis software tool and was run in both generic and test case modes. The knowledge quality assessment involved development of a questionnaire around the ERICA assessment approach, which was distributed to a range of experts in the fields of non-human biota dose assessment and radioactive waste disposal assessments. Combined, these assessments enabled critical model features and parameters that are both sensitive (i.e. have a large influence on model output) and of low knowledge quality to be identified for each of the three test cases. The output of this project is intended to provide information on those parameters that may need to be considered in more detail for prospective site-specific biota dose assessments for GDFs. Such information should help users to enhance the quality of their assessments and build greater confidence in the results. (orig.)

  15. Lansoprazole induces sensitivity to suboptimal doses of paclitaxel in human melanoma.

    Science.gov (United States)

    Azzarito, Tommaso; Venturi, Giulietta; Cesolini, Albino; Fais, Stefano

    2015-01-28

    Tumor acidity is now considered an important determinant of drug-resistance and tumor progression, and anti-acidic approaches, such as Proton Pump inhibitors (PPIs), have demonstrated promising antitumor and chemo-sensitizing efficacy. The main purpose of the present study was to evaluate the possible PPI-induced sensitization of human melanoma cells to Paclitaxel (PTX). Our results show that PTX and the PPI Lansoprazole (LAN) combination was extremely efficient against metastatic melanoma cells, as compared to the single treatments, both in vitro and in vivo. We also showed that acidity plays an important role on the anti-tumor activity of these drugs, being detrimental for PTX activity, while crucial for the synergistic effect of PTX following pretreatment with LAN, due to its nature of pro-drug needing protonation for a full activation. We obtained straightforward results in a human melanoma xenograft model combining well tolerated LAN doses with suboptimal and poorly toxic doses of PTX. With this study we provide a clear evidence that the PPI LAN may be included in new combined therapy of human melanoma together with low doses of PTX. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  16. A UV-sensitive human clonal cell line, RSa, which has low repair activity

    International Nuclear Information System (INIS)

    Suzuki, N.; Fuse, A.

    1981-01-01

    The repair activity of a human transformed cell line, RSa, which was found to be highly sensitive to the lethal effects of 254 mm far-ultraviolet radiation, was compared with that of HeLa cells by evaluating the range of UV-induced incorporation of [methyl- 3 H]thymidine ([ 3 H]dThd) or 5-[6- 3 H]bromodeoxyuridine ([ 3 H]BrdUrd) into deoxyribonucleic acid. Direct scintillation counting was used for measuring the extent of unscheduled DNA synthesis (UDS) in UV-irradiated cells, which were treated with hydroxyurea or with arginine deprivation. More quantitative measurements were made by using the density labeling and equilibrium centrifugation method for assaying repair replication. All the amounts of UDS and repair replication in RSa cells were markedly below those in HeLa cells. The possible relationships of the low repair activity to abnormally high UV sensitivity in RSa cells are discussed. (orig.)

  17. Sensitivity of hyperthermia-treated human cells to killing by ultraviolet or gamma radiation

    International Nuclear Information System (INIS)

    Mitchel, R.E.; Smith, B.P.; Wheatly, N.; Chan, A.; Child, S.; Paterson, M.C.

    1985-01-01

    Human xeroderma pigmentosum (XP) or Fanconi anemia (FA) fibroblasts displayed shouldered 45 0 C heat survival curves not significantly different from normal fibroblasts, a result similar to that previously found for ataxia telangiectasia (AT) cells, indicating heat resistance is not linked to either uv or low-LET ionizing radiation resistance. Hyperthermia (45 0 C) sensitized normal and XP fibroblasts to killing by gamma radiation but failed to sensitize the cells to the lethal effects of 254 nm uv radiation. Thermal inhibition of repair of ionizing radiation lesions but not uv-induced lesions appears to contribute synergistically to cell death. The thermal enhancement ratio (TER) for the synergistic interaction of hyperthermia (45 0 C, 30 min) and gamma radiation was significantly lower in one FA and two strains (TER = 1.7-1.8) than that reported previously for three normal strains (TER = 2.5-3.0). These XP and FA strains may be more gamma sensitive than normal human fibroblasts. Since hyperthermia treatment only slightly increases the gamma-radiation sensitivity of ataxia telangiectasia (AT) fibroblasts compared to normal strains, it is possible that the degree of thermal enhancement attainable reflects the genetically inherent ionizing radiation repair capacity of the cells. The data indicate that both repair inhibition and particular lesion types are required for lethal synergism between heat and radiation. We therefore postulate that the transient thermal inhibition of repair results in the conversion of gamma-induced lesions to irrepairable lethal damage, while uv-type damage can remain unaltered during this period

  18. Using citizen science data to identify the sensitivity of species to human land use.

    Science.gov (United States)

    Todd, Brian D; Rose, Jonathan P; Price, Steven J; Dorcas, Michael E

    2016-12-01

    Conservation practitioners must contend with an increasing array of threats that affect biodiversity. Citizen scientists can provide timely and expansive information for addressing these threats across large scales, but their data may contain sampling biases. We used randomization procedures to account for possible sampling biases in opportunistically reported citizen science data to identify species' sensitivities to human land use. We analyzed 21,044 records of 143 native reptile and amphibian species reported to the Carolina Herp Atlas from North Carolina and South Carolina between 1 January 1990 and 12 July 2014. Sensitive species significantly associated with natural landscapes were 3.4 times more likely to be legally protected or treated as of conservation concern by state resource agencies than less sensitive species significantly associated with human-dominated landscapes. Many of the species significantly associated with natural landscapes occurred primarily in habitats that had been nearly eradicated or otherwise altered in the Carolinas, including isolated wetlands, longleaf pine savannas, and Appalachian forests. Rare species with few reports were more likely to be associated with natural landscapes and 3.2 times more likely to be legally protected or treated as of conservation concern than species with at least 20 reported occurrences. Our results suggest that opportunistically reported citizen science data can be used to identify sensitive species and that species currently restricted primarily to natural landscapes are likely at greatest risk of decline from future losses of natural habitat. Our approach demonstrates the usefulness of citizen science data in prioritizing conservation and in helping practitioners address species declines and extinctions at large extents. © 2016 Society for Conservation Biology.

  19. Amphetamine sensitization alters reward processing in the human striatum and amygdala.

    Directory of Open Access Journals (Sweden)

    Owen G O'Daly

    Full Text Available Dysregulation of mesolimbic dopamine transmission is implicated in a number of psychiatric illnesses characterised by disruption of reward processing and goal-directed behaviour, including schizophrenia, drug addiction and impulse control disorders associated with chronic use of dopamine agonists. Amphetamine sensitization (AS has been proposed to model the development of this aberrant dopamine signalling and the subsequent dysregulation of incentive motivational processes. However, in humans the effects of AS on the dopamine-sensitive neural circuitry associated with reward processing remains unclear. Here we describe the effects of acute amphetamine administration, following a sensitising dosage regime, on blood oxygen level dependent (BOLD signal in dopaminoceptive brain regions during a rewarded gambling task performed by healthy volunteers. Using a randomised, double-blind, parallel-groups design, we found clear evidence for sensitization to the subjective effects of the drug, while rewarded reaction times were unchanged. Repeated amphetamine exposure was associated with reduced dorsal striatal BOLD signal during decision making, but enhanced ventromedial caudate activity during reward anticipation. The amygdala BOLD response to reward outcomes was blunted following repeated amphetamine exposure. Positive correlations between subjective sensitization and changes in anticipation- and outcome-related BOLD signal were seen for the caudate nucleus and amygdala, respectively. These data show for the first time in humans that AS changes the functional impact of acute stimulant exposure on the processing of reward-related information within dopaminoceptive regions. Our findings accord with pathophysiological models which implicate aberrant dopaminergic modulation of striatal and amygdala activity in psychosis and drug-related compulsive disorders.

  20. Phenylbutyrate Sensitizes Human Glioblastoma Cells Lacking Wild-Type P53 Function to Ionizing Radiation

    International Nuclear Information System (INIS)

    Lopez, Carlos A.; Feng, Felix Y.; Herman, Joseph M.; Nyati, Mukesh K.; Lawrence, Theodore S.; Ljungman, Mats

    2007-01-01

    Purpose: Histone deacetylase (HDAC) inhibitors induce growth arrest, differentiation, and apoptosis in cancer cells. Phenylbutyrate (PB) is a HDAC inhibitor used clinically for treatment of urea cycle disorders. Because of its low cytotoxicity, cerebrospinal fluid penetration, and high oral bioavailability, we investigated PB as a potential radiation sensitizer in human glioblastoma cell lines. Methods and Materials: Four glioblastoma cell lines were selected for this study. Phenylbutyrate was used at a concentration of 2 mM, which is achievable in humans. Western blots were used to assess levels of acetylated histone H3 in tumor cells after treatment with PB. Flow cytometry was used for cell cycle analysis. Clonogenic assays were performed to assess the effect of PB on radiation sensitivity. We used shRNA against p53 to study the role of p53 in radiosensitization. Results: Treatment with PB alone resulted in hyperacetylation of histones, confirmed by Western blot analysis. The PB alone resulted in cytostatic effects in three cell lines. There was no evidence of G 1 arrest, increase in sub-G 1 fraction or p21 protein induction. Clonogenic assays showed radiosensitization in two lines harboring p53 mutations, with enhancement ratios (± SE) of 1.5 (± 0.2) and 1.3 (± 0.1), respectively. There was no radiopotentiating effect in two cell lines with wild-type p53, but knockdown of wild-type p53 resulted in radiosensitization by PB. Conclusions: Phenylbutyrate can produce p21-independent cytostasis, and enhances radiation sensitivity in p53 mutant human glioblastoma cells in vitro. This suggests the potential application of combined PB and radiotherapy in glioblastoma harboring mutant p53

  1. MicroRNA-203 Modulates the Radiation Sensitivity of Human Malignant Glioma Cells

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Ji Hyun [Department of Radiation Oncology, Graduate School of Medicine, Seoul National University, Seoul (Korea, Republic of); Hwang, Yeo Hyun; Lee, David J.; Kim, Dan Hyo; Park, Ji Min [Medical Science Research Institute, Seoul National University Bundang Hospital, Kyeonggido (Korea, Republic of); Wu, Hong-Gyun [Department of Radiation Oncology, Graduate School of Medicine, Seoul National University, Seoul (Korea, Republic of); Kim, In Ah, E-mail: inah228@snu.ac.kr [Department of Radiation Oncology, Graduate School of Medicine, Seoul National University, Seoul (Korea, Republic of); Medical Science Research Institute, Seoul National University Bundang Hospital, Kyeonggido (Korea, Republic of); Cancer Research Institute, Seoul National University, Seoul (Korea, Republic of)

    2016-02-01

    Purpose: We investigated whether miR-203 could modulate the radiation sensitivity of glioblastoma (GBM) cells and which target gene(s) could be involved. Methods and Materials: Three human malignant glioma (MG) cell lines and normal human astrocytes were transfected with control microRNA, pre-miR-203, or antisense miR-203. Real-time PCR (RT-PCR), clonogenic assays, immunofluorescence, and invasion/migration assays were performed. To predict the target(s), bioinformatics analyses using microRNA target databases were performed. Results: Overexpression of miR-203 increased the radiation sensitivity of all 3 human MG cell lines and prolonged radiation-induced γ-H2AX foci formation. Bioinformatics analyses suggested that miR-203 could be involved in post-transcriptional control of DNA repair, PI3K/AKT, SRC, and JAK/STAT3 and the vascular signaling pathway. Western blot analysis validated the fact that miR-203 downregulated ATM, RAD51, SRC, PLD2, PI3K-AKT, JAK-STAT3, VEGF, HIF-1α, and MMP2. Overexpression of miR-203 inhibited invasion and migration potentials, downregulated SLUG and Vimentin, and upregulated Claudin-1 and ZO1. Conclusions: These data demonstrate that miR-203 potentially controls DNA damage repair via the PI3K/AKT and JAK/STAT3 pathways and may collectively contribute to the modulation of radiation sensitivity in MG cells by inhibiting DNA damage repair, prosurvival signaling, and epithelium-mesenchyme transition. Taken together, these findings demonstrate that miR-203 could be a target for overcoming the radiation resistance of GBM.

  2. MicroRNA-203 Modulates the Radiation Sensitivity of Human Malignant Glioma Cells

    International Nuclear Information System (INIS)

    Chang, Ji Hyun; Hwang, Yeo Hyun; Lee, David J.; Kim, Dan Hyo; Park, Ji Min; Wu, Hong-Gyun; Kim, In Ah

    2016-01-01

    Purpose: We investigated whether miR-203 could modulate the radiation sensitivity of glioblastoma (GBM) cells and which target gene(s) could be involved. Methods and Materials: Three human malignant glioma (MG) cell lines and normal human astrocytes were transfected with control microRNA, pre-miR-203, or antisense miR-203. Real-time PCR (RT-PCR), clonogenic assays, immunofluorescence, and invasion/migration assays were performed. To predict the target(s), bioinformatics analyses using microRNA target databases were performed. Results: Overexpression of miR-203 increased the radiation sensitivity of all 3 human MG cell lines and prolonged radiation-induced γ-H2AX foci formation. Bioinformatics analyses suggested that miR-203 could be involved in post-transcriptional control of DNA repair, PI3K/AKT, SRC, and JAK/STAT3 and the vascular signaling pathway. Western blot analysis validated the fact that miR-203 downregulated ATM, RAD51, SRC, PLD2, PI3K-AKT, JAK-STAT3, VEGF, HIF-1α, and MMP2. Overexpression of miR-203 inhibited invasion and migration potentials, downregulated SLUG and Vimentin, and upregulated Claudin-1 and ZO1. Conclusions: These data demonstrate that miR-203 potentially controls DNA damage repair via the PI3K/AKT and JAK/STAT3 pathways and may collectively contribute to the modulation of radiation sensitivity in MG cells by inhibiting DNA damage repair, prosurvival signaling, and epithelium-mesenchyme transition. Taken together, these findings demonstrate that miR-203 could be a target for overcoming the radiation resistance of GBM.

  3. O6-alkylguanine-DNA-alkyltransferase activity and nitrosourea sensitivity in human cancer cell lines.

    OpenAIRE

    Walker, M. C.; Masters, J. R.; Margison, G. P.

    1992-01-01

    The DNA repair enzyme, O6-alkylguanine-DNA-alkyltransferase (ATase), is thought to be the principal mechanism controlling resistance to nitrosoureas and related alkylating agents. We compared the sensitivities of five human testis and five bladder tumour cell lines to two nitrosoureas (N-nitroso-N-methylurea (MNU) and mitozolomide) with cellular levels of ATase. Enzyme levels ranged from 3 to 206 fmol mg-1 protein (0.1 x 10(4) to 5.1 x 10(4) molecules/cell) in the testis lines and from 11 to ...

  4. The Arctic Alzheimer mutation enhances sensitivity to toxic stress in human neuroblastoma cells

    DEFF Research Database (Denmark)

    Sennvik, Kristina; Nilsberth, Camilla; Stenh, Charlotte

    2002-01-01

    The E693G (Arctic) mutation of the amyloid precursor protein was recently found to lead to early-onset Alzheimer's disease in a Swedish family. In the present study, we report that the Arctic mutation decreases cell viability in human neuroblastoma cells. The cell viability, as measured by the MTT...... their secretion of beta-secretase cleaved amyloid precursor protein. The enhanced sensitivity to toxic stress in cells with the Arctic mutation most likely contributes to the pathogenic pathway leading to Alzheimer's disease....

  5. Differentiation-associated decrease in muscarinic receptor sensitivity in human neuroblastoma cells

    International Nuclear Information System (INIS)

    Heikkilae, J.E.; Scott, J.G.; Suominen, L.A.; Akerman, K.E.O.

    1987-01-01

    Muscarinic receptor-linked increases in intracellular free Ca 2+ as measured with quin-2 and Ca 2+ release from monolayers of cells have been measured in the human neuroblastoma cell line SH-SY5Y. Induction of differentiation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) leads to a decrease in the sensitivity of the cells to low concentrations of agonists with respect to the induced increase in cytosolic free Ca 2+ and stimulation of Ca 2+ efflux. No decrease in agonist binding affinity was observed when the displacement of a labelled antagonist, 3 H-NMS, by a non-labelled agonist was studied

  6. The human milk protein-lipid complex HAMLET sensitizes bacterial pathogens to traditional antimicrobial agents.

    Science.gov (United States)

    Marks, Laura R; Clementi, Emily A; Hakansson, Anders P

    2012-01-01

    The fight against antibiotic resistance is one of the most significant challenges to public health of our time. The inevitable development of resistance following the introduction of novel antibiotics has led to an urgent need for the development of new antibacterial drugs with new mechanisms of action that are not susceptible to existing resistance mechanisms. One such compound is HAMLET, a natural complex from human milk that kills Streptococcus pneumoniae (the pneumococcus) using a mechanism different from common antibiotics and is immune to resistance-development. In this study we show that sublethal concentrations of HAMLET potentiate the effect of common antibiotics (penicillins, macrolides, and aminoglycosides) against pneumococci. Using MIC assays and short-time killing assays we dramatically reduced the concentrations of antibiotics needed to kill pneumococci, especially for antibiotic-resistant strains that in the presence of HAMLET fell into the clinically sensitive range. Using a biofilm model in vitro and nasopharyngeal colonization in vivo, a combination of HAMLET and antibiotics completely eradicated both biofilms and colonization in mice of both antibiotic-sensitive and resistant strains, something each agent alone was unable to do. HAMLET-potentiation of antibiotics was partially due to increased accessibility of antibiotics to the bacteria, but relied more on calcium import and kinase activation, the same activation pathway HAMLET uses when killing pneumococci by itself. Finally, the sensitizing effect was not confined to species sensitive to HAMLET. The HAMLET-resistant respiratory species Acinetobacter baumanii and Moraxella catarrhalis were all sensitized to various classes of antibiotics in the presence of HAMLET, activating the same mechanism as in pneumococci. Combined these results suggest the presence of a conserved HAMLET-activated pathway that circumvents antibiotic resistance in bacteria. The ability to activate this pathway may extend

  7. Comparison of two different techniques on the human lymphocytes morphology and sensitivity to gamma radiation

    International Nuclear Information System (INIS)

    Kol, R.

    1985-02-01

    results that were obtained with regard to the sensitivity of human lymphocytes to radiation. (Author)

  8. Selenite Treatment Inhibits LAPC-4 Tumor Growth and Prostate-Specific Antigen Secretion in a Xenograft Model of Human Prostate Cancer

    International Nuclear Information System (INIS)

    Bhattacharyya, Rumi S.; Husbeck, Bryan; Feldman, David; Knox, Susan J.

    2008-01-01

    Purpose: Selenium compounds have known chemopreventive effects on prostate cancer. However selenite, an inorganic form of selenium, has not been extensively studied as a treatment option for prostate cancer. Our previous studies have demonstrated the inhibition of androgen receptor expression and androgen stimulated prostate-specific antigen (PSA) expression by selenite in human prostate cancer cell lines. In this study, we investigated the in vivo effects of selenite as a therapy to treat mice with established LAPC-4 tumors. Methods and Materials: Male mice harboring androgen-dependent LAPC-4 xenograft tumors were treated with selenite (2 mg/kg intraperitoneally three times per week) or vehicle for 42 days. In addition, androgen-independent LAPC-4 xenograft tumors were generated in female mice over 4 to 6 months. Once established, androgen-independent LAPC-4 tumor fragments were passaged into female mice and were treated with selenite or vehicle for 42 days. Changes in tumor volume and serum PSA levels were assessed. Results: Selenite significantly decreased androgen-dependent LAPC-4 tumor growth in male mice over 42 days (p < 0.001). Relative tumor volume was decreased by 41% in selenite-treated animals compared with vehicle-treated animals. The inhibition of LAPC-4 tumor growth corresponded to a marked decrease in serum PSA levels (p < 0.01). In the androgen-independent LAPC-4 tumors in female mice, selenite treatment decreased tumor volume by 58% after 42 days of treatment (p < 0.001). Conclusions: These results suggest that selenite may have potential as a novel therapeutic agent to treat both androgen-dependent and androgen-independent prostate cancer

  9. Decoding of faces and face components in face-sensitive human visual cortex

    Directory of Open Access Journals (Sweden)

    David F Nichols

    2010-07-01

    Full Text Available A great challenge to the field of visual neuroscience is to understand how faces are encoded and represented within the human brain. Here we show evidence from functional magnetic resonance imaging (fMRI for spatially distributed processing of the whole face and its components in face-sensitive human visual cortex. We used multi-class linear pattern classifiers constructed with a leave-one-scan-out verification procedure to discriminate brain activation patterns elicited by whole faces, the internal features alone, and the external head outline alone. Furthermore, our results suggest that whole faces are represented disproportionately in the fusiform cortex (FFA whereas the building blocks of faces are represented disproportionately in occipitotemporal cortex (OFA. Faces and face components may therefore be organized with functional clustering within both the FFA and OFA, but with specialization for face components in the OFA and the whole face in the FFA.

  10. Sensitive and Flexible Polymeric Strain Sensor for Accurate Human Motion Monitoring.

    Science.gov (United States)

    Khan, Hassan; Razmjou, Amir; Ebrahimi Warkiani, Majid; Kottapalli, Ajay; Asadnia, Mohsen

    2018-02-01

    Flexible electronic devices offer the capability to integrate and adapt with human body. These devices are mountable on surfaces with various shapes, which allow us to attach them to clothes or directly onto the body. This paper suggests a facile fabrication strategy via electrospinning to develop a stretchable, and sensitive poly (vinylidene fluoride) nanofibrous strain sensor for human motion monitoring. A complete characterization on the single PVDF nano fiber has been performed. The charge generated by PVDF electrospun strain sensor changes was employed as a parameter to control the finger motion of the robotic arm. As a proof of concept, we developed a smart glove with five sensors integrated into it to detect the fingers motion and transfer it to a robotic hand. Our results shows that the proposed strain sensors are able to detect tiny motion of fingers and successfully run the robotic hand.

  11. Localization and function of ATP-sensitive potassium channels in human skeletal muscle

    DEFF Research Database (Denmark)

    Nielsen, Jens Jung; Kristensen, Michael; Hellsten, Ylva

    2003-01-01

    The present study investigated the localization of ATP-sensitive K+ (KATP) channels in human skeletal muscle and the functional importance of these channels for human muscle K+ distribution at rest and during muscle activity. Membrane fractionation based on the giant vesicle technique...... or the sucrose-gradient technique in combination with Western blotting demonstrated that the KATP channels are mainly located in the sarcolemma. This localization was confirmed by immunohistochemical measurements. With the microdialysis technique, it was demonstrated that local application of the KATP channel...... to in vitro conditions, the present study demonstrated that under in vivo conditions the KATP channels are active at rest and contribute to the accumulation of interstitial K+....

  12. Sensitive and Flexible Polymeric Strain Sensor for Accurate Human Motion Monitoring

    Directory of Open Access Journals (Sweden)

    Hassan Khan

    2018-02-01

    Full Text Available Flexible electronic devices offer the capability to integrate and adapt with human body. These devices are mountable on surfaces with various shapes, which allow us to attach them to clothes or directly onto the body. This paper suggests a facile fabrication strategy via electrospinning to develop a stretchable, and sensitive poly (vinylidene fluoride nanofibrous strain sensor for human motion monitoring. A complete characterization on the single PVDF nano fiber has been performed. The charge generated by PVDF electrospun strain sensor changes was employed as a parameter to control the finger motion of the robotic arm. As a proof of concept, we developed a smart glove with five sensors integrated into it to detect the fingers motion and transfer it to a robotic hand. Our results shows that the proposed strain sensors are able to detect tiny motion of fingers and successfully run the robotic hand.

  13. UV-sensitive photoreceptor protein OPN5 in humans and mice.

    Science.gov (United States)

    Kojima, Daisuke; Mori, Suguru; Torii, Masaki; Wada, Akimori; Morishita, Rika; Fukada, Yoshitaka

    2011-01-01

    A variety of animal species utilize the ultraviolet (UV) component of sunlight as their environmental cues, whereas physiological roles of UV photoreception in mammals, especially in human beings, remain open questions. Here we report that mouse neuropsin (OPN5) encoded by the Opn5 gene exhibited an absorption maximum (λmax) at 380 nm when reconstituted with 11-cis-retinal. Upon UV-light illumination, OPN5 was converted to a blue-absorbing photoproduct (λmax 470 nm), which was stable in the dark and reverted to the UV-absorbing state by the subsequent orange light illumination, indicating its bistable nature. Human OPN5 also had an absorption maximum at 380 nm with spectral properties similar to mouse OPN5, revealing that OPN5 is the first and hitherto unknown human opsin with peak sensitivity in the UV region. OPN5 was capable of activating heterotrimeric G protein Gi in a UV-dependent manner. Immuno-blotting analyses of mouse tissue extracts identified the retina, the brain and, unexpectedly, the outer ears as the major sites of OPN5 expression. In the tissue sections of mice, OPN5 immuno-reactivities were detected in a subset of non-rod/non-cone retinal neurons as well as in the epidermal and muscle cells of the outer ears. Most of these OPN5-immuno-reactivities in mice were co-localized with positive signals for the alpha-subunit of Gi. These results demonstrate the first example of UV photoreceptor in human beings and strongly suggest that OPN5 triggers a UV-sensitive Gi-mediated signaling pathway in the mammalian tissues.

  14. Sensitive method for the determination of nitrated polycyclic aromatic hydrocarbons in the human diet

    Energy Technology Data Exchange (ETDEWEB)

    Siegmund, B.; Weiss, R.; Pfannhauser, W. [Graz Univ. of Technology (Austria). Inst. of Food Chemistry

    2003-01-01

    The presence of nitrated polycyclic aromatic hydrocarbons (nitro-PAHs) as contaminants in foods is nowadays a legitimate cause of concern, as they are reported to be strong direct-acting mutagens and carcinogens that may pose a risk to human health. Nevertheless, their concentrations in foods are in the ultra-trace region. In this study, we present a very sensitive analytical method for the determination of this compound class in complex food matrices. Special emphasis was put on sample extraction and clean up. Recoveries of 75% could be obtained for 1- and 2-nitronaphthalene; for all other compounds investigated the recoveries were {>=}94%. The determination of the compounds was performed by gas chromatography-mass spectrometry. The results show that the use of negative chemical ionization (NCI) increases the sensitivity by one order of magnitude in comparison to electron impact ionization (EI) (limits of detection 0.07-0.25 {mu}g kg{sup -1} for EI and 0.01-0.02 {mu}g kg{sup -1} for NCI). The influence of the sample matrix on the sensitivity of the method is demonstrated with extracts of a sample from a duplicate diet study. Due to the lack of a certified reference material the suitability of the method is demonstrated with pumpkin seed oil that was spiked with nitro-PAHs at two different concentration levels. (orig.)

  15. Dynamic modulation of thymidylate synthase gene expression and fluorouracil sensitivity in human colorectal cancer cells.

    Directory of Open Access Journals (Sweden)

    Kentaro Wakasa

    Full Text Available Biomarkers have revolutionized cancer chemotherapy. However, many biomarker candidates are still in debate. In addition to clinical studies, a priori experimental approaches are needed. Thymidylate synthase (TS expression is a long-standing candidate as a biomarker for 5-fluorouracil (5-FU treatment of cancer patients. Using the Tet-OFF system and a human colorectal cancer cell line, DLD-1, we first constructed an in vitro system in which TS expression is dynamically controllable. Quantitative assays have elucidated that TS expression in the transformant was widely modulated, and that the dynamic range covered 15-fold of the basal level. 5-FU sensitivity of the transformant cells significantly increased in response to downregulated TS expression, although being not examined in the full dynamic range because of the doxycycline toxicity. Intriguingly, our in vitro data suggest that there is a linear relationship between TS expression and the 5-FU sensitivity in cells. Data obtained in a mouse model using transformant xenografts were highly parallel to those obtained in vitro. Thus, our in vitro and in vivo observations suggest that TS expression is a determinant of 5-FU sensitivity in cells, at least in this specific genetic background, and, therefore, support the possibility of TS expression as a biomarker for 5-FU-based cancer chemotherapy.

  16. Hydrogen-bond network and pH sensitivity in human transthyretin

    Energy Technology Data Exchange (ETDEWEB)

    Yokoyama, Takeshi, E-mail: tyokoya3@pha.u-toyama.ac.jp; Mizuguchi, Mineyuki; Nabeshima, Yuko [University of Toyama, 2630 Sugitani, Toyama 930-0914 (Japan); Kusaka, Katsuhiro; Yamada, Taro [Ibaraki University, 162-1 Shirakata, Tokai, Ibaraki 319-1106 (Japan); Hosoya, Takaaki [Ibaraki University, 162-1 Shirakata, Tokai, Ibaraki 319-1106 (Japan); Ibaraki University, 4-12-1 Naka-Narusawa, Hitachi, Ibaraki 316-8511 (Japan); Ohhara, Takashi [Comprehensive Research Organization for Science and Society, 162-1 Shirakata, Tokai, Ibaraki 319-1106 (Japan); Kurihara, Kazuo [Japan Atomic Energy Agency, 2-4 Shirakata, Tokai, Ibaraki 319-1195 (Japan); Tanaka, Ichiro [Ibaraki University, 162-1 Shirakata, Tokai, Ibaraki 319-1106 (Japan); Ibaraki University, 4-12-1 Naka-Narusawa, Hitachi, Ibaraki 316-8511 (Japan); Niimura, Nobuo [Ibaraki University, 162-1 Shirakata, Tokai, Ibaraki 319-1106 (Japan)

    2013-11-01

    The neutron crystal structure of human transthyretin is presented. Transthyretin (TTR) is a tetrameric protein. TTR misfolding and aggregation are associated with human amyloid diseases. Dissociation of the TTR tetramer is believed to be the rate-limiting step in the amyloid fibril formation cascade. Low pH is known to promote dissociation into monomer and the formation of amyloid fibrils. In order to reveal the molecular mechanisms underlying pH sensitivity and structural stabilities of TTR, neutron diffraction studies were conducted using the IBARAKI Biological Crystal Diffractometer with the time-of-flight method. Crystals for the neutron diffraction experiments were grown up to 2.5 mm{sup 3} for four months. The neutron crystal structure solved at 2.0 Å revealed the protonation states of His88 and the detailed hydrogen-bond network depending on the protonation states of His88. This hydrogen-bond network is involved in monomer–monomer and dimer–dimer interactions, suggesting that the double protonation of His88 by acidification breaks the hydrogen-bond network and causes the destabilization of the TTR tetramer. Structural comparison with the X-ray crystal structure at acidic pH identified the three amino acid residues responsible for the pH sensitivity of TTR. Our neutron model provides insights into the molecular stability related to amyloidosis.

  17. Evaluation of cloned cells, animal model, and ATRA sensitivity of human testicular yolk sac tumor

    Directory of Open Access Journals (Sweden)

    Zhao Junfeng

    2012-03-01

    Full Text Available Abstract The testicular yolk sac tumor (TYST is the most common neoplasm originated from germ cells differentiated abnormally, a major part of pediatric malignant testicular tumors. The present study aimed at developing and validating the in vitro and vivo models of TYST and evaluating the sensitivity of TYST to treatments, by cloning human TYST cells and investigating the histology, ultra-structure, growth kinetics and expression of specific proteins of cloned cells. We found biological characteristics of cloned TYST cells were similar to the yolk sac tumor and differentiated from the columnar to glandular-like or goblet cells-like cells. Chromosomes for tumor identification in each passage met nature of the primary tumor. TYST cells were more sensitive to all-trans-retinoic acid which had significantly inhibitory effects on cell proliferation. Cisplatin induced apoptosis of TYST cells through the activation of p53 expression and down-regulation of Bcl- expression. Thus, we believe that cloned TYST cells and the animal model developed here are useful to understand the molecular mechanism of TYST cells and develop potential therapies for human TYST.

  18. Sensitivity to ionizing radiation and chemotherapeutic agents in gemcitabine-resistant human tumor cell lines

    International Nuclear Information System (INIS)

    Bree, Chris van; Kreder, Natasja Castro; Loves, Willem J.P.; Franken, Nicolaas A.P.; Peters, Godefridus J.; Haveman, Jaap

    2002-01-01

    Purpose: To determine cross-resistance to anti-tumor treatments in 2',2'difluorodeoxycytidine (dFdC, gemcitabine)-resistant human tumor cells. Methods and Materials: Human lung carcinoma cells SW-1573 (SWp) were made resistant to dFdC (SWg). Sensitivity to cisplatin (cDDP), paclitaxel, 5-fluorouracil (5-FU), methotrexate (MTX), cytarabine (ara-C), and dFdC was measured by a proliferation assay. Radiosensitivity and radioenhancement by dFdC of this cell panel and the human ovarian carcinoma cell line A2780 and its dFdC-resistant variant AG6000 were determined by clonogenic assay. Bivariate flowcytometry was performed to study cell cycle changes. Results: In the SWg, a complete deoxycytidine kinase (dCK) deficiency was found on mRNA and protein level. This was accompanied by a 10-fold decrease in dCK activity which resulted in the >1000-fold resistance to dFdC. Sensitivity to other anti-tumor drugs was not altered, except for ara-C (>100-fold resistance). Radiosensitivity was not altered in the dFdC-resistant cell lines SWg and AG6000. High concentrations (50-100 μM dFdC) induced radioenhancement in the dFdC-resistant cell lines similar to the radioenhancement obtained at lower concentrations (10 nM dFdC) in the parental lines. An early S-phase arrest was found in all cell lines after dFdC treatment where radioenhancement was achieved. Conclusions: In the dFdC-resistant lung tumor cell line SWg, the deficiency in dCK is related to the resistance to dFdC and ara-C. No cross-resistance was observed to other anti-tumor drugs used for the treatment in lung cancer. Sensitivity to ionizing radiation was not altered in two different dFdC-resistant cell lines. Resistance to dFdC does not eliminate the ability of dFdC to sensitize cells to radiation

  19. Development of nanobody-based flow injection chemiluminescence immunoassay for sensitive detection of human prealbumin.

    Science.gov (United States)

    Ma, Lei; Sun, Yanyan; Kang, Xuejun; Wan, Yakun

    2014-11-15

    Nanobodies, derived from camelid heavy-chain antibodies, have novel and impactful applications in clinical diagnostics. Our objective is to develop a nanobody-based chemiluminescence immunoassay for sensitive detection of human prealbumin (PA). In this context, a phage display nanobody library is constructed via immunizing dromedary camel with human prealbumin. Three nanobodies have been identified by five successive bio-panning steps. Based on their high expression level and good affinity, two out of three are chosen for further study. Magnetic beads (MBs) were functionalized with PEI by acylamide bond formed between the carboxyl group on the surface of the MB. Then, an anti-PA nanobody (Nb1) can be effectively immobilized onto the surface of the functionalized MB using glutaradehyde as the link. The modified MBs with Nb1 can specifically capture the target PA and reacted with silica nanoparticles with co-immobilized HRP and anti-PA nanobody (Nb2). The concentration of PA was detected by flow injection chemiluminescence. When using MB/PEI as the carrier of anti-PA Nb1, the CL signal significantly increased to 4-fold compared with the signal using MB without PEI modification. The CL signal was further amplified to 5-fold when Si/Nb2 was used as the signal probe. Under optimized conditions, the present immunoassay exhibited a wide quantitative range from 0.05 to 1000 μg L(-1) with a detection limit of 0.01 μg L(-1). The sensitivity of the proposed immunoassay offers great promises in providing a sensitive, specific, time saving, and potential method for detecting PA in clinical settings. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Dietary Sodium Restriction Decreases Insulin Secretion Without Affecting Insulin Sensitivity in Humans

    Science.gov (United States)

    Byrne, Loretta M.; Yu, Chang; Wang, Thomas J.; Brown, Nancy J.

    2014-01-01

    Context: Interruption of the renin-angiotensin-aldosterone system prevents incident diabetes in high-risk individuals, although the mechanism remains unclear. Objective: To test the hypothesis that activation of the endogenous renin-angiotensin-aldosterone system or exogenous aldosterone impairs insulin secretion in humans. Design: We conducted a randomized, blinded crossover study of aldosterone vs vehicle and compared the effects of a low-sodium versus a high-sodium diet. Setting: Academic clinical research center. Participants: Healthy, nondiabetic, normotensive volunteers. Interventions: Infusion of exogenous aldosterone (0.7 μg/kg/h for 12.5 h) or vehicle during low or high sodium intake. Low sodium (20 mmol/d; n = 12) vs high sodium (160 mmol/d; n = 17) intake for 5–7 days. Main Outcome Measures: Change in acute insulin secretory response assessed during hyperglycemic clamps while in sodium balance during a low-sodium vs high-sodium diet during aldosterone vs vehicle. Results: A low-sodium diet increased endogenous aldosterone and plasma renin activity, and acute glucose-stimulated insulin (−16.0 ± 5.6%; P = .007) and C-peptide responses (−21.8 ± 8.4%; P = .014) were decreased, whereas the insulin sensitivity index was unchanged (−1.0 ± 10.7%; P = .98). Aldosterone infusion did not affect the acute insulin response (+1.8 ± 4.8%; P = .72) or insulin sensitivity index (+2.0 ± 8.8%; P = .78). Systolic blood pressure and serum potassium were similar during low and high sodium intake and during aldosterone infusion. Conclusions: Low dietary sodium intake reduces insulin secretion in humans, independent of insulin sensitivity. PMID:25029426

  1. Functional Study of the P32T ITPA Variant Associated with Drug Sensitivity in Humans

    Science.gov (United States)

    Stepchenkova, Elena I.; Tarakhovskaya, Elena R.; Spitler, Kathryn; Frahm, Christin; Menezes, Miriam R.; Simone, Peter D.; Kolar, Carol; Marky, Luis A.; Borgstahl, Gloria E. O.; Pavlov, Youri I.

    2009-01-01

    Sanitization of the cellular nucleotide pools from mutagenic base analogs is necessary for the accuracy of transcription and replication of genetic material and plays a substantial role in cancer prevention. The undesirable mutagenic, recombinogenic and toxic incorporation of purine base analogs (i.e. ITP, dITP, XTP, dXTP or 6-hydroxyaminopurine (HAP) deoxynucleoside triphosphate) into nucleic acids is prevented by inosine triphosphate pyrophosphatase (ITPA). The ITPA gene is a highly conserved, moderately expressed gene. Defects in ITPA orthologs in model organisms cause severe sensitivity to HAP and chromosome fragmentation. A human polymorphic allele 94C->A encodes for the enzyme with a P32T amino acid change and leads to accumulation of non-hydrolyzed ITP. ITPase activity is not detected in erythrocytes of these patients. The P32T polymorphism has also been associated with adverse sensitivity to purine base analog drugs. We have found that the ITPA-P32T mutant is a dimer in solution, as is wild-type ITPA, and has normal ITPA activity in vitro, but the melting point of ITPA-P32T is 5 degrees C lower than that of wild-type. ITPA-P32T is also fully functional in vivo in model organisms as determined by a HAP mutagenesis assay and its complementation of a bacterial ITPA defect. The amount of ITPA protein detected by western blot is severely diminished in a human fibroblast cell line with the 94C->A change. We propose that the P32T mutation exerts its effect in certain human tissues by cumulative effects of destabilization of transcripts, protein stability and availability. PMID:19631656

  2. Ultra-Sensitive HIV-1 Latency Viral Outgrowth Assays Using Humanized Mice.

    Science.gov (United States)

    Schmitt, Kimberly; Akkina, Ramesh

    2018-01-01

    In the current quest for a complete cure for HIV/AIDS, highly sensitive HIV-1 latency detection methods are critical to verify full viral eradication. Until now, the in vitro quantitative viral outgrowth assays (qVOA) have been the gold standard for assessing latent HIV-1 viral burden. However, these assays have been inadequate in detecting the presence of ultralow levels of latent virus in a number of patients who were initially thought to have been cured, but eventually showed viral rebound. In this context, new approaches utilizing in vivo mouse-based VOAs are promising. In the murine VOA (mVOA), large numbers of CD4 + T cells or PBMC from aviremic subjects are xenografted into immunodeficient NSG mice, whereas in the humanized mouse-based VOA (hmVOA) patient CD4 + T cell samples are injected into BLT or hu-hematopoetic stem cells (hu-HSC) humanized mice. While latent virus could be recovered in both of these systems, the hmVOA provides higher sensitivity than the mVOA using a fewer number of input cells. In contrast to the mVOA, the hmVOA provides a broader spectrum of highly susceptible HIV-1 target cells and enables newly engrafted cells to home into preformed human lymphoid organs where they can infect cells in situ after viral activation. Hu-mice also allow for both xenograft- and allograft-driven cell expansions with less severe GvH providing a longer time frame for potential viral outgrowth from cells with a delayed latent viral activation. Based on these advantages, the hmVOA has great potential in playing an important role in HIV-1 latency and cure research.

  3. Do dogs (Canis lupus familiaris) make counterproductive choices because they are sensitive to human ostensive cues?

    Science.gov (United States)

    Marshall-Pescini, Sarah; Passalacqua, Chiara; Miletto Petrazzini, Maria Elena; Valsecchi, Paola; Prato-Previde, Emanuela

    2012-01-01

    Dogs appear to be sensitive to human ostensive communicative cues in a variety of situations, however there is still a measure of controversy as to the way in which these cues influence human-dog interactions. There is evidence for instance that dogs can be led into making evaluation errors in a quantity discrimination task, for example losing their preference for a larger food quantity if a human shows a preference for a smaller one, yet there is, so far, no explanation for this phenomenon. Using a modified version of this task, in the current study we investigated whether non-social, social or communicative cues (alone or in combination) cause dogs to go against their preference for the larger food quantity. Results show that dogs' evaluation errors are indeed caused by a social bias, but, somewhat contrary to previous studies, they highlight the potent effect of stimulus enhancement (handling the target) in influencing the dogs' response. A mild influence on the dog's behaviour was found only when different ostensive cues (and no handling of the target) were used in combination, suggesting their cumulative effect. The discussion addresses possible motives for discrepancies with previous studies suggesting that both the intentionality and the directionality of the action may be important in causing dogs' social biases.

  4. Protein phosphorylation in isolated human adipocytes - Adrenergic control of the phosphorylation of hormone-sensitive lipase

    International Nuclear Information System (INIS)

    Smiley, R.M.; Paul, S.; Browning, M.D.; Leibel, R.L.; Hirsch, J.

    1990-01-01

    The effect of adrenergic agents on protein phosphorylation in human adipocytes was examined. Freshly isolated human fat cells were incubated with 32 PO 4 in order to label intracellular ATP, then treated with a variety of adrenergic and other pharmacologic agents. Treatment with the β-adrenergic agonist isoproterenol led to a significant increase in phosphate content of at least five protein bands (M r 52, 53, 63, 67, 84 kDa). The increase in phosphorylation was partially inhibited by the α-2 agonist clonidine. Epinephrine, a combined α and β agonist, was less effective at increasing phosphate content of the proteins than was isoproterenol. Neither insulin nor the α-1 agonist phenylephrine had any discernible effect on the pattern of protein phosphorylation. The 84 kDa phosphorylated peptide band appears to contain hormone-sensitive lipase, a key enzyme in the lipolytic pathway which is activated by phosphorylation. These results are somewhat different than previously reported results for rat adipocytes, and represent the first report of overall pattern and adrenergic modulation of protein phosphorylation in human adipocytes

  5. Sensitive liquid chromatography-tandem mass spectrometry method for quantification of hydrochlorothiazide in human plasma.

    Science.gov (United States)

    Ramakrishna, N V S; Vishwottam, K N; Manoj, S; Koteshwara, M; Wishu, S; Varma, D P

    2005-12-01

    A simple, rapid, sensitive and specific liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of hydrochlorothiazide (I), a common diuretic and anti-hypertensive agent. The analyte and internal standard, tamsulosin (II) were extracted by liquid-liquid extraction with diethyl ether-dichloromethane (70:30, v/v) using a Glas-Col Multi-Pulse Vortexer. The chromatographic separation was performed on a reversed-phase column (Waters symmetry C18) with a mobile phase of 10 mm ammonium acetate-methanol (15:85, v/v). The protonated analyte was quantitated in negative ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 296.1 solidus in circle 205.0 and m/z 407.2 solidus in circle 184.9 were used to measure I and II, respectively. The assay exhibited a linear dynamic range of 0.5-200 ng/mL for hydrochlorothiazide in human plasma. The lower limit of quantitation was 500 pg/mL, with a relative standard deviation of less than 9%. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. A run time of 2.5 min for each sample made it possible to analyze a throughput of more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. (c) 2005 John Wiley & Sons, Ltd.

  6. Acquisition of anoikis resistance in human osteosarcoma cells does not alter sensitivity to chemotherapeutic agents

    International Nuclear Information System (INIS)

    Díaz-Montero, C Marcela; McIntyre, Bradley W

    2005-01-01

    Chemotherapy-induced cell death can involve the induction of apoptosis. Thus, aberrant function of the pathways involved might result in chemoresistance. Since cell adhesion to the extracellular matrix acts as a survival factor that homeostatically maintains normal tissue architecture, it was tested whether acquisition of resistance to deadhesion-induced apoptosis (anoikis) in human osteosarcoma would result in resistance to chemotherapy. Osteosarcoma cell lines (SAOS-2 and TE-85) obtained from ATCC and were maintained in complete Eagle's MEM medium. Suspension culture was established by placing cells in tissue culture wells coated with poly-HEMA. Cell cytotoxicity was determined using a live/dead cytotoxicity assay. Cell cycle/apoptosis analyses were performed using propidium iodide (PI) staining with subsequent FACS analysis. Apoptosis was also assayed by Annexin-FITC/PI staining. Etoposide, adriamycin, vinblastine, cisplatin and paclitaxel were able to induce apoptosis in human osteosarcoma cells SAOS-2 regardless of their anoikis resistance phenotype or the culture conditions (adhered vs. suspended). Moreover, suspended anoikis resistant TE-85 cells (TE-85ar) retained their sensitivity to chemotherapy as well. Acquisition of anoikis resistance in human osteosarcoma cells does not result in a generalized resistance to all apoptotic stimuli, including chemotherapy. Moreover, our results suggest that the pathways regulating anoikis resistance and chemotherapy resistance might involve the action of different mediators

  7. Acquisition of anoikis resistance in human osteosarcoma cells does not alter sensitivity to chemotherapeutic agents

    Directory of Open Access Journals (Sweden)

    McIntyre Bradley W

    2005-04-01

    Full Text Available Abstract Background Chemotherapy-induced cell death can involve the induction of apoptosis. Thus, aberrant function of the pathways involved might result in chemoresistance. Since cell adhesion to the extracellular matrix acts as a survival factor that homeostatically maintains normal tissue architecture, it was tested whether acquisition of resistance to deadhesion-induced apoptosis (anoikis in human osteosarcoma would result in resistance to chemotherapy. Methods Osteosarcoma cell lines (SAOS-2 and TE-85 obtained from ATCC and were maintained in complete Eagle's MEM medium. Suspension culture was established by placing cells in tissue culture wells coated with poly-HEMA. Cell cytotoxicity was determined using a live/dead cytotoxicity assay. Cell cycle/apoptosis analyses were performed using propidium iodide (PI staining with subsequent FACS analysis. Apoptosis was also assayed by Annexin-FITC/PI staining. Results Etoposide, adriamycin, vinblastine, cisplatin and paclitaxel were able to induce apoptosis in human osteosarcoma cells SAOS-2 regardless of their anoikis resistance phenotype or the culture conditions (adhered vs. suspended. Moreover, suspended anoikis resistant TE-85 cells (TE-85ar retained their sensitivity to chemotherapy as well. Conclusion Acquisition of anoikis resistance in human osteosarcoma cells does not result in a generalized resistance to all apoptotic stimuli, including chemotherapy. Moreover, our results suggest that the pathways regulating anoikis resistance and chemotherapy resistance might involve the action of different mediators.

  8. Radiation sensitization by dihydroartemisinin on human HeLa cells of cervical cancer

    International Nuclear Information System (INIS)

    Chen Xialin; Cao Jianping; Ji Rong; Zhu Wei; Liu Yang; Gong Xiaomei; Tang Yan; Pan Chunyan; Fan Saijun

    2009-01-01

    Objective: To investigate the radiosensitizing effects of dihydroartemisinin (DHA) on human HeLa cells of cervical cancer irradiated by X rays. Methods: Cell growth kinetics was determined using MTF assay. Cell survival was analyzed by elonogenic assay. The change of cell cycle and apeptosis was measured by flow cytometry. Results: Dihydroartemisinin inhibited the growth of HeLa cells of human cervical cancer and showed a dose-dependent and time-dependent manner. Dihydroartemisinin (20 μmol/L) showed the radiosensitizing effects on HeLa cells, and the sensitizing enhancement ratio (SER) was 1.47. Dihydroartemisinin abrogated radiation-induced G 2 arrest of the tested HeLa cells, the G 2 ratio of medicine + radiation group dechned from 73.58% to 48.31%. Dihydroartemisinin enhanced the apoptosis of HeLa cells by X-irradiation, the apoptosis rates of medicine + radiation group significantly increased from 29.46%, 48.04%, 70.21% to 45.79%, 66.36% and 79.58%, respectively for 2, 4 and 6 Gy. Conclusions: Dihydroartemisinin could increase the radiosensitivity of HeLa cells of human cervical cancer. Abrogation of radiation-induced C 2 arrest could be part of the mechanism. (authors)

  9. Highly sensitive electrochemical detection of human telomerase activity based on bio-barcode method.

    Science.gov (United States)

    Li, Ying; Liu, Bangwei; Li, Xia; Wei, Qingli

    2010-07-15

    In the present study, an electrochemical method for highly sensitive detection of human telomerase activity was developed based on bio-barcode amplification assay. Telomerase was extracted from HeLa cells, then the extract was mixed with telomerase substrate (TS) primer to perform extension reaction. The extension product was hybridized with the capture DNA immobilized on the Au electrode and then reacted with the signal DNA on Au nanoparticles to form a sandwich hybridization mode. Electrochemical signals were generated by chronocoulometric interrogation of [Ru(NH(3))(6)](3+) that quantitatively binds to the DNA on Au nanoparticles via electrostatic interaction. This method can detect the telomerase activity from as little as 10 cultured cancer cells without the polymerase chain reaction (PCR) amplification of telomerase extension product. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  10. Acquisition and processing method for human sensorial, sensitive, motory and phonatory circuits reaction times

    International Nuclear Information System (INIS)

    Doche, Claude

    1972-01-01

    This work describes a storage and acquisition device and a method for human sensorial and sensitive motory and phonatory reaction times. The considered circuits are those made with the visual, auditory and sensory receptor organs and the motory or phonatory effector organs. The anatomo-physiological localization of these circuits allows us to appreciate the possibilities of the central nervous system for different angles. The experimental population is made of normal and pathological individuals (individuals having tumoral or vascular, localized or diffused cerebral lesions or parkinsonian individuals). The parameter processing method is based on the multivariate analysis results and allows us to position each individual compared to a normal individual and to appreciate the weight of each circuit in this positioning. Clinical exploitation results give to this method a prognosis and therapeutic interest. It seems though untimely to talk about its diagnosis value. (author) [fr

  11. Butyrate down regulates BCL-XL and sensitizes human fibroblasts to radiation and chemotherapy induced apoptosis

    International Nuclear Information System (INIS)

    Chung, Diana H.; Ljungman, Mats; Zhang Fenfen; Chen Feng; McLaughlin, William P.

    1997-01-01

    Purpose/Objective: Butyrate is a short chain fatty acid that has been implicated in the induction of cell cycle arrest, cell differentiation and apoptosis. The purpose of this study was to determine if butyrate treatment sensitizes cells to radiation or chemotherapy induced apoptosis. Materials and Methods: Normal neonatal human diploid fibroblasts were used throughout this study. Apoptosis was scored and quantified using three different methods. First, cell morphology using propidium iodide and fluorescence microscopy was used to qualitatively determine apoptosis and to quantify the percentage of cells undergoing apoptosis. Second, apoptosis induced DNA degradation was scored by quantifying the amount of cells appearing in a sub-G1 peak using fixed and PI-stained cells and flow cytometry. Third, apoptosis-induced DNA degradation was examined by using an assay involving direct lysis of cells in the wells of agarose gels followed by conventional gel electrophoresis. Western blotting was used to quantify the cellular levels of the apoptosis regulators, Bcl-2, Bcl-XL and Bax. Results: Human diploid fibroblasts, which were resistant to radiation induced apoptosis, were found to undergo massive apoptosis when radiation was combined with butyrate treatment. Sensitization was obtained when butyrate was added before or after radiation although the combination of both pre and post-treatment was the most effective. Butyrate was also found to enhance UV light and cisplatin-induced apoptosis. These findings correlated with a reduction of the apoptosis antagonist Bcl-XL. Bcl-XL levels significantly dropped in a time and dose dependent manner. In addition, butyrate effectively blocked UV-induced accumulation of p53. Conclusion: Our results suggest that butyrate may be an attractive agent to use in combination with radiation or chemotherapy to lower the apoptotic threshold of tumor cells, regardless of the p53 status of the tumor cells

  12. Methylation-Sensitive Amplification Length Polymorphism (MS-AFLP) Microarrays for Epigenetic Analysis of Human Genomes.

    Science.gov (United States)

    Alonso, Sergio; Suzuki, Koichi; Yamamoto, Fumiichiro; Perucho, Manuel

    2018-01-01

    Somatic, and in a minor scale also germ line, epigenetic aberrations are fundamental to carcinogenesis, cancer progression, and tumor phenotype. DNA methylation is the most extensively studied and arguably the best understood epigenetic mechanisms that become altered in cancer. Both somatic loss of methylation (hypomethylation) and gain of methylation (hypermethylation) are found in the genome of malignant cells. In general, the cancer cell epigenome is globally hypomethylated, while some regions-typically gene-associated CpG islands-become hypermethylated. Given the profound impact that DNA methylation exerts on the transcriptional profile and genomic stability of cancer cells, its characterization is essential to fully understand the complexity of cancer biology, improve tumor classification, and ultimately advance cancer patient management and treatment. A plethora of methods have been devised to analyze and quantify DNA methylation alterations. Several of the early-developed methods relied on the use of methylation-sensitive restriction enzymes, whose activity depends on the methylation status of their recognition sequences. Among these techniques, methylation-sensitive amplification length polymorphism (MS-AFLP) was developed in the early 2000s, and successfully adapted from its original gel electrophoresis fingerprinting format to a microarray format that notably increased its throughput and allowed the quantification of the methylation changes. This array-based platform interrogates over 9500 independent loci putatively amplified by the MS-AFLP technique, corresponding to the NotI sites mapped throughout the human genome.

  13. Effect of growth temperature on lipid composition and ultraviolet sensitivity of human cells

    International Nuclear Information System (INIS)

    McAleer, M.A.; Moore, S.P.; Moss, S.H.

    1987-01-01

    Human skin fibroblasts were incubated at either 25 or 37 0 C before UV irradiation. Cells incubated at 25 0 C were more resistant to near UV radiation than cells grown at 37 0 C, but cells grown at the lower temperature were more sensitive to 254 nm radiation. Fatty acid analysis of membranes of cells showed that cells incubated at the lower temperature contained significantly higher amounts of linoleic acid (18:2) and linolenic acid (18:3) than cells incubated at 37 0 C. To determine if this difference in fatty acid content of the membranes was responsible for the UV survival characteristics of cells incubated at different temperatures, cells were enriched with either linoleate or linolenate during a 37 0 C incubation period. Gas chromatography revealed that cells incorporated the supplied fatty acid. Fatty acid enriched cells were then irradiated with near UV, and survival characteristics were compared to those obtained with cells grown at the lower incubation temperature. The results suggest that the different proportion of fatty acid content of the cells is not the cause of different UV sensitivities of cells grown at 25 0 C compared to cells grown at 37 0 C. (author)

  14. Inhibition of SRC-3 enhances sensitivity of human cancer cells to histone deacetylase inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Zou, Zhengzhi, E-mail: zouzhengzhi@m.scnu.edu.cn [MOE Key Laboratory of Laser Life Science and Institute of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou 510000 (China); Luo, Xiaoyong [Department of Oncology, The Affiliated Luoyang Central Hospital of Zhengzhou University, Luoyang 471000 (China); Nie, Peipei [KingMed Diagnostics and KingMed School of Laboratory Medicine, Guangzhou Medical University, Guangzhou 510000 (China); Wu, Baoyan; Zhang, Tao; Wei, Yanchun [MOE Key Laboratory of Laser Life Science and Institute of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou 510000 (China); Wang, Wenyi [Xiamen Cancer Center, Department of Medical Oncology, The First Affiliated Hospital of Xiamen University, Xiamen 361000 (China); Geng, Guojun; Jiang, Jie [Xiamen Cancer Center, Department of Thoracic Surgery, The First Affiliated Hospital of Xiamen University, Xiamen 361000 (China); Mi, Yanjun, E-mail: myjgj_77@163.com [Xiamen Cancer Center, Department of Medical Oncology, The First Affiliated Hospital of Xiamen University, Xiamen 361000 (China)

    2016-09-09

    SRC-3 is widely expressed in multiple tumor types and involved in cancer cell proliferation and apoptosis. Histone deacetylase (HDAC) inhibitors are promising antitumor drugs. However, the poor efficacy of HDAC inhibitors in solid tumors has restricted its further clinical application. Here, we reported the novel finding that depletion of SRC-3 enhanced sensitivity of breast and lung cancer cells to HDAC inhibitors (SAHA and romidepsin). In contrast, overexpression of SRC-3 decreased SAHA-induced cancer cell apoptosis. Furthermore, we found that SRC-3 inhibitor bufalin increased cancer cell apoptosis induced by HDAC inhibitors. The combination of bufalin and SAHA was particular efficient in attenuating AKT activation and reducing Bcl-2 levels. Taken together, these accumulating data might guide development of new breast and lung cancer therapies. - Highlights: • Depletion of SRC-3 enhanced sensitivity of breast and lung cancer cells to HDAC inhibitors. • Overexpression of SRC-3 enhanced cancer cell resistance to HDAC inhibitors. • SRC-3 inhibitor bufalin increased cancer cell apoptosis induced by HDAC inhibitors. • Bufalin synergized with HDAC inhibitor attenuated AKT activation and reduced Bcl-2 levels in human cancer cell.

  15. Reduced sensitivity to sooner reward during intertemporal decision-making following insula damage in humans

    Directory of Open Access Journals (Sweden)

    Manuela eSellitto

    2016-01-01

    Full Text Available During intertemporal choice, humans tend to prefer small-sooner rewards over larger-delayed rewards, reflecting temporal discounting (TD of delayed outcomes. Functional neuroimaging evidence has implicated the insular cortex in time-sensitive decisions, yet it is not clear whether activity in this brain region is crucial for, or merely associated with, TD behaviour. Here, patients with damage to the insula (Insular patients, control patients with lesions outside the insula, and healthy individuals chose between smaller-sooner and larger-later monetary rewards. Insular patients were less sensitive to sooner rewards than were the control groups, exhibiting reduced TD. A Voxel-based Lesion-Symptom Mapping (VLSM analysis confirmed a statistically significant association between insular damage and reduced TD. These results indicate that the insular cortex is crucial for intertemporal choice. We suggest that he insula may be necessary to anticipate the bodily/emotional effects of receiving rewards at different delays, influencing the computation of their incentive value. Devoid of such input, insular patients’ choices would be governed by a heuristic of quantity, allowing patients to wait for larger options.

  16. Ascorbic acid glycation of lens proteins produces UVA sensitizers similar to those in human lens

    International Nuclear Information System (INIS)

    Ortwerth, B.J.; Linetsky, Mikhail; Olesen, P.R.

    1995-01-01

    Soluble calf lens proteins were extensively glycated during a 4 week incubation with ascorbic acid in the presence of oxygen. Amino acids analysis of the dialyzed proteins removed at weekly intervals showed an increasing loss of lysine, arginine and histidine, consistent with the extensive protein cross-linking observed. Irradiation of the dialyzed samples with UVA light (1.0 kJ/cm 2 total illumination through a 338 nm cutoff filter) caused an increasing loss of tryptophan, an additional loss of histidine and the production of micromolar concentrations of hydrogen peroxide. No alteration in amino acid content and no photolytic effects were seen in proteins incubated without ascorbic acid in proteins incubated with glucose for 4 weeks. The rate of hydrogen peroxide formation was linear with each glycated sample with a maximum production of 25 nmol/mg protein illuminated. The possibility that the sensitizer activity was due to an ascorbate-induced oxidation of tryptophan was eliminated by the presence of a heavy metal ion chelator during the incubation and by showing equivalent effects with ascorbate-incubated ribonuclease A, which is devoid of tryptophan. The ascorbate-incubated samples displayed increasing absorbance at wavelengths above 300 nm and increasing fluorescence (340/430) as glycation proceeded. The spectra of the 4 week glycated proteins were identical to those obtained with a solubilized water-insoluble fraction from human lens, which is known to have UVA sensitizer activity. (Author)

  17. Skin Sensitive Difference of Human Body Sections under Clothing-Smirnov Test of Skin Surface Temperatures' Dynamic Changing

    Institute of Scientific and Technical Information of China (English)

    LI Jun; WU Hai-yan; WANG Yun-yi

    2004-01-01

    Skin sensitive difference of human body sections under clothing is the theoretic foundation of thermal insulation clothing design.By a new method of researching on clothing comfort perception,the skin temperature live changing procedure of human body sections affected by the same cold stimulation is inspected.Furthermore with the Smirnov test the skin temperatures dynamic changing patterns of main human body sections are obtained.

  18. Functional organization of the face-sensitive areas in human occipital-temporal cortex.

    Science.gov (United States)

    Shao, Hanyu; Weng, Xuchu; He, Sheng

    2017-08-15

    Human occipital-temporal cortex features several areas sensitive to faces, presumably forming the biological substrate for face perception. To date, there are piecemeal insights regarding the functional organization of these regions. They have come, however, from studies that are far from homogeneous with regard to the regions involved, the experimental design, and the data analysis approach. In order to provide an overall view of the functional organization of the face-sensitive areas, it is necessary to conduct a comprehensive study that taps into the pivotal functional properties of all the face-sensitive areas, within the context of the same experimental design, and uses multiple data analysis approaches. In this study, we identified the most robustly activated face-sensitive areas in bilateral occipital-temporal cortices (i.e., AFP, aFFA, pFFA, OFA, pcSTS, pSTS) and systemically compared their regionally averaged activation and multivoxel activation patterns to 96 images from 16 object categories, including faces and non-faces. This condition-rich and single-image analysis approach critically samples the functional properties of a brain region, allowing us to test how two basic functional properties, namely face-category selectivity and face-exemplar sensitivity are distributed among these regions. Moreover, by examining the correlational structure of neural responses to the 96 images, we characterize their interactions in the greater face-processing network. We found that (1) r-pFFA showed the highest face-category selectivity, followed by l-pFFA, bilateral aFFA and OFA, and then bilateral pcSTS. In contrast, bilateral AFP and pSTS showed low face-category selectivity; (2) l-aFFA, l-pcSTS and bilateral AFP showed evidence of face-exemplar sensitivity; (3) r-OFA showed high overall response similarities with bilateral LOC and r-pFFA, suggesting it might be a transitional stage between general and face-selective information processing; (4) r-aFFA showed high

  19. New sensitive direct radioimmunoassay for human plasma renin and its clinical application

    International Nuclear Information System (INIS)

    Higaki, J.; Ogihara, T.; Imai, N.; Kumahara, Y.; Hontani, S.; Nishiura, M.; Ogawa, H.; Hirose, S.; Murakami, K.

    1984-01-01

    A new sensitive direct radioimmunoassay for human plasma renin has been developed. Renin was purified from Haas' preparation utilizing a pepstatin-C 6 -Sepharose affinity chromatography. Antiserum, prepared by immunizing rabbits with the purified renin, was used for the direct radioimmunoassay at a final dilution of 1:30,000. The antibody was specific for human renal and plasma renin, but did not cross-react with cathepsin D, trypsin, or renins of mouse, dog, and rat. Radioimmunoassay was performed by the double antibody technique using the delayed tracer addition method. In this method, a standard curve was obtained over a range from 0.2 to 8.0 ng/ml. The values from this assay correlated well with total renin activity measured as the generation rate of angiotensin I after trypsin activation, but correlated weakly with active renin activity. This finding disclosed that both active and inactive renin were detected by this method. In normal participants, plasma renin concentration determined by direct radioimmunoassay was increased by standing and furosemide injection. The plasma renin concentration determined by direct radioimmunoassay of patients with essential hypertension was not significantly different from values in normal controls. The values were higher in patients with renovascular hypertension, malignant hypertension and Bartter's syndrome, but lower in patients with primary aldosteronism than in normal controls. 20 references, 7 figures

  20. Treatment of system dependencies and human interactions in PRA studies: a review and sensitivity study

    International Nuclear Information System (INIS)

    Orvis, D.D.; Joksimovich, V.; Worledge, D.H.

    1985-01-01

    The Electric Power Research Institute sponsored the review and comparison of five PRA studies: Arkansas Nuclear One - Unit 1, Big Rock Point, Grand Gulf, Limerick, and Zion - Unit 1. The review has been conducted in two phases. The Phase I review may be characterized as a qualitative look into many aspects of a PRA study. The Phase II review was performed to quantify the extent that differences in analytical techniques or key assumptions in these areas affect the differences in study results. In each of the PRA studies reviewed, the general descriptions of analytical approaches and descriptions of the analyses of event tree, fault tree and human interaction analyses that affected the dominant core damage sequences were reviewed. When these descriptions aroused interest because of seeming inconsistencies within the study or with other studies, they were pursued in some depth. The approaches or assumptions were contrasted to similar elements from other studies, and sensitivity analyses were performed in many cases to test the significance of results to the analytical models or assumptions. Inferences were drawn from the results regarding significance of the item to plant-specific results and, where possible, were generalized to other PRAs. This paper describes the results of the review of system dependencies and human interactions

  1. Characterization of lipid metabolism in insulin-sensitive adipocytes differentiated from immortalized human mesenchymal stem cells

    International Nuclear Information System (INIS)

    Prawitt, Janne; Niemeier, Andreas; Kassem, Moustapha; Beisiegel, Ulrike; Heeren, Joerg

    2008-01-01

    There is a great demand for cell models to study human adipocyte function. Here we describe the adipogenic differentiation of a telomerase-immortalized human mesenchymal stem cell line (hMSC-Tert) that maintains numerous features of terminally differentiated adipocytes even after prolonged withdrawal of the peroxisome proliferator activated receptor γ (PPARγ) agonist rosiglitazone. Differentiated hMSC-Tert developed the characteristic monolocular phenotype of mature adipocytes. The expression of adipocyte specific markers was highly increased during differentiation. Most importantly, the presence of the PPARγ agonist rosiglitazone was not required for the stable expression of lipoprotein lipase, adipocyte fatty acid binding protein and perilipin on mRNA and protein levels. Adiponectin expression was post-transcriptionally down-regulated in the absence of rosiglitazone. Insulin sensitivity as measured by insulin-induced phosphorylation of Akt and S6 ribosomal protein was also independent of rosiglitazone. In addition to commonly used adipogenic markers, we investigated further PPARγ-stimulated proteins with a role in lipid metabolism. We observed an increase of lipoprotein receptor (VLDLR, LRP1) and apolipoprotein E expression during differentiation. Despite this increased expression, the receptor-mediated endocytosis of lipoproteins was decreased in differentiated adipocytes, suggesting that these proteins may have an additional function in adipose tissue beyond lipoprotein uptake

  2. Human body temperature and new approaches to constructing temperature-sensitive bacterial vaccines.

    Science.gov (United States)

    White, Matthew D; Bosio, Catharine M; Duplantis, Barry N; Nano, Francis E

    2011-09-01

    Many of the live human and animal vaccines that are currently in use are attenuated by virtue of their temperature-sensitive (TS) replication. These vaccines are able to function because they can take advantage of sites in mammalian bodies that are cooler than the core temperature, where TS vaccines fail to replicate. In this article, we discuss the distribution of temperature in the human body, and relate how the temperature differential can be exploited for designing and using TS vaccines. We also examine how one of the coolest organs of the body, the skin, contains antigen-processing cells that can be targeted to provoke the desired immune response from a TS vaccine. We describe traditional approaches to making TS vaccines, and highlight new information and technologies that are being used to create a new generation of engineered TS vaccines. We pay particular attention to the recently described technology of substituting essential genes from Arctic bacteria for their homologues in mammalian pathogens as a way of creating TS vaccines.

  3. Androgen-Sensitized Apoptosis of HPr-1AR Human Prostate Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Congcong Chen

    Full Text Available Androgen receptor (AR signaling is crucial to the development and homeostasis of the prostate gland, and its dysregulation mediates common prostate pathologies. The mechanisms whereby AR regulates growth suppression and differentiation of luminal epithelial cells in the prostate gland and proliferation of malignant versions of these cells have been investigated in human and rodent adult prostate. However, the cellular stress response of human prostate epithelial cells is not well understood, though it is central to prostate health and pathology. Here, we report that androgen sensitizes HPr-1AR and RWPE-AR human prostate epithelial cells to cell stress agents and apoptotic cell death. Although 5α-dihydrotestosterone (DHT treatment alone did not induce cell death, co-treatment of HPr-1AR cells with DHT and an apoptosis inducer, such as staurosporine (STS, TNFt, or hydrogen peroxide, synergistically increased cell death in comparison to treatment with each apoptosis inducer by itself. We found that the synergy between DHT and apoptosis inducer led to activation of the intrinsic/mitochondrial apoptotic pathway, which is supported by robust cleavage activation of caspase-9 and caspase-3. Further, the dramatic depolarization of the mitochondrial membrane potential that we observed upon co-treatment with DHT and STS is consistent with increased mitochondrial outer membrane permeabilization (MOMP in the pro-apoptotic mechanism. Interestingly, the synergy between DHT and apoptosis inducer was abolished by AR antagonists and inhibitors of transcription and protein synthesis, suggesting that AR mediates pro-apoptotic synergy through transcriptional regulation of MOMP genes. Expression analysis revealed that pro-apoptotic genes (BCL2L11/BIM and AIFM2 were DHT-induced, whereas pro-survival genes (BCL2L1/BCL-XL and MCL1 were DHT-repressed. Hence, we propose that the net effect of these AR-mediated expression changes shifts the balance of BCL2-family proteins

  4. Glucose Metabolism of Human Prostate Cancer Mouse Xenografts

    Directory of Open Access Journals (Sweden)

    Hossein Jadvar

    2005-04-01

    Full Text Available We hypothesized that the glucose metabolism of prostate cancer is modulated by androgen. We performed in vivo biodistribution and imaging studies of [F-18] fluorodeoxyglucose (FDG accumulation in androgen-sensitive (CWR-22 and androgen-independent (PC-3 human prostate cancer xenografts implanted in castrated and noncastrated male athymic mice. The growth pattern of the CWR-22 tumor was best approximated by an exponential function (tumor size in mm3 = 14.913 e0.108 × days, R2 = .96, n = 5. The growth pattern of the PC-3 tumor was best approximated by a quadratic function (tumor size in mm3 = 0.3511 × days2 + 49.418 × day −753.33, R2 = .96, n = 3. The FDG accumulation in the CWR-22 tumor implanted in the castrated mice was significantly lower, by an average of 55%, in comparison to that implanted in the noncastrated host (1.27 vs. 2.83, respectively, p < .05. The 3-week maximal standardized uptake value (SUVmax was 0.99 ± 0.43 (mean ± SD for CWR-22 and 1.21 ± 0.32 for PC-3, respectively. The 5-week SUVmax was 1.22 ± 0.08 for CWR-22 and 1.35 ± 0.17 for PC-3, respectively. The background muscle SUVmax was 0.53 ± 0.11. Glucose metabolism was higher in the PC-3 tumor than in the CWR-22 tumor at both the 3-week (by 18% and the 5-week (by 9.6% micro-PET imaging sessions. Our results support the notions that FDG PET may be useful in the imaging evaluation of response to androgen ablation therapy and in the early prediction of hormone refractoriness in men with metastatic prostate cancer.

  5. Contrasting effects of ultraviolet-A and ultraviolet B exposure on induction of contact sensitivity in human skin

    DEFF Research Database (Denmark)

    Skov, Lone; Hansen, Henrik; Barker, J. N.

    1997-01-01

    Ultraviolet-B (UVB), in addition to direct effects on DNA, induces immunological changes in the skin that predispose to the development of skin cancer. Whether ultraviolet-A (UVA) induces similar changes is unknown. This effect can be investigated in humans in vivo using epicutaneous antigens...... as a model of tumour antigens. Volunteers (n = 46) were randomly assigned to received no sensitization, sensitization with the allergen diphenylcyclopropenone (DPCP) on non-UV-exposed normal skin, or sensitization with DPCP on skin exposed to three minimal erythema doses (MED) of either UVA or UVB radiation...... the immunization rate compared with sensitization on non-irradiated skin (P UVA radiation did not result in a decreased immunization rate compared with non-irradiated skin. These results indicate that in humans erythemagenic...

  6. Study of radiation sensitization of artesunate on human HeLa cells of cervical cancer

    International Nuclear Information System (INIS)

    Ji Rong; Cao Jianping; Chen Xialin; Zhu Wei; Jiang Qing; Pan Chunyan; Zhou Yuanyuan; Feng Yang; Peng Xiaomei; Liu Yang; Fan Saijun

    2010-01-01

    Objective: To investigate the radiosensitizing effects of artesunate on human HeLa cells of cervical cancer in vitro. Methods: Hela cells irradiated with 60 Co γ-rays. The dose rate was 0.635 Gy/min and the radiation dose was 0, 1, 2, 4, 6 Gy, respectively. The anti-proliferation activities of artesunate on HeLa cells were evaluated with MTT assay, to determine the most appropriate drug concentration. The effect of radiosensitivity was observed by using clonogenic assay. The single-hit multi-target model was used to plot the HeLa cell's dose-survival curve, to calculate mean lethal dose, quasi-threshold dose and sensitization enhancement rate, and to evaluate its radiosensitization effect. The apoptosis was analyzed with flow cytometry (FCM) to further test the radiation sensitization of artesunate on HeLa cells. Results: The inhibition of artesunate on HeLa cells increased with concentration. In radiation group, the cell cloning efficiency were 91.67%, 82.02%, 58.06%, 25.01%, respectively, and in artesunate (2.0 μmol/L) + radiation group, the cell cloning efficiency were 74.93%, 60.53%, 22.38%, 5.05%. In radiation group and artesunate (2.0 μmol/L) + radiation group, the mean lethal dose (D 0 ) was 2.95 and 2.07 Gy, respectively, while the qusai-threshold dose (D q ) were 2.01 and 1.24 Gy, respectively, and SER was 1.43. Compared with 2 and 6 Gy radiation group, the apoptosis rate of drug + radiation group increased from 12.26%, 40.08% to 22.71%, 59.92. Conclusions: The inhibiting effect of artesunate on HeLa cells is concentration-dependent. Artesunate has radiosensitizing effect on HeLa cells in vitro. (authors)

  7. Sensitivity of low energy brachytherapy Monte Carlo dose calculations to uncertainties in human tissue composition

    Energy Technology Data Exchange (ETDEWEB)

    Landry, Guillaume; Reniers, Brigitte; Murrer, Lars; Lutgens, Ludy; Bloemen-Van Gurp, Esther; Pignol, Jean-Philippe; Keller, Brian; Beaulieu, Luc; Verhaegen, Frank [Department of Radiation Oncology (MAASTRO), GROW-School for Oncology and Developmental Biology, Maastricht University Medical Center, Maastricht 6201 BN (Netherlands); Department of Radiation Oncology, Sunnybrook Health Sciences Centre, University of Toronto, Toronto, Ontario M4N 3M5 (Canada); Departement de Radio-Oncologie et Centre de Recherche en Cancerologie, de l' Universite Laval, CHUQ, Pavillon L' Hotel-Dieu de Quebec, Quebec G1R 2J6 (Canada) and Departement de Physique, de Genie Physique et d' Optique, Universite Laval, Quebec G1K 7P4 (Canada); Department of Radiation Oncology (MAASTRO), GROW-School for Oncology and Developmental Biology, Maastricht University Medical Center, Maastricht 6201 BN (Netherlands) and Medical Physics Unit, McGill University, Montreal General Hospital, Montreal, Quebec H3G 1A4 (Canada)

    2010-10-15

    Purpose: The objective of this work is to assess the sensitivity of Monte Carlo (MC) dose calculations to uncertainties in human tissue composition for a range of low photon energy brachytherapy sources: {sup 125}I, {sup 103}Pd, {sup 131}Cs, and an electronic brachytherapy source (EBS). The low energy photons emitted by these sources make the dosimetry sensitive to variations in tissue atomic number due to the dominance of the photoelectric effect. This work reports dose to a small mass of water in medium D{sub w,m} as opposed to dose to a small mass of medium in medium D{sub m,m}. Methods: Mean adipose, mammary gland, and breast tissues (as uniform mixture of the aforementioned tissues) are investigated as well as compositions corresponding to one standard deviation from the mean. Prostate mean compositions from three different literature sources are also investigated. Three sets of MC simulations are performed with the GEANT4 code: (1) Dose calculations for idealized TG-43-like spherical geometries using point sources. Radial dose profiles obtained in different media are compared to assess the influence of compositional uncertainties. (2) Dose calculations for four clinical prostate LDR brachytherapy permanent seed implants using {sup 125}I seeds (Model 2301, Best Medical, Springfield, VA). The effect of varying the prostate composition in the planning target volume (PTV) is investigated by comparing PTV D{sub 90} values. (3) Dose calculations for four clinical breast LDR brachytherapy permanent seed implants using {sup 103}Pd seeds (Model 2335, Best Medical). The effects of varying the adipose/gland ratio in the PTV and of varying the elemental composition of adipose and gland within one standard deviation of the assumed mean composition are investigated by comparing PTV D{sub 90} values. For (2) and (3), the influence of using the mass density from CT scans instead of unit mass density is also assessed. Results: Results from simulation (1) show that variations

  8. Development of a sensitive and specific epitope-blocking ELISA for universal detection of antibodies to human enterovirus 71 strains.

    Directory of Open Access Journals (Sweden)

    Fang He

    Full Text Available BACKGROUND: Human Enterovirus 71 (EV71 is a common cause of hand, foot and mouth disease (HFMD in young children. It is often associated with severe neurological diseases and mortalities in recent outbreaks across the Asia Pacific region. Currently, there is no efficient universal antibody test available to detect EV71 infections. METHODOLOGY/PRINCIPAL FINDING: In the present study, an epitope-blocking ELISA was developed to detect specific antibodies to human EV71 viruses in human or animal sera. The assay relies on a novel monoclonal antibody (Mab 1C6 that specifically binds to capsid proteins in whole EV71 viruses without any cross reaction to any EV71 capsid protein expressed alone. The sensitivity and specificity of the epitope-blocking ELISA for EV71 was evaluated and compared to microneutralization using immunized animal sera to multiple virus genotypes of EV71 and coxsackieviruses. Further, 200 serum sample from human individuals who were potentially infected with EV71 viruses were tested in both the blocking ELISA and microneutralization. Results indicated that antibodies to EV71 were readily detected in immunized animals or human sera by the epitope blocking ELISA whereas specimens with antibodies to other enteroviruses yielded negative results. This assay is not only simpler to perform but also shows higher sensitivity and specificity as compared to microneutralization. CONCLUSION: The epitope-blocking ELISA based on a unique Mab 1C6 provided highly sensitive and 100% specific detection of antibodies to human EV71 viruses in human sera.

  9. Sensitization of human cells by inhibitors of DNA synthesis following the action of DNA-damaging agents

    Energy Technology Data Exchange (ETDEWEB)

    Filatov, M.V.; Noskin, L.A. (Leningrad Inst. of Nuclear Physics, Gatchina (USSR))

    1983-08-01

    Inhibitors of DNA synthesis 1-..beta..-arabinofuranosylcytosine (Ac) and hydroxyurea (Hu) taken together drastically sensitized human cells to the killing effect of DNA-damaging agents. For UV-irradiation this sensitization depended on the cells' ability for excision repair. By using viscoelastometric methods of measurement of double-strand breaks (DSB) in the genome, it was established that the first DSB were generated after incubation of the damaged cells in the mixture of inhibitors at about the same dose when sensitization appeared. A scheme is proposed to describe molecular events associated with the phenomenon studied. 35 refs.

  10. Human Neuronal Calcium Sensor-1 Protein Avoids Histidine Residues To Decrease pH Sensitivity.

    Science.gov (United States)

    Gong, Yehong; Zhu, Yuzhen; Zou, Yu; Ma, Buyong; Nussinov, Ruth; Zhang, Qingwen

    2017-01-26

    pH is highly regulated in mammalian central nervous systems. Neuronal calcium sensor-1 (NCS-1) can interact with numerous target proteins. Compared to that in the NCS-1 protein of Caenorhabditis elegans, evolution has avoided the placement of histidine residues at positions 102 and 83 in the NCS-1 protein of humans and Xenopus laevis, possibly to decrease the conformational sensitivity to pH gradients in synaptic processes. We used all-atom molecular dynamics simulations to investigate the effects of amino acid substitutions between species on human NCS-1 by substituting Arg102 and Ser83 for histidine at neutral (R102H and S83H) and acidic pHs (R102H p and S83H p ). Our cumulative 5 μs simulations revealed that the R102H mutation slightly increases the structural flexibility of loop L2 and the R102H p mutation decreases protein stability. Community network analysis illustrates that the R102H and S83H mutations weaken the interdomain and strengthen the intradomain communications. Secondary structure contents in the S83H and S83H p mutants are similar to those in the wild type, whereas the global structural stabilities and salt-bridge probabilities decrease. This study highlights the conformational dynamics effects of the R102H and S83H mutations on the local structural flexibility and global stability of NCS-1, whereas protonated histidine decreases the stability of NCS-1. Thus, histidines at positions 102 and 83 may not be compatible with the function of NCS-1 whether in the neutral or protonated state.

  11. Does the A-not-B error in adult pet dogs indicate sensitivity to human communication?

    Science.gov (United States)

    Kis, Anna; Topál, József; Gácsi, Márta; Range, Friederike; Huber, Ludwig; Miklósi, Adám; Virányi, Zsófia

    2012-07-01

    Recent dog-infant comparisons have indicated that the experimenter's communicative signals in object hide-and-search tasks increase the probability of perseverative (A-not-B) errors in both species (Topál et al. 2009). These behaviourally similar results, however, might reflect different mechanisms in dogs and in children. Similar errors may occur if the motor response of retrieving the object during the A trials cannot be inhibited in the B trials or if the experimenter's movements and signals toward the A hiding place in the B trials ('sham-baiting') distract the dogs' attention. In order to test these hypotheses, we tested dogs similarly to Topál et al. (2009) but eliminated the motor search in the A trials and 'sham-baiting' in the B trials. We found that neither an inability to inhibit previously rewarded motor response nor insufficiencies in their working memory and/or attention skills can explain dogs' erroneous choices. Further, we replicated the finding that dogs have a strong tendency to commit the A-not-B error after ostensive-communicative hiding and demonstrated the crucial effect of socio-communicative cues as the A-not-B error diminishes when location B is ostensively enhanced. These findings further support the hypothesis that the dogs' A-not-B error may reflect a special sensitivity to human communicative cues. Such object-hiding and search tasks provide a typical case for how susceptibility to human social signals could (mis)lead domestic dogs.

  12. Sensitivity of PCR assays for murine gammaretroviruses and mouse contamination in human blood samples.

    Directory of Open Access Journals (Sweden)

    Li Ling Lee

    Full Text Available Gammaretroviruses related to murine leukemia virus (MLV have variously been reported to be present or absent in blood from chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME patients and healthy controls. Using subjects from New York State, we have investigated by PCR methods whether MLV-related sequences can be identified in nucleic acids isolated from whole blood or from peripheral blood mononuclear cells (PBMCs or following PBMC culture. We have also passaged the prostate cancer cell line LNCaP following incubation with plasma from patients and controls and assayed nucleic acids for viral sequences. We have used 15 sets of primers that can effectively amplify conserved regions of murine endogenous and exogenous retrovirus sequences. We demonstrate that our PCR assays for MLV-related gag sequences and for mouse DNA contamination are extremely sensitive. While we have identified MLV-like gag sequences following PCR on human DNA preparations, we are unable to conclude that these sequences originated in the blood samples.

  13. Screening, sensitivity, and uncertainty for the CREAM method of Human Reliability Analysis

    International Nuclear Information System (INIS)

    Bedford, Tim; Bayley, Clare; Revie, Matthew

    2013-01-01

    This paper reports a sensitivity analysis of the Cognitive Reliability and Error Analysis Method for Human Reliability Analysis. We consider three different aspects: the difference between the outputs of the Basic and Extended methods, on the same HRA scenario; the variability in outputs through the choices made for common performance conditions (CPCs); and the variability in outputs through the assignment of choices for cognitive function failures (CFFs). We discuss the problem of interpreting categories when applying the method, compare its quantitative structure to that of first generation methods and discuss also how dependence is modelled with the approach. We show that the control mode intervals used in the Basic method are too narrow to be consistent with the Extended method. This motivates a new screening method that gives improved accuracy with respect to the Basic method, in the sense that (on average) halves the uncertainty associated with the Basic method. We make some observations on the design of a screening method that are generally applicable in Risk Analysis. Finally, we propose a new method of combining CPC weights with nominal probabilities so that the calculated probabilities are always in range (i.e. between 0 and 1), while satisfying sensible properties that are consistent with the overall CREAM method

  14. A novel BCI based on ERP components sensitive to configural processing of human faces

    Science.gov (United States)

    Zhang, Yu; Zhao, Qibin; Jing, Jin; Wang, Xingyu; Cichocki, Andrzej

    2012-04-01

    This study introduces a novel brain-computer interface (BCI) based on an oddball paradigm using stimuli of facial images with loss of configural face information (e.g., inversion of face). To the best of our knowledge, till now the configural processing of human faces has not been applied to BCI but widely studied in cognitive neuroscience research. Our experiments confirm that the face-sensitive event-related potential (ERP) components N170 and vertex positive potential (VPP) have reflected early structural encoding of faces and can be modulated by the configural processing of faces. With the proposed novel paradigm, we investigate the effects of ERP components N170, VPP and P300 on target detection for BCI. An eight-class BCI platform is developed to analyze ERPs and evaluate the target detection performance using linear discriminant analysis without complicated feature extraction processing. The online classification accuracy of 88.7% and information transfer rate of 38.7 bits min-1 using stimuli of inverted faces with only single trial suggest that the proposed paradigm based on the configural processing of faces is very promising for visual stimuli-driven BCI applications.

  15. Coupled multiview autoencoders with locality sensitivity for three-dimensional human pose estimation

    Science.gov (United States)

    Yu, Jialin; Sun, Jifeng; Luo, Shasha; Duan, Bichao

    2017-09-01

    Estimating three-dimensional (3D) human poses from a single camera is usually implemented by searching pose candidates with image descriptors. Existing methods usually suppose that the mapping from feature space to pose space is linear, but in fact, their mapping relationship is highly nonlinear, which heavily degrades the performance of 3D pose estimation. We propose a method to recover 3D pose from a silhouette image. It is based on the multiview feature embedding (MFE) and the locality-sensitive autoencoders (LSAEs). On the one hand, we first depict the manifold regularized sparse low-rank approximation for MFE and then the input image is characterized by a fused feature descriptor. On the other hand, both the fused feature and its corresponding 3D pose are separately encoded by LSAEs. A two-layer back-propagation neural network is trained by parameter fine-tuning and then used to map the encoded 2D features to encoded 3D poses. Our LSAE ensures a good preservation of the local topology of data points. Experimental results demonstrate the effectiveness of our proposed method.

  16. Assessing the sensitivity of human skin hyperspectral responses to increasing anemia severity levels

    Science.gov (United States)

    Baranoski, Gladimir V. G.; Dey, Ankita; Chen, Tenn F.

    2015-09-01

    Anemia is a prevalent medical condition that seriously affects millions of people all over the world. In many regions, not only its initial detection but also its monitoring are hindered by limited access to laboratory facilities. This situation has motivated the development of a wide range of optical devices and procedures to assist physicians in these tasks. Although noticeable progress has been achieved in this area, the search for reliable, low-cost, and risk-free solutions still continues, and the strengthening of the knowledge base about this disorder and its effects is essential for the success of these initiatives. We contribute to these efforts by closely examining the sensitivity of human skin hyperspectral responses (within and outside the visible region of the light spectrum) to reduced hemoglobin concentrations associated with increasing anemia severity levels. This investigation, which involves skin specimens with distinct biophysical and morphological characteristics, is supported by controlled in silico experiments performed using a predictive light transport model and measured data reported in the biomedical literature. We also propose a noninvasive procedure to be employed in the monitoring of this condition at the point-of-care.

  17. Resveratrol-Sensitized UVA Induced Apoptosis in Human Keratinocytes through Mitochondrial Oxidative Stress and Pore Opening

    Science.gov (United States)

    Boyer, Jean Z; Jandova, Jana; Janda, Jaroslav; Vleugels, Frank R; Elliott, David; Sligh, James E

    2012-01-01

    Resveratrol (3, 5, 4′-trihydroxy- trans- stilbene), a polyphenol compound, is derived from natural products such as the skin of red grapes, blueberries and cranberries. Resveratrol not only exhibits antioxidant, cardioprotection, and anti-aging properties, but can also inhibit cancer cell growth and induce apoptosis. It has been shown that resveratrol inhibits the activation of Nf-kB and subsequently down regulates the expression of Nf-kB regulated genes such as interleukin-2 and Bcl-2, leading to cell cycle arrest and increased apoptosis in multiple myeloma cells. In the skin, resveratrol has been reported to sensitize keratinocytes to UVA induced apoptosis. However, the effect of resveratrol on opening of the mitochondrial permeability transition pore has not been previously examined. Our data show that UVA (14J/cm2) along with resveratrol causes massive oxidative stress in mitochondria. As a consequence of oxidative stress, the mitochondrial membrane potential decreases which results in opening of the mitochondrial pores ultimately leading to apoptosis in human keratinocytes. These results may have clinical implications for development of future chemotherapeutic treatment for tumors of the skin. PMID:22673012

  18. Widespread molecular patterns associated with drug sensitivity in breast cancer cell lines, with implications for human tumors.

    Directory of Open Access Journals (Sweden)

    Chad J Creighton

    Full Text Available BACKGROUND: Recent landmark studies have profiled cancer cell lines for molecular features, along with measuring the corresponding growth inhibitory effects for specific drug compounds. These data present a tool for determining which subsets of human cancer might be more responsive to particular drugs. To this end, the NCI-DREAM-sponsored DREAM7: Drug Sensitivity Prediction Challenge (sub-challenge 1 set out to predict the sensitivities of 18 breast cancer cell lines to 31 previously untested compounds, on the basis of molecular profiling data and a training subset of cell lines. METHODS AND RESULTS: With 47 teams submitting blinded predictions, team Creighton scored third in terms of overall accuracy. Team Creighton's method was simple and straightforward, incorporated multiple expression data types (RNA-seq, gene array, RPPA, and incorporated all profiled features (not only the "best" predictive ones. As an extension of the approach, cell line data, from public datasets of expression profiling coupled with drug sensitivities (Barretina, Garnett, Heiser were used to "predict" the drug sensitivities in human breast tumors (using data from The Cancer Genome Atlas. Drug sensitivity correlations within human breast tumors showed differences by expression-based subtype, with many associations in line with the expected (e.g. Lapatinib sensitivity in HER2-enriched cancers and others inviting further study (e.g. relative resistance to PI3K inhibitors in basal-like cancers. CONCLUSIONS: Molecular patterns associated with drug sensitivity are widespread, with potentially hundreds of genes that could be incorporated into making predictions, as well as offering biological clues as to the mechanisms involved. Applying the cell line patterns to human tumor data may help generate hypotheses on what tumor subsets might be more responsive to therapies, where multiple cell line datasets representing various drugs may be used, in order to assess consistency of

  19. Thymidine kinase deficient human cells have increased UV sensitivity in their capacity to support herpes simplex virus but normal UV sensitivity for colony formation

    International Nuclear Information System (INIS)

    Rainbow, A.J.

    1989-01-01

    A thymidine kinase deficient (tk - ) and two thymidine kinase proficient (tk + ) human cell lines were compared for UV sensitivity using colony-forming ability as well as their capacity to support the plaque formation of herpes simplex type 1 (HSV-1).The tk - line (143 cells) was a derivative of one of the tk + lines (R970-5), whereas the other tk + line (AC4 cells) was a derivative of the 143 cells obtained by transfection with purified sheared HSV-2 DNA encoding the viral tk gene. 143, R970-5 and AC4 cells showed a similar UV sensitivity for colony-forming ability. In contrast, the capacity to support HSV-1 plaque formation immediately (within 1 h) afte UV-irradiation was reduced to a greater extent in the 143 cells compared to the R970-5 and AC4 cells. Capacity curves for plaque formation of the HSV-1: KOS wild-type (tk + ) strain were similar to those for the HSV-1: PTK3B mutant (tk - ) strain were similar to those for the HSV-1: PTK3B mutant (tk - ) strain in the 3 cell strains, indicating that the viral tk gene does not influence the ability of HSV-1 to form plaques in UV-irradiated compared to unirradiated human cells. Cellular capacity for HSV-1 plaque formation was found to recover in both tk - and tk + cells for cultures infected 24 h after UV-irradiation. These results suggest that repair of UV-damaged DNA takes place to a similar extent in both tk - and tk + human cells, but the kinetics of repair are initially slower in tk - compared to tk + human cells. (author). 33 refs.; 3 figs.; 1 tab

  20. Skeletal muscle phosphatidylcholine and phosphatidylethanolamine are related to insulin sensitivity and respond to acute exercise in humans.

    Science.gov (United States)

    Newsom, Sean A; Brozinick, Joseph T; Kiseljak-Vassiliades, Katja; Strauss, Allison N; Bacon, Samantha D; Kerege, Anna A; Bui, Hai Hoang; Sanders, Phil; Siddall, Parker; Wei, Tao; Thomas, Melissa; Kuo, Ming Shang; Nemkov, Travis; D'Alessandro, Angelo; Hansen, Kirk C; Perreault, Leigh; Bergman, Bryan C

    2016-06-01

    Several recent reports indicate that the balance of skeletal muscle phosphatidylcholine (PC) and phosphatidylethanolamine (PE) is a key determinant of muscle contractile function and metabolism. The purpose of this study was to determine relationships between skeletal muscle PC, PE and insulin sensitivity, and whether PC and PE are dynamically regulated in response to acute exercise in humans. Insulin sensitivity was measured via intravenous glucose tolerance in sedentary obese adults (OB; n = 14), individuals with type 2 diabetes (T2D; n = 15), and endurance-trained athletes (ATH; n = 15). Vastus lateralis muscle biopsies were obtained at rest, immediately after 90 min of cycle ergometry at 50% maximal oxygen consumption (V̇o2 max), and 2-h postexercise (recovery). Skeletal muscle PC and PE were measured via infusion-based mass spectrometry/mass spectrometry analysis. ATH had greater levels of muscle PC and PE compared with OB and T2D (P insulin sensitivity (both P insulin sensitivity among the entire cohort (r = -0.43, P = 0.01). Muscle PC and PE were altered by exercise, particularly after 2 h of recovery, in a highly group-specific manner. However, muscle PC:PE ratio remained unchanged in all groups. In summary, total muscle PC and PE are positively related to insulin sensitivity while PC:PE ratio is inversely related to insulin sensitivity in humans. A single session of exercise significantly alters skeletal muscle PC and PE levels, but not PC:PE ratio. Copyright © 2016 the American Physiological Society.

  1. Evidence for the speed-value trade-off: human and monkey decision making is magnitude sensitive.

    Science.gov (United States)

    Pirrone, Angelo; Azab, Habiba; Hayden, Benjamin Y; Stafford, Tom; Marshall, James A R

    2018-04-01

    Complex natural systems from brains to bee swarms have evolved to make adaptive multifactorial decisions. Recent theoretical and empirical work suggests that many evolved systems may take advantage of common motifs across multiple domains. We are particularly interested in value sensitivity (i.e., sensitivity to the magnitude or intensity of the stimuli or reward under consideration) as a mechanism to resolve deadlocks adaptively. This mechanism favours long-term reward maximization over accuracy in a simple manner, because it avoids costly delays associated with ambivalence between similar options; speed-value trade-offs have been proposed to be evolutionarily advantageous for many kinds of decision. A key prediction of the value-sensitivity hypothesis is that choices between equally-valued options will proceed faster when the options have a high value than when they have a low value. However, value-sensitivity is not part of idealised choice models such as diffusion to bound. Here we examine two different choice behaviours in two different species, perceptual decisions in humans and economic choices in rhesus monkeys, to test this hypothesis. We observe the same value sensitivity in both human perceptual decisions and monkey value-based decisions. These results endorse the idea that neural decision systems make use of the same basic principle of value-sensitivity in order to resolve costly deadlocks and thus improve long-term reward intake.

  2. Sensitive and simultaneous quantification of zinc pyrithione and climbazole deposition from anti-dandruff shampoos onto human scalp

    NARCIS (Netherlands)

    Chen, G.; Miao, M.; Hoptroff, M.; Fei, X.; Collins, L.Z.; Jones, A.; Janssen, H.G.

    2015-01-01

    A sensitive ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method has been developed and validated for simultaneous quantification of zinc pyrithione (ZPT) and climbazole (CBZ) deposited onto human scalp from anti-dandruff (AD) shampoos. Scrubbing with a buffer

  3. Proteasome inhibitor MG132 sensitizes HPV-positive human cervical cancer cells to rhTRAIL-induced apoptosis

    NARCIS (Netherlands)

    Hougardy, BMT; Maduro, JH; van der Zee, AGJ; de Groot, DJA; van den Heuvel, FAJ; de Vries, EGE; de Jong, S

    2006-01-01

    In cervical carcinogenesis, the p53 tumor suppressor pathway is disrupted by HPV (human papilloma virus) E6 oncogene expression. E6 targets p53 for rapid proteasome-mediated degradation. We therefore investigated whether proteasome inhibition by MG132 could restore wild-type p53 levels and sensitize

  4. Study on chemotherapeutic sensitizing effect of nimotuzumab on different human esophageal squamous carcinoma cells.

    Science.gov (United States)

    Yang, Xiaoyu; Ji, Yinghua; Kang, Xiaochun; Chen, Meiling; Kou, Weizheng; Jin, Cailing; Lu, Ping

    2016-02-01

    Esophageal cancer is one of the leading causes of mortality worldwide. Although, surgery, radio- and chemotherapy are used to treat the disease, the identification of new drugs is crucial to increase the curative effect. The aim of the present study was to examine the chemotherapeutic sensitizing effect of nimotuzumab (h-R3) and cisplatin cytotoxic drugs cisplatin (DDP) and 5-fluorouracil (5-FU) on esophageal carcinoma cells with two different epidermal growth factor receptor (EGFR) expressions. The expression of EGFR was detected in the human EC1 or EC9706 esophageal squamous cell carcinoma cell line using immunohistochemistry. The inhibitory effect of DDP and 5-FU alone or combined with h-R3 on EC1 or EC9706 cell proliferation was detected using an MTT assay. Flow cytometry and the TUNEL assay were used to determine the effect of single or combined drug treatment on cell apoptosis. The results showed that the expression of EGFR was low in EC1 cells but high in EC9706 cells. The inhibitory effect of the single use of h-R3 on EC1 or EC9706 cell proliferation was decreased. The inhibitory effect between single use of h-R3 alone and combined use of the chemotherapy drugs showed no statistically significant difference (P>0.05) on the EC1 cell growth rate, but showed a statistically significant difference (a=0.05) on EC9706 cell growth rate. The results detected by flow cytometry and TUNEL assay showed that the difference between single use of h-R3 alone and the control group was statistically significant with regard to the EC1 apoptosis rate effect (P0.05). However, statistically significant differences were identified in the apoptotic rate of EC9706 cells between the h-R3 combined chemotherapy group and single chemotherapy group (P0.05). In conclusion, the sensitization effect of h-R3 on chemotherapy drugs is associated with the expression level of EGFR in EC1 or EC9706 cells. The cell killing effect of the combined use of h-R3 with DDP and 5-FU showed no obvious

  5. Sensitive cell-based assay for determination of human immunodeficiency virus type 1 coreceptor tropism.

    Science.gov (United States)

    Weber, Jan; Vazquez, Ana C; Winner, Dane; Gibson, Richard M; Rhea, Ariel M; Rose, Justine D; Wylie, Doug; Henry, Kenneth; Wright, Alison; King, Kevin; Archer, John; Poveda, Eva; Soriano, Vicente; Robertson, David L; Olivo, Paul D; Arts, Eric J; Quiñones-Mateu, Miguel E

    2013-05-01

    CCR5 antagonists are a powerful new class of antiretroviral drugs that require a companion assay to evaluate the presence of CXCR4-tropic (non-R5) viruses prior to use in human immunodeficiency virus (HIV)-infected individuals. In this study, we have developed, characterized, verified, and prevalidated a novel phenotypic test to determine HIV-1 coreceptor tropism (VERITROP) based on a sensitive cell-to-cell fusion assay. A proprietary vector was constructed containing a near-full-length HIV-1 genome with the yeast uracil biosynthesis (URA3) gene replacing the HIV-1 env coding sequence. Patient-derived HIV-1 PCR products were introduced by homologous recombination using an innovative yeast-based cloning strategy. The env-expressing vectors were then used in a cell-to-cell fusion assay to determine the presence of R5 and/or non-R5 HIV-1 variants within the viral population. Results were compared with (i) the original version of Trofile (Monogram Biosciences, San Francisco, CA), (ii) population sequencing, and (iii) 454 pyrosequencing, with the genotypic data analyzed using several bioinformatics tools, i.e., the 11/24/25 rule, Geno2Pheno (2% to 5.75%, 3.5%, or 10% false-positive rate [FPR]), and webPSSM. VERITROP consistently detected minority non-R5 variants from clinical specimens, with an analytical sensitivity of 0.3%, with viral loads of ≥1,000 copies/ml, and from B and non-B subtypes. In a pilot study, a 73.7% (56/76) concordance was observed with the original Trofile assay, with 19 of the 20 discordant results corresponding to non-R5 variants detected using VERITROP and not by the original Trofile assay. The degree of concordance of VERITROP and Trofile with population and deep sequencing results depended on the algorithm used to determine HIV-1 coreceptor tropism. Overall, VERITROP showed better concordance with deep sequencing/Geno2Pheno at a 0.3% detection threshold (67%), whereas Trofile matched better with population sequencing (79%). However, 454

  6. Highly sensitive detection of human IgG using a novel bio-barcode assay combined with DNA chip technology

    International Nuclear Information System (INIS)

    Liu, Zhenbao; Zhou, Bo; Wang, Haiqing; Lu, Feng; Liu, Tianjun; Song, Cunxian; Leng, Xigang

    2013-01-01

    A simple and ultrasensitive detection of human IgG based on signal amplification using a novel bio-barcode assay and DNA chip technology was developed. The sensing platform was a sandwich system made up of antibody-modified magnetic microparticles (Ab-MMPs)/human IgG/Cy3-labeled single-stranded DNA and antibody-modified gold nanoparticles (Cy3-ssDNA-Ab-AuNPs). The MMPs (2.5 μm in diameter) modified with mouse anti-human IgG monoclonal-antibodies could capture human IgG and further be separated and enriched via a magnetic field. The AuNPs (13 nm in diameter) conjugated with goat anti-human IgG polyclonal-antibodies and Cy3-ssDNA could further combine with the human IgG/Ab-MMP complex. The Cy3-ssDNA on AuNPs was then released by TCEP to hybridize with the DNA chip, thus generating a detectable signal by the fluorescence intensity of Cy3. In order to improve detection sensitivity, a three-level cascaded signal amplification was developed: (1) The MMP enrichment as the first-level; (2) Large quantities of Cy3-ssDNA on AuNPs as the second-level; (3) The Cy3-ssDNA conjugate with DNA chip as the third-level. The highly sensitive technique showed an increased response of the fluorescence intensity to the increased concentration of human IgG through a detection range from 1 pg mL −1 to 10 ng mL −1 . This sensing technique could not only improve the detection sensitivity for the low concentration of human IgG but also present a robust and efficient signal amplification model. The detection method has good stability, specificity, and reproducibility and could be applied in the detection of human IgG in the real samples

  7. Highly sensitive detection of human IgG using a novel bio-barcode assay combined with DNA chip technology

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhenbao [Central South University, School of Pharmaceutical Sciences (China); Zhou, Bo, E-mail: zhoubo1771@163.com [The Affiliated Zhongda Hospital of Southeast University, Department of Gerontology (China); Wang, Haiqing; Lu, Feng; Liu, Tianjun; Song, Cunxian; Leng, Xigang, E-mail: lengxigyky@163.com [Institute of Biomedical Engineering, Chinese Academy of Medical Sciences and Peking Union Medical College (China)

    2013-09-15

    A simple and ultrasensitive detection of human IgG based on signal amplification using a novel bio-barcode assay and DNA chip technology was developed. The sensing platform was a sandwich system made up of antibody-modified magnetic microparticles (Ab-MMPs)/human IgG/Cy3-labeled single-stranded DNA and antibody-modified gold nanoparticles (Cy3-ssDNA-Ab-AuNPs). The MMPs (2.5 {mu}m in diameter) modified with mouse anti-human IgG monoclonal-antibodies could capture human IgG and further be separated and enriched via a magnetic field. The AuNPs (13 nm in diameter) conjugated with goat anti-human IgG polyclonal-antibodies and Cy3-ssDNA could further combine with the human IgG/Ab-MMP complex. The Cy3-ssDNA on AuNPs was then released by TCEP to hybridize with the DNA chip, thus generating a detectable signal by the fluorescence intensity of Cy3. In order to improve detection sensitivity, a three-level cascaded signal amplification was developed: (1) The MMP enrichment as the first-level; (2) Large quantities of Cy3-ssDNA on AuNPs as the second-level; (3) The Cy3-ssDNA conjugate with DNA chip as the third-level. The highly sensitive technique showed an increased response of the fluorescence intensity to the increased concentration of human IgG through a detection range from 1 pg mL{sup -1} to 10 ng mL{sup -1}. This sensing technique could not only improve the detection sensitivity for the low concentration of human IgG but also present a robust and efficient signal amplification model. The detection method has good stability, specificity, and reproducibility and could be applied in the detection of human IgG in the real samples.

  8. Highly sensitive detection of human IgG using a novel bio-barcode assay combined with DNA chip technology

    Science.gov (United States)

    Liu, Zhenbao; Zhou, Bo; Wang, Haiqing; Lu, Feng; Liu, Tianjun; Song, Cunxian; Leng, Xigang

    2013-09-01

    A simple and ultrasensitive detection of human IgG based on signal amplification using a novel bio-barcode assay and DNA chip technology was developed. The sensing platform was a sandwich system made up of antibody-modified magnetic microparticles (Ab-MMPs)/human IgG/Cy3-labeled single-stranded DNA and antibody-modified gold nanoparticles (Cy3-ssDNA-Ab-AuNPs). The MMPs (2.5 μm in diameter) modified with mouse anti-human IgG monoclonal-antibodies could capture human IgG and further be separated and enriched via a magnetic field. The AuNPs (13 nm in diameter) conjugated with goat anti-human IgG polyclonal-antibodies and Cy3-ssDNA could further combine with the human IgG/Ab-MMP complex. The Cy3-ssDNA on AuNPs was then released by TCEP to hybridize with the DNA chip, thus generating a detectable signal by the fluorescence intensity of Cy3. In order to improve detection sensitivity, a three-level cascaded signal amplification was developed: (1) The MMP enrichment as the first-level; (2) Large quantities of Cy3-ssDNA on AuNPs as the second-level; (3) The Cy3-ssDNA conjugate with DNA chip as the third-level. The highly sensitive technique showed an increased response of the fluorescence intensity to the increased concentration of human IgG through a detection range from 1 pg mL-1 to 10 ng mL-1. This sensing technique could not only improve the detection sensitivity for the low concentration of human IgG but also present a robust and efficient signal amplification model. The detection method has good stability, specificity, and reproducibility and could be applied in the detection of human IgG in the real samples.

  9. Sensitive and specific detection of the non-human sialic Acid N-glycolylneuraminic acid in human tissues and biotherapeutic products.

    Directory of Open Access Journals (Sweden)

    Sandra L Diaz

    Full Text Available Humans are genetically defective in synthesizing the common mammalian sialic acid N-glycolylneuraminic acid (Neu5Gc, but can metabolically incorporate it from dietary sources (particularly red meat and milk into glycoproteins and glycolipids of human tumors, fetuses and some normal tissues. Metabolic incorporation of Neu5Gc from animal-derived cells and medium components also results in variable contamination of molecules and cells intended for human therapies. These Neu5Gc-incorporation phenomena are practically significant, because normal humans can have high levels of circulating anti-Neu5Gc antibodies. Thus, there is need for the sensitive and specific detection of Neu5Gc in human tissues and biotherapeutic products. Unlike monoclonal antibodies that recognize Neu5Gc only in the context of underlying structures, chicken immunoglobulin Y (IgY polyclonal antibodies can recognize Neu5Gc in broader contexts. However, prior preparations of such antibodies (including our own suffered from some non-specificity, as well as some cross-reactivity with the human sialic acid N-acetylneuraminic acid (Neu5Ac.We have developed a novel affinity method utilizing sequential columns of immobilized human and chimpanzee serum sialoglycoproteins, followed by specific elution from the latter column by free Neu5Gc. The resulting mono-specific antibody shows no staining in tissues or cells from mice with a human-like defect in Neu5Gc production. It allows sensitive and specific detection of Neu5Gc in all underlying glycan structural contexts studied, and is applicable to immunohistochemical, enzyme-linked immunosorbent assay (ELISA, Western blot and flow cytometry analyses. Non-immune chicken IgY is used as a reliable negative control. We show that these approaches allow sensitive detection of Neu5Gc in human tissue samples and in some biotherapeutic products, and finally show an example of how Neu5Gc might be eliminated from such products, by using a human cell

  10. Human umbilical vein: involvement of cyclooxygenase-2 pathway in bradykinin B1 receptor-sensitized responses.

    Science.gov (United States)

    Errasti, A E; Rey-Ares, V; Daray, F M; Rogines-Velo, M P; Sardi, S P; Paz, C; Podestá, E J; Rothlin, R P

    2001-08-01

    In isolated human umbilical vein (HUV), the contractile response to des-Arg9-bradykinin (des-Arg9-BK), selective BK B1 receptor agonist, increases as a function of the incubation time. Here, we evaluated whether cyclooxygenase (COX) pathway is involved in BK B1-sensitized response obtained in 5-h incubated HUV rings. The effect of different concentrations of indomethacin, sodium salicylate, ibuprofen, meloxicam, lysine clonixinate or NS-398 administrated 30 min before concentration-response curves (CRC) was studied. All treatments produced a significant rightward shift of the CRC to des-Arg9-BK in a concentration-dependent manner, which provides pharmacological evidence that COX pathway is involved in the BK B1 responses. Moreover, in this tissue, the NS-398 pKb (5.2) observed suggests that COX-2 pathway is the most relevant. The strong correlation between published pIC50 for COX-2 and the NSAIDs' pKbs estimated further supports the hypothesis that COX-2 metabolites are involved in BK B1 receptor-mediated responses. In other rings, indomethacin (30, 100 micromol/l) or NS-398 (10, 30 micromol/l) produced a significant rightward shift of the CRC to BK, selective BK B2 agonist, and its pKbs were similar to the values to inhibit BK B1 receptor responses, suggesting that COX-2 pathway also is involved in BK B2 receptor responses. Western blot analysis shows that COX-1 and COX-2 isoenzymes are present before and after 5-h in vitro incubation and apparently COX-2 does not suffer additional induction.

  11. Sensitive spectrophotometric determination of metoclopramide hydrochloride in dosage forms and spiked human urine using vanillin

    Directory of Open Access Journals (Sweden)

    O. Zenita Devi

    2016-09-01

    Full Text Available A new spectrophotometric method which is simple, sensitive, selective and rapid is described for the determination of metoclopramide hydrochloride (MCP in bulk drug and in dosage forms using vanillin as the chromogenic agent. The method is based on the condensation reaction between primary aromatic amine group present in MCP with aromatic aldehyde, vanillin to produce an intense yellow colored product. The resulting Schiff’s base shows an absorption maximum at 410 nm and the reaction product is stable for more than one day. The reaction was carried out in acetic acid and perchloric acid medium. Beer’s law was obeyed in the concentration range 1.5–15.0 μg ml−1 MCP with a molar absorptivity of 1.89 × 104 l mol−1 cm−1. The limit of detection (LOD and limit of quantification (LOQ were found to be 0.51 and 1.55 μg ml−1, respectively. The method was statistically evaluated by calculating percent relative error (% RE for accuracy and percent relative standard deviation (% RSD for precision, and was applied successfully to the determination of MCP in tablets, in injection and also in spiked human urine. No interference was observed from common additives found in pharmaceutical preparations. The results obtained by the proposed method were validated statistically by comparing the results with those of the reference method by applying the Student’s t-test and F-test. The accuracy and reliability of the method were further ascertained by performing recovery tests via standard-addition technique.

  12. Sensitive microculture method for isolation of human immunodeficiency virus type 1 from blood leukocytes.

    Science.gov (United States)

    Erice, A; Sannerud, K J; Leske, V L; Aeppli, D; Balfour, H H

    1992-02-01

    A study was conducted to compare our standard culture with a new microculture procedure for isolation of human immunodeficiency virus type 1 (HIV-1) from blood leukocytes. A total of 137 blood specimens from 102 HIV-1 antibody-positive individuals (52 were asymptomatic, 31 were symptomatic, and 19 had AIDS) were cultured in a microculture system in which 10(6) of the patients' peripheral blood mononuclear cells (PBMC) were cocultured with 10(6) phytohemagglutinin (PHA)-stimulated PBMC from an HIV-1 antibody-negative blood donor in 1.2 ml of culture medium. Results were compared with those of a historical control group of 139 standard HIV-1 cultures from 108 HIV-1 antibody-positive subjects (58 were asymptomatic, 36 were symptomatic, and 14 had AIDS). For standard cultures, 10 x 10(6) of the patients' PBMC were cocultured with 5 x 10(6) PHA-stimulated PBMC from an HIV-1 antibody-negative blood donor in 15 ml of culture medium. HIV-1 was isolated in 128 (93%) microcultures and 133 (96%) standard cultures. Both methods identified more than 75% of the positive cultures within 7 days and 100% of the positive cultures within 14 days. The isolation rates for HIV-1 in microcultures compared with standard cultures were 91 versus 93% (specimens from asymptomatic individuals), 93 versus 96% (specimens from symptomatic individuals), and 97 versus 100% (specimens from patients with AIDS). The median time to positivity for both culture methods was 7 days, and this correlated significantly with symptoms and CD4+ cell counts. The microculture method is a sensitive and less expensive system for isolation of HIV-1 from PBMC of HIV-1 antibody-positive individuals, and we recommend it as the culture method of choice, especially for children and patients with AIDS and severe anemia or leukopenia whose blood volume is an important consideration.

  13. An automated system for the objective evaluation of human gustatory sensitivity using tongue biopotential recordings.

    Directory of Open Access Journals (Sweden)

    Danilo Pani

    Full Text Available The goal of this work is to develop an automatic system for the evaluation of the gustatory sensitivity of patients using an electrophysiological recording of the response of bud cells to taste stimuli. In particular, the study aims to evaluate the effectiveness and limitations of supervised classifiers in the discrimination between subjects belonging to the three 6-n-propylthiouracil (PROP taster categories (supertasters, medium tasters, and non-tasters, exploiting features extracted from electrophysiological recordings of the tongue. Thirty-nine subjects (equally divided into the three PROP status classes by standard non-objective scaling methods underwent a non-invasive, differential, biopotential recording of their tongues during stimulation with PROP by using a custom-made, flexible, silver electrode. Two different classifiers were trained to recognize up to seven different features extracted from the recorded depolarization signal. The classification results indicate that the identified set of features allows to distinguish between PROP tasters and non-tasters (average accuracy of 80% ± 18% and up to 94% ± 15% when only supertasters and non-tasters are considered, but medium tasters were difficult to identify. However, these apparent classification errors are related to uncertainty in the labeling procedures, which are based on non-objective tests, in which the subjects provided borderline evaluations. Thus, using the proposed method, it is possible, for the first time, to automatically achieve objective PROP taster status identification with high accuracy. The simplicity of the recording technique allows for easy reproduction of the experimental setting; thus the technique can be used in future studies to evaluate other gustatory stimuli. The proposed approach represents the first objective and automatic method to directly measure human gustatory responses and a milestone for physiological taste studies, with applications ranging from

  14. [Knockdown of ATG5 enhances the sensitivity of human renal carcinoma cells to sunitinib].

    Science.gov (United States)

    Li, Peng; Han, Qi; Tang, Ming; Zhang, Keqin

    2017-03-01

    Objective To investigate the expression levels of autophagy-related gene 5 (ATG5) and microtubule-associated protein 1 light chain 3 (LC3) and their effects on sunitinib resistance in human renal carcinoma cells. Methods After clinic-pathologic feature and survival analysis, 99 renal clear cell carcinoma tissues with different histological grades were used to detect the expression of ATG5 and LC3 by immunohistochemistry. Renal carcinoma cell line A-498 was infected with lentivirus-mediated ATG5 shRNA. Western blot analysis was performed to confirm the efficiency of ATG5 knockdown. Proliferation rate of A-498 cells in control group and ATG5 low expression group was determined by flow cytometry. Finally, the survival rate was detected by MTT assay after A-498 cells were treated with different concentrations of sunitinib. Results The expression levels of ATG5 and LC3 in renal clear cell carcinoma tissues were significantly higher than those in para-tumor tissues. The expression levels of ATG5 and LC3 were associated with classification, histological grade, TNM stage and survival rate, rather than gender, age, location, tumor size. Compared with the control group, the protein expressions of ATG5 and LC3 significantly decreased in A-498 cells with ATG5 low expression. The cell proliferation rate in ATG5 downregulation group was lower than that in the control group. Compared with control group, the survival rate in ATG5 low expression group were significantly reduced in a dose-dependent manner after sunitinib treatment. Conclusion Autophagy is active in renal clear cell carcinoma, and the drug sensitivity to sunitinib in renal cancer cells can be enhanced by the downregulation of ATG5.

  15. Insulin sensitivity is independent of lipid binding protein trafficking at the plasma membrane in human skeletal muscle

    DEFF Research Database (Denmark)

    Jordy, Andreas Børsting; Serup, Annette Karen; Karstoft, Kristian

    2014-01-01

    The aim of the present study was to investigate lipid-induced regulation of lipid binding proteins in human skeletal muscle and the impact hereof on insulin sensitivity. Eleven healthy male subjects underwent a 3-day hyper-caloric and high-fat diet regime. Muscle biopsies were taken before......-regulated by increased fatty acid availability. This suggests a time dependency in the up-regulation of FAT/CD36 and FABPpm protein during high availability of plasma fatty acids. Furthermore, we did not detect FATP1 and FATP4 protein in giant sarcolemmal vesicles obtained from human skeletal muscle. In conclusion......, this study shows that a short-term lipid-load increases mRNA content of key lipid handling proteins in human muscle. However, decreased insulin sensitivity after high-fat diet is not accompanied with relocation of FAT/CD36 or FABPpm protein to the sarcolemma. Finally, FATP1 and FATP4 protein could...

  16. An ancestral haplotype of the human PERIOD2 gene associates with reduced sensitivity to light-induced melatonin suppression.

    Directory of Open Access Journals (Sweden)

    Tokiho Akiyama

    Full Text Available Humans show various responses to the environmental stimulus in individual levels as "physiological variations." However, it has been unclear if these are caused by genetic variations. In this study, we examined the association between the physiological variation of response to light-stimulus and genetic polymorphisms. We collected physiological data from 43 subjects, including light-induced melatonin suppression, and performed haplotype analyses on the clock genes, PER2 and PER3, exhibiting geographical differentiation of allele frequencies. Among the haplotypes of PER3, no significant difference in light sensitivity was found. However, three common haplotypes of PER2 accounted for more than 96% of the chromosomes in subjects, and 1 of those 3 had a significantly low-sensitive response to light-stimulus (P < 0.05. The homozygote of the low-sensitive PER2 haplotype showed significantly lower percentages of melatonin suppression (P < 0.05, and the heterozygotes of the haplotypes varied their ratios, indicating that the physiological variation for light-sensitivity is evidently related to the PER2 polymorphism. Compared with global haplotype frequencies, the haplotype with a low-sensitive response was more frequent in Africans than in non-Africans, and came to the root in the phylogenetic tree, suggesting that the low light-sensitive haplotype is the ancestral type, whereas the other haplotypes with high sensitivity to light are the derived types. Hence, we speculate that the high light-sensitive haplotypes have spread throughout the world after the Out-of-Africa migration of modern humans.

  17. Enhanced expression of IL-8 in normal human keratinocytes and human keratinocyte cell line HaCaT in vitro after stimulation with contact sensitizers, tolerogens and irritants.

    Science.gov (United States)

    Mohamadzadeh, M; Müller, M; Hultsch, T; Enk, A; Saloga, J; Knop, J

    1994-12-01

    To investigate the interleukin-8 production of keratinocytes after stimulation in vitro we have used various agents: (i) contact sensitizer (2,4-dinitrofluorobenzene, 3-n-pentadecylcatechol); (ii) tolerogen (5-methyl-3-n-pentadecylcatechol); (iii) irritant (sodium lauryl sulfate). Interleukin-8 gene expression was assessed by northern blot hybridization of the total cytoplasmic RNA extracted from subconfluent normal human keratinocyte cultures and the keratinocyte cell line HaCaT using a radiolabeled DNA probe specific for human interleukin-8. Interleukin-8 gene expression was markedly increased upon in vitro stimulation after 1-6 h with contact sensitizers, tolerogen and the irritant. In contrast, interleukin-8 production was not detectable in unstimulated normal human keratinocytes or the HaCaT keratinocyte cell line. These results suggest that the induction and production of interleukin-8 is a response to nonspecific stimuli and may play a critical role in the early response to immunogenic or inflammatory signals in man.

  18. [Variability of the sensitivity of human lymphocytes to the antiproliferative action of alkylating agents].

    Science.gov (United States)

    Veremko, L N; Telegin, L Iu; Pevnitskii, L A

    1983-05-01

    A study was made of variability of the sensitivity of peripheral blood lymphocytes from different donors to an antiproliferative action of cyclophosphamide and thiophosphamide. A similar degree of the sensitivity was revealed to alkylating agents differing in the action mode, with this degree being independent of the "stimulation index" magnitude.

  19. Inhibition of c-Kit signaling is associated with reduced heat and cold pain sensitivity in humans.

    Science.gov (United States)

    Ceko, Marta; Milenkovic, Nevena; le Coutre, Philipp; Westermann, Jörg; Lewin, Gary R

    2014-07-01

    The tyrosine kinase receptor c-Kit is critically involved in the modulation of nociceptive sensitivity in mice. Ablation of the c-Kit gene results in hyposensitivity to thermal pain, whereas activation of c-Kit produces hypersensitivity to noxious heat, without altering sensitivity to innocuous mechanical stimuli. In this study, we investigated the role of c-Kit signaling in human pain perception. We hypothesized that subjects treated with Imatinib or Nilotinib, potent inhibitors of tyrosine kinases including c-Kit but also Abl1, PDFGFRα, and PDFGFRβ, that are used to treat chronic myeloid leukemia (CML), would experience changes in thermal pain sensitivity. We examined 31 asymptomatic CML patients (14 male and 17 female) receiving Imatinib/Nilotinib treatment and compared them to 39 age- and sex-matched healthy controls (12 male and 27 female). We used cutaneous heat and cold stimulation to test normal and noxious thermal sensitivity, and a grating orientation task to assess tactile acuity. Thermal pain thresholds were significantly increased in the Imatinib/Nilotinib-treated group, whereas innocuous thermal and tactile thresholds were unchanged compared to those in the control group. In conclusion, our findings suggest that the biological effects of c-Kit inhibition are comparable in mice and humans in that c-Kit activity is required to regulate thermal pain sensitivity but does not affect innocuous thermal and mechanical sensation. The effect on experimental heat pain observed in our study is comparable to those of several common analgesics; thus modulation of the c-Kit pathway can be used to specifically modulate noxious heat and cold sensitivity in humans. Copyright © 2014 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  20. Lysophosphatidylcholine hydrolases of human erythrocytes, lymphocytes, and brain: Sensitive targets of conserved specificity for organophosphorus delayed neurotoxicants

    International Nuclear Information System (INIS)

    Vose, Sarah C.; Holland, Nina T.; Eskenazi, Brenda; Casida, John E.

    2007-01-01

    Brain neuropathy target esterase (NTE), associated with organophosphorus (OP)-induced delayed neuropathy, has the same OP inhibitor sensitivity and specificity profiles assayed in the classical way (paraoxon-resistant, mipafox-sensitive hydrolysis of phenyl valerate) or with lysophosphatidylcholine (LysoPC) as the substrate. Extending our earlier observation with mice, we now examine human erythrocyte, lymphocyte, and brain LysoPC hydrolases as possible sensitive targets for OP delayed neurotoxicants and insecticides. Inhibitor profiling of human erythrocytes and lymphocytes gave the surprising result of essentially the same pattern as with brain. Human erythrocyte LysoPC hydrolases are highly sensitive to OP delayed neurotoxicants, with in vitro IC 50 values of 0.13-85 nM for longer alkyl analogs, and poorly sensitive to the current OP insecticides. In agricultural workers, erythrocyte LysoPC hydrolyzing activities are similar for newborn children and their mothers and do not vary with paraoxonase status but have high intersample variation that limits their use as a biomarker. Mouse erythrocyte LysoPC hydrolase activity is also of low sensitivity in vitro and in vivo to the OP insecticides whereas the delayed neurotoxicant ethyl n-octylphosphonyl fluoride inhibits activity in vivo at 1-3 mg/kg. Overall, inhibition of blood LysoPC hydrolases is as good as inhibition of brain NTE as a predictor of OP inducers of delayed neuropathy. NTE and lysophospholipases (LysoPLAs) both hydrolyze LysoPC, yet they are in distinct enzyme families with no sequence homology and very different catalytic sites. The relative contributions of NTE and LysoPLAs to LysoPC hydrolysis and clearance from erythrocytes, lymphocytes, and brain remain to be defined

  1. Central sensitization in spinal cord injured humans assessed by reflex receptive fields

    DEFF Research Database (Denmark)

    Biurrun Manresa, José Alberto; Finnerup, Nanna Susanne Brix; Johannesen, Inger Lauge

    2014-01-01

    OBJECTIVE: To investigate the effects of central sensitization, elicited by intramuscular injection of capsaicin, by comparing the reflex receptive fields (RRF) of spinally-intact volunteers and spinal cord injured volunteers that present presensitized spinal nociceptive mechanisms. METHODS...... after an intramuscular injection of capsaicin in the foot sole in order to induce central sensitization. RESULTS: Both groups presented RRF expansion and lowered NWR thresholds immediately after capsaicin injection, reflected by the enlargement of RRF sensitivity areas and RRF probability areas....... Moreover, the topography of the RRF sensitivity and probability areas were significantly different in SCI volunteers compared to NI volunteers in terms of size and shape. CONCLUSIONS: SCI volunteers can develop central sensitization, despite adaptive/maladaptive changes in synaptic plasticity and lack...

  2. A conceptual framework for predicting temperate ecosystem sensitivity to human impacts on fire regimes

    Science.gov (United States)

    D. B. McWethy; P. E. Higuera; C. Whitlock; T. T. Veblen; D. M. J. S. Bowman; G. J. Cary; S. G. Haberle; R. E. Keane; B. D. Maxwell; M. S. McGlone; G. L. W. Perry; J. M. Wilmshurst

    2013-01-01

    The increased incidence of large fires around much of the world in recent decades raises questions about human and non-human drivers of fire and the likelihood of increased fire activity in the future. The purpose of this paper is to outline a conceptual framework for examining where human-set fires and feedbacks are likely to be most pronounced in temperate forests...

  3. Vitamins C and K3 sensitize human urothelial tumors to gemcitabine.

    Science.gov (United States)

    Kassouf, Wassim; Highshaw, Ralph; Nelkin, Gina M; Dinney, Colin P; Kamat, Ashish M

    2006-10-01

    We evaluated the antitumor effects of vitamins C and K3 for human urothelial carcinoma and the potential use of the combination of vitamins C plus K3 as a sensitizing agent for conventional chemotherapy for urothelial carcinoma. The antiproliferative and apoptotic effects of vitamin C alone, vitamin K3 alone, vitamins C plus K3, gemcitabine alone and gemcitabine plus vitamins C plus K3 were assessed in vitro by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, propidium iodide staining and flow cytometry. For in vivo studies we implanted UMUC-14 tumorigenic urothelial carcinoma cells into the subcutis of nude mice. One week later we treated 10 mice each with saline (control), vitamins C plus K3, gemcitabine or gemcitabine plus vitamins C plus K3. Treatment was continued for 4 weeks, followed by necropsy. Tumor volume was measured and tumor kinetics were established. Apoptosis and proliferation were evaluated in tumor sections using immunohistochemistry and TUNEL assay. Vitamins C plus K3 induced cytostasis and caused apoptosis to a greater degree than either vitamin alone (p Vitamins C plus K3 also substantially augmented the effects of gemcitabine in vitro. There were 32.3% apoptosis with gemcitabine plus vitamins C plus K3, 5.3% with gemcitabine alone and 15.8% with vitamins C plus K3 alone (p vitamins C plus K3 compared with that in the control or for either agent alone. Mean tumor weight and growth rate in the gemcitabine plus vitamins C plus K3 group (237 mg and 11.3 mm3 daily) were decreased compared with those in the control (530 mg and 34.3 mm3 daily), and those for vitamins C plus K3 alone (490 mg and 25.2 mm3 daily) and gemcitabine alone (400 mg and 21.3 mm3 daily) (p Vitamins C and K3 have significant antiproliferative and apoptotic effects when used in combination. This combination enhances the efficacy of gemcitabine against bladder cancer in vivo.

  4. Sensitive quantification of apomorphine in human plasma using a LC-ESI-MS-MS method.

    Science.gov (United States)

    Abe, Emuri; Alvarez, Jean-Claude

    2006-06-01

    An analytical method based on liquid chromatography coupled with ion trap mass spectrometry (MS) detection with electrospray ionization interface has been developed for the identification and quantification of apomorphine in human plasma. Apomorphine was isolated from 0.5 mL of plasma using a liquid-liquid extraction with diethyl ether and boldine as internal standard, with satisfactory extraction recoveries. Analytes were separated on a 5-microm C18 Highpurity (Thermohypersil) column (150 mm x 2.1 mm I.D.) maintained at 30 degrees C, coupled to a precolumn (C18, 5-microm, 10 mm x 2.0 mm I.D., Thermo). The elution was achieved isocratically with a mobile phase of 2 mM NH4COOH buffer pH 3.8/acetonitrile (50/50, vol/vol) at a flow rate of 200 microL per minute. Data were collected either in full-scan MS mode at m/z 150 to 500 or in full-scan tandem mass spectrometry mode, selecting the [M+H]ion at m/z 268.0 for apomorphine and m/z 328.0 for boldine. The most intense daughter ion of apomorphine (m/z 237.1) and boldine (m/z 297.0) were used for quantification. Retention times were 2.03 and 2.11 minutes for boldine and apomorphine, respectively. Calibration curves were linear in the 0.025 to 20 ng/mL range. The limits of detection and quantification were 0.010 ng/mL and 0.025 ng/mL, respectively. Accuracy and precision of the assay were measured by analyzing 54 quality control samples for 3 days. At concentrations of 0.075, 1.5, and 15 ng/mL, intraday precisions were less than 10.1%, 5.3%, and 3.8%, and interday precisions were less than 4.8%, 6.6%, and 6.5%, respectively. Accuracies were in the 99.5 to 104.2% range. An example of a patient who was given 6 mg of apomorphine subcutaneously is shown, with concentrations of 14.1 ng/mL after 30 minutes and 0.20 ng/mL after 6 hours. The method described enables the unambiguous identification and quantification of apomorphine with very good sensitivity using only 0.5 mL of sample, and is very convenient for therapeutic drug

  5. Accounting for human variability and sensitivity in setting standards for electromagnetic fields.

    Science.gov (United States)

    Bailey, William H; Erdreich, Linda S

    2007-06-01

    Biological sensitivity and variability are key issues for risk assessment and standard setting. Variability encompasses general inter-individual variations in population responses, while sensitivity relates to unusual or extreme responses based on genetic, congenital, medical, or environmental conditions. For risk assessment and standard setting, these factors affect estimates of thresholds for effects and dose-response relationships and inform efforts to protect the more sensitive members of the population, not just the typical or average person. While issues of variability and sensitivity can be addressed by experimental and clinical studies of electromagnetic fields, investigators have paid little attention to these important issues. This paper provides examples that illustrate how default assumptions regarding variability can be incorporated into estimates of 60-Hz magnetic field exposures with no risk of cardiac stimulation and how population thresholds and variability of peripheral nerve stimulation responses at 60-Hz can be estimated from studies of pulsed gradient magnetic fields in magnetic resonance imaging studies. In the setting of standards for radiofrequency exposures, the International Commission for Non-Ionizing Radiation Protection uses inter-individual differences in thermal sensitivity as one of the considerations in the development of "safety factors." However, neither the range of sensitivity nor the sufficiency or excess of the 10-fold and the additional 5-fold safety factors have been assessed quantitatively. Data on the range of responses between median and sensitive individuals regarding heat stress and cognitive function should be evaluated to inform a reassessment of these safety factors and to identify data gaps.

  6. Nasal insulin changes peripheral insulin sensitivity simultaneously with altered activity in homeostatic and reward-related human brain regions.

    Science.gov (United States)

    Heni, M; Kullmann, S; Ketterer, C; Guthoff, M; Linder, K; Wagner, R; Stingl, K T; Veit, R; Staiger, H; Häring, H-U; Preissl, H; Fritsche, A

    2012-06-01

    Impaired insulin sensitivity is a major factor leading to type 2 diabetes. Animal studies suggest that the brain is involved in the regulation of insulin sensitivity. We investigated whether insulin action in the human brain regulates peripheral insulin sensitivity and examined which brain areas are involved. Insulin and placebo were given intranasally. Plasma glucose, insulin and C-peptide were measured in 103 participants at 0, 30 and 60 min. A subgroup (n = 12) was also studied with functional MRI, and blood sampling at 0, 30 and 120 min. For each time-point, the HOMA of insulin resistance (HOMA-IR) was calculated as an inverse estimate of peripheral insulin sensitivity. Plasma insulin increased and subsequently decreased. This excursion was accompanied by slightly decreased plasma glucose, resulting in an initially increased HOMA-IR. At 1 h after insulin spray, the HOMA-IR subsequently decreased and remained lower up to 120 min. An increase in hypothalamic activity was observed, which correlated with the increased HOMA-IR at 30 min post-spray. Activity in the putamen, right insula and orbitofrontal cortex correlated with the decreased HOMA-IR at 120 min post-spray. Central insulin action in specific brain areas, including the hypothalamus, may time-dependently regulate peripheral insulin sensitivity. This introduces a potential novel mechanism for the regulation of peripheral insulin sensitivity and underlines the importance of cerebral insulin action for the whole organism.

  7. Repair of closely opposed cyclobutyl pyrimidine dimers in UV-sensitive human diploid fibroblasts

    International Nuclear Information System (INIS)

    Lam, L.H.; Reynolds, R.J.

    1986-01-01

    An enzyme-sensitive site assay has been used to examine the fate of closely opposed pyrimidine dimers in fibroblasts from individuals afflicted with various genetic disorders that confer increased cellular sensitivity to UV radiation. The disappearance of bifilar enzyme-sensitive sites was found to be normal in cells from individuals with Fanconi's anemia, Cockayne's syndrome, dyskeratosis congenita and the variant form of xeroderma pigmentosum. The rate of bifilar enzyme-sensitive site removal in XP cells assigned to complementation group C was reduced by an amount similar to that observed for the repair of isolated dimers. Our results indicate that the initiation of repair at closely opposed dimers is slow in XP-C cells but normal in all other cells examined. (Auth.)

  8. Effects of Lactobacillus acidophilus NCFM on insulin sensitivity and the systemic inflammatory response in human subjects

    DEFF Research Database (Denmark)

    Andreasen, Anne Sofie; Larsen, Nadja; Pedersen-Skovsgaard, Theis

    2010-01-01

    According to animal studies, intake of probiotic bacteria may improve glucose homeostasis. We hypothesised that probiotic bacteria improve insulin sensitivity by attenuating systemic inflammation. Therefore, the effects of oral supplementation with the probiotic bacterium Lactobacillus acidophilus...

  9. Retained sensitivity to cytotoxic pyrimidine nucleoside analogs in thymidine kinase 2 deficient human fibroblasts

    OpenAIRE

    Bjerke, Mia; Solaroli, Nicola; Lesko, Nicole; Balzarini, Jan; Johansson, Magnus; Karlsson, Anna

    2010-01-01

    Thymidine kinase 2 (TK2) is a mitochondrial deoxyribonucleoside kinase that phosphorylates several nucleoside analogs used in anti-viral and anti-cancer therapy. A fibroblast cell line with decreased TK2 activity was investigated in order to obtain insights in the effects of TK2 deficiency on nucleotide metabolism. The role of TK2 for the sensitivity against cytotoxic nucleoside analogs was also investigated. The TK2 deficient cells retained their sensitivity against all pyrimidine nucleoside...

  10. Diagnostic sensitivity of two radio receptor assays (TRAK Assay and TRAK Dyno human) for the detection of TSH receptor antibodies

    International Nuclear Information System (INIS)

    Paunkovic, N.; Paunkovic, J.

    2003-01-01

    Radio receptor assays for the detection of TSH receptor antibodies in serum are typically based on binding the competition of TSH-R antibodies and 125I -labelled-TSH for membrane preparation of thyrocytes (TBII tests). The sensitivity of the available tests utilizing porcine cell membranes was found to be around 80%. A new test (TRAK Dyno human, BRAHMS) utilizes human recombinant TSH receptor and human standard material that is supposed to improve the performance of the test. We have compared the results of these two assays. The sensitivity of the TRAK Assay tested in 356 patients with untreated Grave's disease was found to be 85%, and 97.5% for TRAK Dyno human in 111 newly diagnosed patients. Both tests were performed from the same serum specimen for 60 of the investigated patients. The TRAK Assay was positive in 50 patients (83.2%) and TRAK Dyno human in 59 patients (98.3%). The specificity of the new radio receptor assay was also improved. (author)

  11. Syringic acid from Tamarix aucheriana possesses antimitogenic and chemo-sensitizing activities in human colorectal cancer cells.

    Science.gov (United States)

    Abaza, Mohamed-Salah; Al-Attiyah, Raja'a; Bhardwaj, Radhika; Abbadi, Ghaneim; Koyippally, Mathew; Afzal, Mohammad

    2013-09-01

    For its variety of biological activities, Tamarix aucheriana (Decne.) Baum. (Tamaricaceae) has an extensive history as a traditional Arab medicine. Antimitogenic and chemo-sensitizing activities of syringic acid (SA) were studied against human colorectal cancer. Chromatographic and spectral data were used for the isolation and identification of SA. MTT, flow cytometry, in vitro invasion and angiogenesis assays, fluoremetry, ELISA and Real Time qPCR were used to test antimitogenic and chemo-sensitizing activities of SA, cell cycle, apoptosis, proteasome and NFκB-DNA-binding activities, cancer cell invasion and angiogenesis, and expression of cell cycle/apoptosis-related genes. SA showed a time- and dose-dependent (IC₅₀ = 0.95-1.2 mg mL⁻¹) antimitogenic effect against cancer cells with little cytotoxicity on normal fibroblasts (≤20%). SA-altered cell cycle (S/G2-M or G1/G2-M phases) in a time-dependent manner, induced apoptosis, inhibited DNA-binding activity of NFκB (p ≤ 0.0001), chymotrypsin-like/PGPH (peptidyl-glutamyl peptide-hydrolyzing) (p ≤ 0.0001) and the trypsin-like (p ≤ 0.002) activities of 26S proteasome and angiogenesis. SA also differentially sensitized cancer cells to standard chemotherapies with a marked increase in their sensitivity to camptothecin (500-fold), 5FU (20,000-fold), doxorubicin (210-fold), taxol (3134-fold), vinblastine (1000-fold), vincristine (130-fold) and amsacrine (107-fold) compared to standard drugs alone. SA exerted its chemotherapeutic and chemo-sensitizing effects through an array of mechanisms including cell-cycle arrest, apoptosis induction, inhibition of cell proliferation, cell migration, angiogenesis, NFκB DNA-binding and proteasome activities. These results demonstrate the potential of SA as an antimitogenic and chemo-sensitizing agent for human colorectal cancer.

  12. pH-sensitive intracellular photoluminescence of carbon nanotube-fluorescein conjugates in human ovarian cancer cells

    International Nuclear Information System (INIS)

    Chen, M T; Ishikawa, F N; Gundersen, M A; Gomez, L M; Vernier, P T; Zhou, C

    2009-01-01

    To add to the understanding of the properties of functionalized carbon nanotubes in biological applications, we report a monotonic pH sensitivity of the intracellular fluorescence emission of single-walled carbon nanotube-fluorescein carbazide (SWCNT-FC) conjugates in human ovarian cancer cells. Light-stimulated intracellular hydrolysis of the amide linkage and localized intracellular pH changes are proposed as mechanisms. SWCNT-FC conjugates may serve as intracellular pH sensors.

  13. Endocrine sensitivity of the receptor-positive T61 human breast carcinoma serially grown in nude mice

    DEFF Research Database (Denmark)

    Brünner, N; Spang-Thomsen, M; Skovgaard Poulsen, H

    1985-01-01

    A study was made on the effect of ovariectomy, 17 beta-oestradiol, and tamoxifen on the oestrogen and progesterone receptor-positive T61 human breast carcinoma grown in nude mice. The effect of the treatment was evaluated by the specific growth delay calculated on the basis of Gompertz growth cur...... but is not a sufficiently clear marker to allow prediction of the endocrine sensitivity of individual breast tumours....

  14. Sensitization of human carcinoma cells to alkylating agents by small interfering RNA suppression of 3-alkyladenine-DNA glycosylase.

    Science.gov (United States)

    Paik, Johanna; Duncan, Tod; Lindahl, Tomas; Sedgwick, Barbara

    2005-11-15

    One of the major cytotoxic lesions generated by alkylating agents is DNA 3-alkyladenine, which can be excised by 3-alkyladenine DNA glycosylase (AAG). Inhibition of AAG may therefore result in increased cellular sensitivity to chemotherapeutic alkylating agents. To investigate this possibility, we have examined the role of AAG in protecting human tumor cells against such agents. Plasmids that express small interfering RNAs targeted to two different regions of AAG mRNA were transfected into HeLa cervical carcinoma cells and A2780-SCA ovarian carcinoma cells. Stable derivatives of both cell types with low AAG protein levels were sensitized to alkylating agents. Two HeLa cell lines with AAG protein levels reduced by at least 80% to 90% displayed a 5- to 10-fold increase in sensitivity to methyl methanesulfonate, N-methyl-N-nitrosourea, and the chemotherapeutic drugs temozolomide and 1,3-bis(2-chloroethyl)-1-nitrosourea. These cells showed no increase in sensitivity to UV light or ionizing radiation. After treatment with methyl methanesulfonate, AAG knockdown HeLa cells were delayed in S phase but accumulated in G2-M. Our data support the hypothesis that ablation of AAG activity in human tumor cells may provide a useful strategy to enhance the efficacy of current chemotherapeutic regimens that include alkylating agents.

  15. Differentiated human midbrain-derived neural progenitor cells express excitatory strychnine-sensitive glycine receptors containing α2β subunits.

    Directory of Open Access Journals (Sweden)

    Florian Wegner

    Full Text Available BACKGROUND: Human fetal midbrain-derived neural progenitor cells (NPCs may deliver a tissue source for drug screening and regenerative cell therapy to treat Parkinson's disease. While glutamate and GABA(A receptors play an important role in neurogenesis, the involvement of glycine receptors during human neurogenesis and dopaminergic differentiation as well as their molecular and functional characteristics in NPCs are largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: Here we investigated NPCs in respect to their glycine receptor function and subunit expression using electrophysiology, calcium imaging, immunocytochemistry, and quantitative real-time PCR. Whole-cell recordings demonstrate the ability of NPCs to express functional strychnine-sensitive glycine receptors after differentiation for 3 weeks in vitro. Pharmacological and molecular analyses indicate a predominance of glycine receptor heteromers containing α2β subunits. Intracellular calcium measurements of differentiated NPCs suggest that glycine evokes depolarisations mediated by strychnine-sensitive glycine receptors and not by D-serine-sensitive excitatory glycine receptors. Culturing NPCs with additional glycine, the glycine-receptor antagonist strychnine, or the Na(+-K(+-Cl(- co-transporter 1 (NKCC1-inhibitor bumetanide did not significantly influence cell proliferation and differentiation in vitro. CONCLUSIONS/SIGNIFICANCE: These data indicate that NPCs derived from human fetal midbrain tissue acquire essential glycine receptor properties during neuronal maturation. However, glycine receptors seem to have a limited functional impact on neurogenesis and dopaminergic differentiation of NPCs in vitro.

  16. In vivo imaging of human oral hard and soft tissues by polarization-sensitive optical coherence tomography

    Science.gov (United States)

    Walther, Julia; Golde, Jonas; Kirsten, Lars; Tetschke, Florian; Hempel, Franz; Rosenauer, Tobias; Hannig, Christian; Koch, Edmund

    2017-12-01

    Since optical coherence tomography (OCT) provides three-dimensional high-resolution images of biological tissue, the benefit of polarization contrast in the field of dentistry is highlighted in this study. Polarization-sensitive OCT (PS OCT) with phase-sensitive recording is used for imaging dental and mucosal tissues in the human oral cavity in vivo. An enhanced polarization contrast of oral structures is reached by analyzing the signals of the co- and crosspolarized channels of the swept source PS OCT system quantitatively with respect to reflectivity, retardation, optic axis orientation, and depolarization. The calculation of these polarization parameters enables a high tissue-specific contrast imaging for the detailed physical interpretation of human oral hard and soft tissues. For the proof-of-principle, imaging of composite restorations and mineralization defects at premolars as well as gingival, lingual, and labial oral mucosa was performed in vivo within the anterior oral cavity. The achieved contrast-enhanced results of the investigated human oral tissues by means of polarization-sensitive imaging are evaluated by the comparison with conventional intensity-based OCT.

  17. Omalizumab Increases the Intrinsic Sensitivity of Human Basophils to IgE-Mediated Stimulation

    Science.gov (United States)

    MacGlashan, Donald; Saini, Sarbjit S.

    2013-01-01

    Background Treatment of allergic patients with omalizumab results in a paradoxical increase in their basophil histamine release response, ex vivo, to crosslinking anti-IgE antibody. It is not known whether this change in response is associated with an increase in intrinsic cellular sensitivity, which would be a paradoxical response. Objective To determine if the increase in response to anti-IgE Ab is a reflection of an increased cellular sensitivity, expressed as molecules of antigen-specific IgE per basophil required to produce a 50% of maximal response. Methods Patients were treated with omalizumab or placebo agent for 12 weeks (NCT01003301 at ClinicalTrials.gov) and the metric of basophil sensitivity was assessed at 4 time points, baseline, 6–8 weeks, 12 weeks (after which treatment stopped) and 24 weeks (12 weeks after the end of treatment). Results As observed previously, treatment with omalizumab resulted in a marked increase in the maximal histamine release induced by crosslinking anti-IgE Ab. This change was accompanied by a marked shift in intrinsic basophil sensitivity, ranging from 2.5 to 125 fold, with an average of 6 fold at the midpoint of the treatment to 12 fold after 12 weeks. The magnitude of the increase in cellular sensitivity was inversely related to the starting sensitivity or the starting maximum histamine release. The increased cellular sensitivity also occurred when using LTC4 secretion as a metric of the basophil response. 12 weeks after the end of treatment, cellular sensitivity was found to shift towards the baseline level although the return to baseline was not yet complete at this time point. Conclusions Treatment with omalizumab results in a markedly increased sensitivity of basophils to IgE-mediated stimulation, in terms of the number of IgE molecules required to produce a given response. These results provide a better quantitative sense of the phenotypic change that occurs in basophils during omalizumab treatment which has

  18. Sexual dimorphism in hepatic, adipose tissue and peripheral tissue insulin sensitivity in obese humans

    Directory of Open Access Journals (Sweden)

    Kasper W. ter Horst

    2015-11-01

    Full Text Available Glucose and lipid metabolism differ between men and women, and women tend to have better whole-body or muscle insulin sensitivity. This may be explained, in part, by differences in sex hormones and adipose tissue distribution. Few studies have investigated gender differences in hepatic, adipose tissue and whole-body insulin sensitivity between severely obese men and women. In this study, we aimed to determine the differences in glucose metabolism between severely obese men and women using tissue-specific measurements of insulin sensitivity. Insulin sensitivity was compared between age and body mass index (BMI-matched obese men and women by a two-step euglycemic hyperinsulinemic clamp with infusion of [6,6-2H2]glucose. Basal endogenous glucose production and insulin sensitivity of the liver, adipose tissue and peripheral tissues were assessed. Liver fat content was assessed by proton magnetic resonance spectroscopy in a subset of included subjects. We included 46 obese men and women (age, 48±2 vs 46±2 years, p=0.591; BMI, 41±1 vs 41±1 kg/m2, p=0.832. There was no difference in basal endogenous glucose production (14.4±1.0 vs 15.3±0.5 µmol•kg fat-free mass-1•min-1, p=0.410, adipose tissue insulin sensitivity (insulin-mediated suppression of free fatty acids, 71.6±3.6 vs 76.1±2.6%, p=0.314 or peripheral insulin sensitivity (insulin-stimulated rate of disappearance of glucose, 26.2±2.1 vs 22.7±1.7 µmol•kg-1•min-1, p=0.211. Obese men were characterized by lower hepatic insulin sensitivity (insulin-mediated suppression of endogenous glucose production, 61.7±4.1 vs 72.8±2.5% in men vs women, resp., p=0.028. Finally, these observations could not be explained by differences in liver fat content (men vs women, 16.5±3.1 vs 16.0±2.5%, p=0.913, n=27.We conclude that obese men have lower hepatic, but comparable adipose tissue and peripheral tissue, insulin sensitivity compared to similarly obese women. Hepatic insulin resistance may

  19. Aging of non-visual spectral sensitivity to light in humans: compensatory mechanisms?

    Directory of Open Access Journals (Sweden)

    Raymond P Najjar

    Full Text Available The deterioration of sleep in the older population is a prevalent feature that contributes to a decrease in quality of life. Inappropriate entrainment of the circadian clock by light is considered to contribute to the alteration of sleep structure and circadian rhythms in the elderly. The present study investigates the effects of aging on non-visual spectral sensitivity to light and tests the hypothesis that circadian disturbances are related to a decreased light transmittance. In a within-subject design, eight aged and five young subjects were exposed at night to 60 minute monochromatic light stimulations at 9 different wavelengths (420-620 nm. Individual sensitivity spectra were derived from measures of melatonin suppression. Lens density was assessed using a validated psychophysical technique. Although lens transmittance was decreased for short wavelength light in the older participants, melatonin suppression was not reduced. Peak of non-visual sensitivity was, however, shifted to longer wavelengths in the aged participants (494 nm compared to young (484 nm. Our results indicate that increased lens filtering does not necessarily lead to a decreased non-visual sensitivity to light. The lack of age-related decrease in non-visual sensitivity to light may involve as yet undefined adaptive mechanisms.

  20. The sensitization potential of sunscreen after ablative fractional skin resurfacing using modified human repeated insult patch test.

    Science.gov (United States)

    Boonchai, Waranya; Sathaworawong, Angkana; Wongpraparut, Chanisada; Wanitphakdeedecha, Rungsima

    2015-10-01

    Ablative fractional skin resurfacing has become popular and proven to be useful in treating scars, photoaging and wrinkles. Although post-inflammatory hyperpigmentation (PIH) is the most common complication especially in dark-skinned patients like Asian. Several modalities have been used to overcome the PIH. To determine the sensitization potential of sunscreen applied immediately after ablative fractional skin resurfacing. Sixty volunteers were recruited. Of these 30 subjects were from previous ablative fractional skin resurfacing study who applied broad-spectrum sunscreen containing anti-inflammatory agent starting on the first day after resurfacing and another 30 non-resurfacing subjects had applied the same sunscreen on the intact skin. All subjects were patch/photopatch tested for sensitization study by using modified human repeated insult patch test (HRIPT). There were significantly higher sensitization rate of UV-filter, octocrylene and the sunscreen in resurfacing group than in non-resurfacing group. Early application of sunscreen after ablative fractional skin resurfacing has increased the incidence of sensitization potential of sunscreen. The sunscreen is recommended to start using from D3 after fractional ablative skin resurfacing to ensure the complete recovery of skin barrier and minimize the risk of sensitization.

  1. Characterization of lipid metabolism in insulin-sensitive adipocytes differentiated from immortalized human mesenchymal stem cells

    DEFF Research Database (Denmark)

    Prawitt, Janne; Niemeier, Andreas; Kassem, Moustapha

    2008-01-01

    There is a great demand for cell models to study human adipocyte function. Here we describe the adipogenic differentiation of a telomerase-immortalized human mesenchymal stem cell line (hMSC-Tert) that maintains numerous features of terminally differentiated adipocytes even after prolonged...

  2. Retained sensitivity to cytotoxic pyrimidine nucleoside analogs in thymidine kinase 2 deficient human fibroblasts.

    Science.gov (United States)

    Bjerke, Mia; Solaroli, Nicola; Lesko, Nicole; Balzarini, Jan; Johansson, Magnus; Karlsson, Anna

    2010-01-01

    Thymidine kinase 2 (TK2) is a mitochondrial deoxyribonucleoside kinase that phosphorylates several nucleoside analogs used in anti-viral and anti-cancer therapy. A fibroblast cell line with decreased TK2 activity was investigated in order to obtain insights in the effects of TK2 deficiency on nucleotide metabolism. The role of TK2 for the sensitivity against cytotoxic nucleoside analogs was also investigated. The TK2 deficient cells retained their sensitivity against all pyrimidine nucleoside analogs tested. This study suggests that nucleoside analog phosphorylation mediated by TK2 may be less important, compared to other deoxyribonucleoside kinases, for the cytotoxic effects of these compounds.

  3. [EVALUATION OF THE HUMAN SENSITIVITY TO SMALLPOX VIRUS BY THE PRIMARY CULTURES OF THE MONOCYTE-MACROPHAGES].

    Science.gov (United States)

    Zamedyanskaya, A S; Titova, K A; Sergeev, Al A; Kabanov, A S; Bulychev, L E; Sergeev, Ar A; Galakhova, D O; Nesterov, A E; Nosareva, O V; Shishkina, L N; Taranov, O S; Omigov, V V; Agafonov, A P; Sergeev, A N

    2016-01-01

    Studies of the primary cultures of granulocytes, mononuclear, and monocyte-macrophage cells derived from human blood were performed using variola virus (VARV) in the doses of 0.001-0.021 PFU/cell (plaques-forming units per cell). Positive dynamics of the virus accumulation was observed only in the monocyte-macrophages with maximum values of virus concentration (5.0-5.5 Ig PFU/ml) mainly within six days after the infection. The fact of VARV replication in the monocyte-macrophages was confirmed by the data of electron microscopy. At the same time, virus vaccines when tested in doses 3.3 and 4.2 Ig PFU/ml did not show the ability to reproduce in these human cells. The people sensitivity to VARV as assessed from the data obtained on human monocyte-macrophages corresponded to -1 PFU (taking into account the smooth interaction of the virus in the body to the cells of this type), which is consistent to previously found theoretical data on the virus sensitivity. The human susceptibility to VARV assessed experimentally can be used to predict the adequacy of developed smallpox models (in vivo) based on susceptible animals. This is necessary for reliable assessment of the efficiency of development of drugs for treatment and prophylaxis of the smallpox.

  4. Effects of inhibition of interleukin-6 signalling on insulin sensitivity and lipoprotein (a levels in human subjects with rheumatoid diseases.

    Directory of Open Access Journals (Sweden)

    Olaf Schultz

    2010-12-01

    Full Text Available Interleukin-6 (IL-6 is a pro-inflammatory cytokine that has been found to be increased in type 2 diabetic subjects. However, it still remains unclear if these elevated IL-6 levels are co-incidental or if this cytokine is causally related to the development of insulin resistance and type 2 diabetes in humans. Therefore, in the present study we examined insulin sensitivity, serum adipokine levels and lipid parameters in human subjects before and after treatment with the IL-6 receptor antibody Tocilizumab.11 non-diabetic patients with rheumatoid disease were included in the study. HOMA-IR was calculated and serum levels for leptin, adiponectin, triglycerides, LDL-cholesterol, HDL-cholesterol and lipoprotein (a (Lp (a were measured before as well as one and three months after Tocilizumab treatment. The HOMA index for insulin resistance decreased significantly. While leptin concentrations were not altered by inhibition of IL-6 signalling, adiponectin concentrations significantly increased. Thus the leptin to adiponectin ratio, a novel marker for insulin resistance, exhibited a significant decrease. Serum triglycerides, LDL-cholesterol and HDL-cholesterol tended to be increased whereas Lp (a levels significantly decreased.Inhibition of IL-6 signalling improves insulin sensitivity in humans with immunological disease suggesting that elevated IL-6 levels in type 2 diabetic subjects might be causally involved in the pathogenesis of insulin resistance. Furthermore, our data indicate that inhibition of IL-6 signalling decreases Lp (a serum levels, which might reduce the cardiovascular risk of human subjects.

  5. Sensitive Tumorigenic Potential Evaluation of Adult Human Multipotent Neural Cells Immortalized by hTERT Gene Transduction.

    Directory of Open Access Journals (Sweden)

    Kee Hang Lee

    Full Text Available Stem cells and therapeutic genes are emerging as a new therapeutic approach to treat various neurodegenerative diseases with few effective treatment options. However, potential formation of tumors by stem cells has hampered their clinical application. Moreover, adequate preclinical platforms to precisely test tumorigenic potential of stem cells are controversial. In this study, we compared the sensitivity of various animal models for in vivo stem cell tumorigenicity testing to identify the most sensitive platform. Then, tumorigenic potential of adult human multipotent neural cells (ahMNCs immortalized by the human telomerase reverse transcriptase (hTERT gene was examined as a stem cell model with therapeutic genes. When human glioblastoma (GBM cells were injected into adult (4-6-week-old Balb/c-nu, adult NOD/SCID, adult NOG, or neonate (1-2-week-old NOG mice, the neonate NOG mice showed significantly faster tumorigenesis than that of the other groups regardless of intracranial or subcutaneous injection route. Two kinds of ahMNCs (682TL and 779TL were primary cultured from surgical samples of patients with temporal lobe epilepsy. Although the ahMNCs were immortalized by lentiviral hTERT gene delivery (hTERT-682TL and hTERT-779TL, they did not form any detectable masses, even in the most sensitive neonate NOG mouse platform. Moreover, the hTERT-ahMNCs had no gross chromosomal abnormalities on a karyotype analysis. Taken together, our data suggest that neonate NOG mice could be a sensitive animal platform to test tumorigenic potential of stem cell therapeutics and that ahMNCs could be a genetically stable stem cell source with little tumorigenic activity to develop regenerative treatments for neurodegenerative diseases.

  6. A novel method of selective removal of human DNA improves PCR sensitivity for detection of Salmonella Typhi in blood samples.

    Science.gov (United States)

    Zhou, Liqing; Pollard, Andrew J

    2012-07-27

    Enteric fever is a major public health problem, causing an estimated 21million new cases and 216,000 or more deaths every year. Current diagnosis of the disease is inadequate. Blood culture only identifies 45 to 70% of the cases and is time-consuming. Serological tests have very low sensitivity and specificity. Clinical samples obtained for diagnosis of enteric fever in the field generally have blood, so that even PCR-based methods, widely used for detection of other infectious diseases, are not a straightforward option in typhoid diagnosis. We developed a novel method to enrich target bacterial DNA by selective removal of human DNA from blood samples, enhancing the sensitivity of PCR tests. This method offers the possibility of improving PCR assays directly using clinical specimens for diagnosis of this globally important infectious disease. Blood samples were mixed with ox bile for selective lysis of human blood cells and the released human DNA was then digested with addition of bile resistant micrococcal nuclease. The intact Salmonella Typhi bacteria were collected from the specimen by centrifugation and the DNA extracted with QIAamp DNA mini kit. The presence of Salmonella Typhi bacteria in blood samples was detected by PCR with the fliC-d gene of Salmonella Typhi as the target. Micrococcal nuclease retained activity against human blood DNA in the presence of up to 9% ox bile. Background human DNA was dramatically removed from blood samples through the use of ox bile lysis and micrococcal nuclease for removal of mammalian DNA. Consequently target Salmonella Typhi DNA was enriched in DNA preparations and the PCR sensitivity for detection of Salmonella Typhi in spiked blood samples was enhanced by 1,000 fold. Use of a combination of selective ox-bile blood cell lysis and removal of human DNA with micrococcal nuclease significantly improves PCR sensitivity and offers a better option for improved typhoid PCR assays directly using clinical specimens in diagnosis of

  7. Traditional Chinese Medicine Curcumin Sensitizes Human Colon Cancer to Radiation by Altering the Expression of DNA Repair-related Genes.

    Science.gov (United States)

    Yang, Guangen; Qiu, Jianming; Wang, Dong; Tao, Yong; Song, Yihuan; Wang, Hongtao; Tang, Juping; Wang, Xing; Sun, Y U; Yang, Zhijian; Hoffman, Robert M

    2018-01-01

    The aim of the present study was to investigate the radio-sensitizing efficacy of curcumin, a traditional Chinese medicine (TCM) on colon cancer cells in vitro and in vivo. Human colon cancer HT-29 cells were treated with curcumin (2.5 μM), irradiation (10 Gy) and the combination of irradiation and curcumin. Cell proliferation was assessed using the MTT assay. Apoptotic cells were detected by Annexin V-PE/7-AAD analysis. PCR was performed to determine differential-expression profiling of 95 DNA-repair genes in irradiated cells and cells treated with both irradiation and curcumin. Differentially-expressed genes were confirmed by Western blotting. In vivo radio-sensitizing efficacy of curcumin was assessed in a xenograft mouse model of HT-29 colon cancer. Curcumin was administrated daily by intraperitoneal injection at 20 mg/kg/dose. Mice received irradiation (10 Gy) twice weekly. Apoptosis of the cancer cells following treatment was determined by TUNEL staining. Irradiation induced proliferation inhibition and apoptosis of HT-29 cells in vitro. Concurrent curcumin treatment sensitized the HT-29 tumor to irradiation (pcurcumin and irradiation compared with irradiation alone (pcurcumin and irradiation resulted in a significantly greater tumor-growth inhibition and apoptosis compared to irradiation treatment alone (pCurcumin sensitizes human colon cancer in vitro and in vivo to radiation. Downregulation of LIG4 and PNKP and upregulation of XRCC5 and CCNH DNA-repair-related genes were involved in the radio-sensitizing efficacy of curcumin in colon cancer. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  8. Radio-sensitization by Piper longumine of human breast adenoma MDA-MB-231 cells in vitro.

    Science.gov (United States)

    Yao, Jian-Xin; Yao, Zhi-Feng; Li, Zhan-Feng; Liu, Yong-Biao

    2014-01-01

    The current study investigated the effects of Piper longumine on radio-sensitization of human breast cancer MDA-MB-231 cells and underlying mechanisms. Human breast cancer MDA-MB-231 cells were cultured in vitro and those in logarithmic growth phase were selected for experiments divided into four groups: control, X-ray exposed, Piper longumine, and Piper longumine combined with X-rays. Conogenic assays were performed to determine the radio-sensitizing effects. Cell survival curves were fitted by single-hit multi-target model and then the survival fraction (SF), average lethal dose (D0), quasi-threshold dose (Dq) and sensitive enhancement ratio (SER) were calculated. Cell apoptosis was analyzed by flow cytometry (FCM).Western blot assays were employed for expression of apoptosis-related proteins (Bc1-2 and Bax) after treatment with Piper longumine and/or X-ray radiation. The intracellular reactive oxygen species (ROS) level was detected by FCM with a DCFH-DA probe. The cloning formation capacity was decreased in the group of piperlongumine plus radiation, which displayed the values of SF2, D0, Dq significantly lower than those of radiation alone group and the sensitive enhancement ratio (SER) of D0 was1.22 and 1.29, respectively. The cell apoptosis rate was increased by the combination treatment of Piper longumine and radiation. Piper longumine increased the radiation-induced intracellular levels of ROS. Compared with the control group and individual group, the combination group demonstrated significantly decreased expression of Bcl-2 with increased Bax. Piper longumine at a non-cytotoxic concentration can enhance the radio-sensitivity of MDA- MB-231cells, which may be related to its regulation of apoptosis-related protein expression and the increase of intracellular ROS level, thus increasing radiation-induced apoptosis.

  9. Muscle triacylglycerol and hormone-sensitive lipase activity in untrained and trained human muscles

    DEFF Research Database (Denmark)

    Helge, Jørn Wulff; Biba, Taus O; Galbo, Henrik

    2006-01-01

    During exercise, triacylglycerol (TG) is recruited in skeletal muscles. We hypothesized that both muscle hormone-sensitive lipase (HSL) activity and TG recruitment would be higher in trained than in untrained subjects in response to prolonged exercise. Healthy male subjects (26 +/- 1 years, body ...

  10. Lipid droplet size and location in human skeletal muscle fibers are associated with insulin sensitivity

    DEFF Research Database (Denmark)

    Nielsen, Joachim; Christensen, Anders E; Nellemann, Birgitte

    2017-01-01

    In skeletal muscle, an accumulation of lipid droplets (LDs) in the subsarcolemmal space is associated with insulin resistance, but the underlying mechanism is not clear. We aimed to investigate how the size, number and location of LDs are associated with insulin sensitivity and muscle fiber types...... are associated with insulin resistance in skeletal muscle....

  11. Sensitivity evaluation of human factors for reliability of the containment spray system

    International Nuclear Information System (INIS)

    Tsujimura, Yasuhiro; Suzuki, Eiji

    1988-01-01

    Evaluation of the human reliability is one of the most difficult problems that deal with the safety and reliability of large systems, especially of the Engineered Safety Features (ESF) of the nuclear power plant. Influences of human factors on the reliability of the Containment Spray System in the ESF were estimated by using the FTA method in this paper. As a result, the adequacy of the system structure and the effects of human factors on variations of the design of the system structure were explained. (author)

  12. The Tactile Modality: A Review of Tactile Sensitivity and Human Tactile Interfaces

    National Research Council Canada - National Science Library

    Myles, Kimberly; Binseel, Mary S

    2007-01-01

    ... the most important. Hearing also is viewed as necessary for interpreting environmental stimuli. In contrast, touch, smell, and taste are largely ignored as being essential to humans' interaction with the environment...

  13. The Tactile Modality: A Review of Tactile Sensitivity and Human Tactile Interfaces

    National Research Council Canada - National Science Library

    Myles, Kimberly; Binseel, Mary S

    2007-01-01

    .... Because humans have a limited capacity to receive, hold in working memory, and cognitively process information taken from the environment, the use of one sensory modality to convey information within...

  14. FENOFIBRATE ADMINISTRATION DOES NOT AFFECT MUSCLE TRIGLYCERIDE CONCENTRATION OR INSULIN SENSITIVITY IN HUMANS

    Science.gov (United States)

    Perreault, Leigh; Bergman, Bryan C.; Hunerdosse, Devon M.; Howard, David J.; Eckel, Robert H.

    2010-01-01

    Objective Animal data suggest that males, in particular, rely on PPAR-α activity to maintain normal muscle triglyceride metabolism. We sought to examine whether this was also true in men vs. women and its relationship to insulin sensitivity. Materials/Methods Normolipidemic obese men (n=9) and women (n=9) underwent an assessment of insulin sensitivity (IVGTT) and intramuscular triglyceride metabolism (GC/MS and GC/C/IRMS from plasma and muscle biopsies taken after infusion of [U-13C]palmitate) before and after 12 weeks of fenofibrate treatment. Results Women were more insulin sensitive (Si; 5.2(0.7 vs. 2.4(0.4 ×10−4/uU/ml, W vs. M, ptriglyceride (IMTG) concentration (41.9(15.5 vs. 30.8(5.1 ug/mg dry weight, W vs. M, p=0.43), and IMTG fractional synthesis rate (FSR; 0.27(0.07 vs. 0.35(0.06/hr, W vs. M, p=0.41) as men. Fenofibrate enhanced FSR in men (0.35(0.06 to 0.54(0.06, p=0.05), with no such change seen in women (0.27(0.07 to 0.32(0.13, p=0.73), and no change in IMTG concentration in either group (23.0(3.9 in M, p=0.26 vs. baseline; 36.3(12.0 in W, p=0.79 vs. baseline). Insulin sensitivity was unaffected by fenofibrate (p>0.68). Lower percent saturation of IMTG in women vs. men before (29.1(2.3 vs. 35.2(1.7%, p=0.06) and after (27.3(2.8 vs. 35.1(1.9%, p=0.04) fenofibrate most closely related to their greater insulin sensitivity (R2=0.34, p=0.10), and was largely unchanged by the drug. Conclusions PPAR-α agonist therapy had little effect on IMTG metabolism in men or women. IMTG saturation, rather than IMTG concentration or FSR, most closely (but not significantly) related to insulin sensitivity and was unchanged by fenofibrate administration. PMID:21306746

  15. Analysis of clonogenic human brain tumour cells: preliminary results of tumour sensitivity testing with BCNU

    Energy Technology Data Exchange (ETDEWEB)

    Rosenblum, M L; Dougherty, D A; Deen, D F; Hoshino, T; Wilson, C B [California Univ., San Francisco (USA). Dept. of Neurology

    1980-04-01

    Biopsies from 6 patients with glioblastoma multiforme were disaggregated and single cells were treated in vitro with various concentrations of 1,3-bis(2-chloroethyl)-1-nitroso urea (BCNU) and plated for cell survival. One patient's cells were sensitive to BCNU in vitro; after a single dose of BCNU her brain scan reverted to normal and she was clinically well. Five tumours demonstrated resistance in vitro. Three of these tumours progressed during the first course of chemotherapy with a nitrosourea and the patients died at 21/2, 4 and 81/2 months after operation. Two patients who showed dramatic responses to radiation therapy were considered unchanged after the first course of nitrosourea therapy (although one demonstrated tumour enlargement on brain scan). The correlation of in vitro testing of tumour cell sensitivity with actual patient response is encouraging enough to warrant further work to determine whether such tests should weigh in decisions on patient therapy.

  16. Human adipocytes are highly sensitive to intermittent hypoxia induced NF-kappaB activity and subsequent inflammatory gene expression

    International Nuclear Information System (INIS)

    Taylor, Cormac T.; Kent, Brian D.; Crinion, Sophie J.; McNicholas, Walter T.; Ryan, Silke

    2014-01-01

    Highlights: • Intermittent hypoxia (IH) leads to NF-κB activation in human primary adipocytes. • Adipocytes bear higher pro-inflammatory potential than other human primary cells. • IH leads to upregulation of multiple pro-inflammatory genes in human adipocytes. - Abstract: Introduction: Intermittent hypoxia (IH)-induced activation of pro-inflammatory pathways is a major contributing factor to the cardiovascular pathophysiology associated with obstructive sleep apnea (OSA). Obesity is commonly associated with OSA although it remains unknown whether adipose tissue is a major source of inflammatory mediators in response to IH. The aim of this study was to test the hypothesis that IH leads to augmented inflammatory responses in human adipocytes when compared to cells of non-adipocyte lineages. Methods and results: Human primary subcutaneous and visceral adipocytes, human primary microvascular pulmonary endothelial cells (HUMEC-L) and human primary small airway epithelial cells (SAEC) were exposed to 0, 6 or 12 cycles of IH or stimulated with tumor necrosis factor (TNF)-α. IH led to a robust increase in NF-κB DNA-binding activity in adipocytes compared with normoxic controls regardless of whether the source of adipocytes was visceral or subcutaneous. Notably, the NF-κB response of adipocytes to both IH and TNF-α was significantly greater than that in HUMEC-L and SAEC. Western blotting confirmed enhanced nuclear translocation of p65 in adipocytes in response to IH, accompanied by phosphorylation of I-κB. Parallel to p65 activation, we observed a significant increase in secretion of the adipokines interleukin (IL)-8, IL-6 and TNF-α with IH in adipocytes accompanied by significant upregulation of mRNA expression. PCR-array suggested profound influence of IH on pro-inflammatory gene expression in adipocytes. Conclusion: Human adipocytes demonstrate strong sensitivity to inflammatory gene expression in response to acute IH and hence, adipose tissue may be a key

  17. Human adipocytes are highly sensitive to intermittent hypoxia induced NF-kappaB activity and subsequent inflammatory gene expression

    Energy Technology Data Exchange (ETDEWEB)

    Taylor, Cormac T. [School of Medicine and Medical Science, The Conway Institute, University College Dublin (Ireland); Kent, Brian D.; Crinion, Sophie J.; McNicholas, Walter T. [School of Medicine and Medical Science, The Conway Institute, University College Dublin (Ireland); Pulmonary and Sleep Disorders Unit, St. Vincent’s University Hospital, Dublin (Ireland); Ryan, Silke, E-mail: silke.ryan@ucd.ie [School of Medicine and Medical Science, The Conway Institute, University College Dublin (Ireland); Pulmonary and Sleep Disorders Unit, St. Vincent’s University Hospital, Dublin (Ireland)

    2014-05-16

    Highlights: • Intermittent hypoxia (IH) leads to NF-κB activation in human primary adipocytes. • Adipocytes bear higher pro-inflammatory potential than other human primary cells. • IH leads to upregulation of multiple pro-inflammatory genes in human adipocytes. - Abstract: Introduction: Intermittent hypoxia (IH)-induced activation of pro-inflammatory pathways is a major contributing factor to the cardiovascular pathophysiology associated with obstructive sleep apnea (OSA). Obesity is commonly associated with OSA although it remains unknown whether adipose tissue is a major source of inflammatory mediators in response to IH. The aim of this study was to test the hypothesis that IH leads to augmented inflammatory responses in human adipocytes when compared to cells of non-adipocyte lineages. Methods and results: Human primary subcutaneous and visceral adipocytes, human primary microvascular pulmonary endothelial cells (HUMEC-L) and human primary small airway epithelial cells (SAEC) were exposed to 0, 6 or 12 cycles of IH or stimulated with tumor necrosis factor (TNF)-α. IH led to a robust increase in NF-κB DNA-binding activity in adipocytes compared with normoxic controls regardless of whether the source of adipocytes was visceral or subcutaneous. Notably, the NF-κB response of adipocytes to both IH and TNF-α was significantly greater than that in HUMEC-L and SAEC. Western blotting confirmed enhanced nuclear translocation of p65 in adipocytes in response to IH, accompanied by phosphorylation of I-κB. Parallel to p65 activation, we observed a significant increase in secretion of the adipokines interleukin (IL)-8, IL-6 and TNF-α with IH in adipocytes accompanied by significant upregulation of mRNA expression. PCR-array suggested profound influence of IH on pro-inflammatory gene expression in adipocytes. Conclusion: Human adipocytes demonstrate strong sensitivity to inflammatory gene expression in response to acute IH and hence, adipose tissue may be a key

  18. Exercise increases human skeletal muscle insulin sensitivity via coordinated increases in microvascular perfusion and molecular signaling

    DEFF Research Database (Denmark)

    Sjøberg, Kim Anker; Frøsig, Christian; Kjøbsted, Rasmus

    2017-01-01

    and increased similarly in both legs during the clamp and L-NMMA had no effect on these insulin-stimulated signaling pathways. Therefore, acute exercise increases insulin sensitivity of muscle by a coordinated increase in insulin-stimulated microvascular perfusion and molecular signaling at the level of TBC1D4...... and glycogen synthase in muscle. This secures improved glucose delivery on the one hand and increased ability to take up and dispose of the delivered glucose on the other hand....

  19. Emotional expectations influence neural sensitivity to fearful faces in humans:An event-related potential study

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    The present study tested whether neural sensitivity to salient emotional facial expressions was influenced by emotional expectations induced by a cue that validly predicted the expression of a subsequently presented target face. Event-related potentials (ERPs) elicited by fearful and neutral faces were recorded while participants performed a gender discrimination task under cued (‘expected’) and uncued (‘unexpected’) conditions. The behavioral results revealed that accuracy was lower for fearful compared with neutral faces in the unexpected condition, while accuracy was similar for fearful and neutral faces in the expected condition. ERP data revealed increased amplitudes in the P2 component and 200–250 ms interval for unexpected fearful versus neutral faces. By contrast, ERP responses were similar for fearful and neutral faces in the expected condition. These findings indicate that human neural sensitivity to fearful faces is modulated by emotional expectations. Although the neural system is sensitive to unpredictable emotionally salient stimuli, sensitivity to salient stimuli is reduced when these stimuli are predictable.

  20. Mechano-sensitive nociceptors are required to detect heat pain thresholds and cowhage itch in human skin.

    Science.gov (United States)

    Weinkauf, B; Dusch, M; van der Ham, J; Benrath, J; Ringkamp, M; Schmelz, M; Rukwied, R

    2016-02-01

    Mechano-sensitive and mechano-insensitive C-nociceptors in human skin differ in receptive field sizes and electrical excitation thresholds, but their distinct functional roles are yet unclear. After blocking the lateral femoral cutaneous nerve (NCFL) in eight healthy male subjects (3-mL Naropin(®) 1%), we mapped the skin innervation territory being anaesthetic to mechanical pin prick but sensitive to painful transcutaneous electrical stimuli. Such 'differentially anaesthetic zones' indicated that the functional innervation with mechano-sensitive nociceptors was absent but the innervation with mechano-insensitive nociceptors remained intact. In these areas, we explored heat pain thresholds, low pH-induced pain, cowhage- and histamine-induced itch, and axon reflex flare. In differentially anaesthetic skin, heat pain thresholds were above the cut-off of 50°C (non-anaesthetized skin 47 ± 0.4°C). Pain ratings to 30 μL pH 4 injections were reduced compared to non-anaesthetized skin (48 ± 9 vs. 79 ± 6 VAS; p pain. The mechano-sensitive nociceptors are crucial for cowhage-induced itch and for the assessment of heat pain thresholds. © 2015 European Pain Federation - EFIC®

  1. Histone Deacetylase Inhibitor Induced Radiation Sensitization Effects on Human Cancer Cells after Photon and Hadron Radiation Exposure

    Directory of Open Access Journals (Sweden)

    Ariungerel Gerelchuluun

    2018-02-01

    Full Text Available Suberoylanilide hydroxamic acid (SAHA is a histone deacetylase inhibitor, which has been widely utilized throughout the cancer research field. SAHA-induced radiosensitization in normal human fibroblasts AG1522 and lung carcinoma cells A549 were evaluated with a combination of γ-rays, proton, and carbon ion exposure. Growth delay was observed in both cell lines during SAHA treatment; 2 μM SAHA treatment decreased clonogenicity and induced cell cycle block in G1 phase but 0.2 μM SAHA treatment did not show either of them. Low LET (Linear Energy Transfer irradiated A549 cells showed radiosensitization effects on cell killing in cycling and G1 phase with 0.2 or 2 μM SAHA pretreatment. In contrast, minimal sensitization was observed in normal human cells after low and high LET radiation exposure. The potentially lethal damage repair was not affected by SAHA treatment. SAHA treatment reduced the rate of γ-H2AX foci disappearance and suppressed RAD51 and RPA (Replication Protein A focus formation. Suppression of DNA double strand break repair by SAHA did not result in the differences of SAHA-induced radiosensitization between human cancer cells and normal cells. In conclusion, our results suggest SAHA treatment will sensitize cancer cells to low and high LET radiation with minimum effects to normal cells.

  2. Radio-sensitivity of the cells from amyotrophic lateral sclerosis model mice transfected with human mutant SOD1

    International Nuclear Information System (INIS)

    Wate, Reika; Ito, Hidefumi; Kusaka, Hirofumi; Takahashi, Sentaro; Kubota, Yoshihisa; Suetomi, Katsutoshi; Sato, Hiroshi; Okayasu, Ryuichi

    2005-01-01

    In order to clarify the possible involvement of oxidative damage induced by ionizing radiation in the onset and/or progression of familial amyotrophic lateral sclerosis (ALS), we studied radio-sensitivity in primary cells derived from ALS model mice expressing human mutant Cu/Zn superoxide dismutase (SOD1). The primary mouse cells expressed both mouse and the mutant human SOD1. The cell survival of the transgenic mice (with mutant SOD1), determined by counting cell numbers at a scheduled time after X-irradiation, is very similar to that of cells from wild type animals. The induction and repair of DNA damage in the transgenic cells, measured by single cell gel electrophoresis and pulsed field gel electrophoresis, are also similar to those of wild type cells. These results indicate that the human mutant SOD1 gene does not seem to contribute to the alteration of radio-sensitivity, at least in the fibroblastic cells used here. Although it is necessary to consider the difference in cell types between fibroblastic and neuronal cells, the present results may suggest that ionizing radiation is not primarily responsible for the onset of familial ALS with the SOD1 mutation, and that the excess risks are probably not a concern for radiation diagnosis and therapy in familial ALS patients. (author)

  3. Hyper-radiation sensitivity of murine scid mutation and mapping of the human homologue HYRC1 gene

    International Nuclear Information System (INIS)

    Komatsu, Kenshi; Ohta, Tohru; Niikawa, Norio; Okumura, Yutaka; Kubota, Nobuo.

    1994-01-01

    The murine severe combined immunodeficient mutation (scid) is characterized by a lack of both B and T cells, due to a defect in lymphoid variable-(diversity)-joining(V(D)J) rearrangement. Scid cells are highly sensitive to both radiation-induced killing and chromosomal aberrations. Present experiments also demonstrated the high sensitivity of scid cells to killing, because of a deficient repair of double strand breaks(DSB). Scid cells can repair only 60% of radiation-induced DSB for 3 hours, while normal cells repair 85% of the DSB. Significantly reduced Do and n values were obtained from survival curves of scid cells and were similar to ataxia-telangiectasia(AT) cells (a unique human disease conferring whole body radiosensitivity). However, the kinetics of DNA synthesis after irradiation were different between the two cell types. In contrast with the radioresistant DNA synthesis of AT cells, DNA synthesis of scid cells was markedly inhibited after irradiation. The existence of different mutations was also supported by evidence of complementation in somatic cell hybrids between scid cells and AT cells. Using these hybrid cells, fragments of human chromosome 8 were introduced into scid cells HPRT mutant via X-irradiation and somatic cell fusion. The resulting hybrid clones contained human DNA fragment(s) which complemented the hyper-radiosensitivity of the scid cells. Alu-PCR products from these hybrids were used for chromosome painting using the technique of chromosome in situ suppression hybridization, allowing assignment of the human HYRC1 (hyper-radiosensitivity of murine scid mutation, complementing 1) gene, a candidate for a V(D)J recombinant gene, to human chromosome 8q11. (author)

  4. Olfactory sensitivity for sperm-attractant aromatic aldehydes: a comparative study in human subjects and spider monkeys.

    Science.gov (United States)

    Kjeldmand, Luna; Salazar, Laura Teresa Hernandez; Laska, Matthias

    2011-01-01

    Using a three-alternative forced-choice ascending staircase procedure, we determined olfactory detection thresholds in 20 human subjects for seven aromatic aldehydes and compared them to those of four spider monkeys tested in parallel using an operant conditioning paradigm. With all seven odorants, both species detected concentrations lyral, and 3-phenylpropanal. No significant correlation between presence/absence of an oxygen-containing moiety attached to the benzene ring or presence/absence of an additional alkyl group next to the functional aldehyde group, and olfactory sensitivity was found in any of the species. However, the presence of a tertiary butyl group in para position (relative to the functional aldehyde group) combined with a lack of an additional alkyl group next to the functional aldehyde group may be responsible for the finding that both species were most sensitive to bourgeonal.

  5. Sensitive method for the quantitative determination of bromocriptine in human plasma by liquid chromatography-tandem mass spectrometry.

    Science.gov (United States)

    Salvador, Arnaud; Dubreuil, Didier; Denouel, Jannick; Millerioux, L

    2005-06-25

    A sensitive LC-MS-MS assay for the quantitative determination of bromocriptine has been developed and validated and is described in this work. The assay involved the extraction of the analyte from 1 ml of human plasma using a solid phase extraction on Oasis MCX cartridges. Chromatography was performed on a Symmetry C18 (2.1 mm x 100 mm, 3.5 microm) column using a mobile phase consisting of 25:75:01 acetonitrile-water-formic acid with a flow rate of 250 microl/min. The linearity was within the concentration range of 2-500 pg/ml. The lower limit of quantification was 2 pg/ml. This method has been demonstrated to be an improvement over existing methods due to its greater sensitivity and specificity.

  6. Suppression of NRF2–ARE activity sensitizes chemotherapeutic agent-induced cytotoxicity in human acute monocytic leukemia cells

    International Nuclear Information System (INIS)

    Peng, Hui; Wang, Huihui; Xue, Peng; Hou, Yongyong; Dong, Jian; Zhou, Tong; Qu, Weidong; Peng, Shuangqing; Li, Jin; Carmichael, Paul L.; Nelson, Bud; Clewell, Rebecca; Zhang, Qiang; Andersen, Melvin E.; Pi, Jingbo

    2016-01-01

    Nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of the antioxidant response element (ARE)-dependent transcription, plays a pivotal role in chemical detoxification in normal and tumor cells. Consistent with previous findings that NRF2–ARE contributes to chemotherapeutic resistance of cancer cells, we found that stable knockdown of NRF2 by lentiviral shRNA in human acute monocytic leukemia (AML) THP-1 cells enhanced the cytotoxicity of several chemotherapeutic agents, including arsenic trioxide (As 2 O 3 ), etoposide and doxorubicin. Using an ARE-luciferase reporter expressed in several human and mouse cells, we identified a set of compounds, including isonicotinic acid amides, isoniazid and ethionamide, that inhibited NRF2–ARE activity. Treatment of THP-1 cells with ethionamide, for instance, significantly reduced mRNA expression of multiple ARE-driven genes under either basal or As 2 O 3 -challenged conditions. As determined by cell viability and cell cycle, suppression of NRF2–ARE by ethionamide also significantly enhanced susceptibility of THP-1 and U937 cells to As 2 O 3 -induced cytotoxicity. In THP-1 cells, the sensitizing effect of ethionamide on As 2 O 3 -induced cytotoxicity was highly dependent on NRF2. To our knowledge, the present study is the first to demonstrate that ethionamide suppresses NRF2–ARE signaling and disrupts the transcriptional network of the antioxidant response in AML cells, leading to sensitization to chemotherapeutic agents. - Highlights: • Identification of novel inhibitors of ARE-dependent transcription • Suppression of NRF2–ARE sensitizes THP-1 cells to chemotherapy. • Ethionamide suppresses ARE-dependent transcriptional activity. • Ethionamide and isoniazid increase the cytotoxicity of As 2 O 3 in AML cells. • Sensitization of THP-1 cells to As 2 O 3 toxicity by ethionamide is NRF2-dependent.

  7. Sensitization of interferon-γ induced apoptosis in human osteosarcoma cells by extracellular S100A4

    International Nuclear Information System (INIS)

    Pedersen, Kjetil Boye; Andersen, Kristin; Fodstad, Øystein; Mælandsmo, Gunhild Mari

    2004-01-01

    S100A4 is a small Ca 2+ -binding protein of the S100 family with metastasis-promoting properties. Recently, secreted S100A4 protein has been shown to possess a number of functions, including induction of angiogenesis, stimulation of cell motility and neurite extension. Cell cultures from two human osteosarcoma cell lines, OHS and its anti-S100A4 ribozyme transfected counterpart II-11b, was treated with IFN-γ and recombinant S100A4 in order to study the sensitizing effects of extracellular S100A4 on IFN-γ mediated apoptosis. Induction of apoptosis was demonstrated by DNA fragmentation, cleavage of poly (ADP-ribose) polymerase and Lamin B. In the present work, we found that the S100A4-expressing human osteosarcoma cell line OHS was more sensitive to IFN-γ-mediated apoptosis than the II-11b cells. S100A4 protein was detected in conditioned medium from OHS cells, but not from II-11b cells, and addition of recombinant S100A4 to the cell medium sensitized II-11b cells to apoptosis induced by IFN-γ. The S100A4/IFN-γ-mediated induction of apoptosis was shown to be independent of caspase activation, but dependent on the formation of reactive oxygen species. Furthermore, addition of extracellular S100A4 was demonstrated to activate nuclear factor-κB (NF-κB). In conclusion, we have shown that S100A4 sensitizes osteosarcoma cells to IFN-γ-mediated induction of apoptosis. Additionally, extracellular S100A4 activates NF-κB, but whether these events are causally related remains unknown

  8. Distribution of ultraviolet-induced DNA repair synthesis in nuclease sensitive and resistant regions of human chromatin

    International Nuclear Information System (INIS)

    Smerdon, M.J.; Tlsty, T.D.; Lieberman, M.W.

    1978-01-01

    The distribution of ultraviolet radiation (uv) induced DNA repair synthesis within chromatin was examined in cultured human diploid fibroblasts (IMR-90). Measurement of the time course of repair synthesis yielded two distinct phases: An initial rapid phase (fast repair) which occurs during the first 2 to 3 h after damage and a slower phase (slow repair) associated with a tenfold decrease in the rate of nucleotide incorporation, which persists for at least 35 h after damage. Staphylococcal nuclease digests of nuclei from cells damaged with uv and labeled during the fast-repair phase revealed a marked preference of fast-repair synthesis for the nuclease-sensitive regions. A new method was developed to analyze the digestion data and showed that approximately 50% of the nucleotides incorporated during the fast-repair phase are located in staphylococcal nuclease-sensitive regions, which comprise about 30% of the genome. Calculations from these data indicate that in the staphylococcal nuclease-sensitive regions the number of newly inserted nucleotides per unit DNA is about twice that of resistant regions. These results were supported by electrophoresis studies which demonstrated a decreased representation of fast-repair synthesis in core particle DNA. In contrast, the distribution within chromatin of nucleotides incorporated during the slow-repair phase was found to be much more homogeneous with about 30% of the repair sites located in 25% of the genome. Digestion studieswith DNase I indicated a slight preference of repair synthesis for regions sensitive to this enzyme; however, no marked difference between the distributions of fast- and slow-repair synthesis was observed. This study provides evidence that the structural constraints placed upon DNA in chromatin also place constraints upon uv-induced DNA repair synthesis in human cells

  9. Integrating human and environmental health in antibiotic risk assessment: A critical analysis of protection goals, species sensitivity and antimicrobial resistance.

    Science.gov (United States)

    Le Page, Gareth; Gunnarsson, Lina; Snape, Jason; Tyler, Charles R

    2017-12-01

    Antibiotics are vital in the treatment of bacterial infectious diseases but when released into the environment they may impact non-target organisms that perform vital ecosystem services and enhance antimicrobial resistance development with significant consequences for human health. We evaluate whether the current environmental risk assessment regulatory guidance is protective of antibiotic impacts on the environment, protective of antimicrobial resistance, and propose science-based protection goals for antibiotic manufacturing discharges. A review and meta-analysis was conducted of aquatic ecotoxicity data for antibiotics and for minimum selective concentration data derived from clinically relevant bacteria. Relative species sensitivity was investigated applying general linear models, and predicted no effect concentrations were generated for toxicity to aquatic organisms and compared with predicted no effect concentrations for resistance development. Prokaryotes were most sensitive to antibiotics but the range of sensitivities spanned up to several orders of magnitude. We show reliance on one species of (cyano)bacteria and the 'activated sludge respiration inhibition test' is not sufficient to set protection levels for the environment. Individually, neither traditional aquatic predicted no effect concentrations nor predicted no effect concentrations suggested to safeguard for antimicrobial resistance, protect against environmental or human health effects (via antimicrobial resistance development). Including data from clinically relevant bacteria and also more species of environmentally relevant bacteria in the regulatory framework would help in defining safe discharge concentrations for antibiotics for patient use and manufacturing that would protect environmental and human health. It would also support ending unnecessary testing on metazoan species. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  10. Polarization sensitive changes in the human macula associated with normal aging and age-related macular degeneration

    Science.gov (United States)

    VanNasdale, Dean Allan, Jr.

    2011-12-01

    The human macula occupies a relatively small, but crucial retinal area, as it is the location responsible for our most acute spatial vision and best color discrimination. Localizing important landmarks in the retina is difficult even in normal eyes where morphological inter-individual variability is high. This becomes even more challenging in the presence of sight-threatening pathology. With respect to the human macula, there remains a significant gap in the understanding of normal structure and function. Even less is known about the pathological mechanisms that occur in sight-threatening diseases including age-related macular degeneration. Because relatively little is known about normal aging changes, it is also difficult to differentiate those changes from changes associated with retinal disease. To better understand normal and pathological changes in the macula, imaging techniques using specific optical signatures are required. Structural features in the macula can be distinguished based on their intrinsic properties using specific light/tissue interactions. Because of the high degree of structural regularity in the macula, polarization sensitive imaging is potentially a useful tool for evaluating the morphology and integrity of the cellular architecture for both normal individuals and those affected by disease. In our investigations, we used polarization sensitive imaging to determining normal landmarks that are important clinically and for research investigations. We found that precision and accuracy in localizing the central macula was greatly improved through the use of polarization sensitive imaging. We also found that specific polarization alterations can be used to demonstrate systematic changes as a function of age, disproportionately affecting the central macular region. When evaluating patients with age-related macular degeneration, we found that precision and accuracy of localizing the central macula was also improved, even when significant pathology

  11. Effect of recombinant adenovirus encoding human p53 tumor suppressor gene (rAd-p53) on the growth and radiotherapeutic sensitivity of human lymphoma cell lines

    International Nuclear Information System (INIS)

    Yu Zeyang; Fan Wo; Li Dongqing; Zhu Ran; Wang Yongqing; Wu Jinchang

    2008-01-01

    Objective: To explore the inhibitory effect and radiation sensitization of recombinant adenovirus encoding human p53 tumor suppressor gene (rAd-p53) on human lymphoma cell lines. Methods: Human lymphoma cell lines Raji and Daudi were treated with rAd-p53, radiation therapy and combined treatment, respectively. The cell growth inhibition was assessed by MTT. The p53 protein expression was detected by Western blotting, and p53 mRNA was detected by BT-PCB. Results: The MTT results showed that the inhibitory effect and radiosensitivity enhancement of rAd-p53 on human lymphoma cell lines were not obvious [Raji: (27.5±4.1)%; Daudi: (28.1±1.6)%]. The results of Western blotting and BT-PCB showed that extrinsic p53 protein and p53 mRNA were expressed to some degree, but not at high-level. In addition, the results didn't demonstrate obvious radiosensitivity enhancement. Conclusions: The role of inhibition and radiosensitivity enhancement of rAd-p53 was not significant on human lymphoma cell lines. (authors)

  12. Accuracy of Diagnostic Methods and Surveillance Sensitivity for Human Enterovirus, South Korea, 1999–2011

    Science.gov (United States)

    Hyeon, Ji-Yeon; Hwang, Seoyeon; Kim, Hyejin; Song, Jaehyoung; Ahn, Jeongbae; Kang, Byunghak; Kim, Kisoon; Choi, Wooyoung; Chung, Jae Keun; Kim, Cheon-Hyun; Cho, Kyungsoon; Jee, Youngmee; Kim, Jonghyun; Kim, Kisang; Kim, Sun-Hee; Kim, Min-Ji

    2013-01-01

    The epidemiology of enteroviral infection in South Korea during 1999–2011 chronicles nationwide outbreaks and changing detection and subtyping methods used over the 13-year period. Of 14,657 patients whose samples were tested, 4,762 (32.5%) samples were positive for human enterovirus (human EV); as diagnostic methods improved, the rate of positive results increased. A seasonal trend of outbreaks was documented. Genotypes enterovirus 71, echovirus 30, coxsackievirus B5, enterovirus 6, and coxsackievirus B2 were the most common genotypes identified. Accurate test results correlated clinical syndromes to enterovirus genotypes: aseptic meningitis to echovirus 30, enterovirus 6, and coxsackievirus B5; hand, foot and mouth disease to coxsackievirus A16; and hand, foot and mouth disease with neurologic complications to enterovirus 71. There are currently no treatments specific to human EV infections; surveillance of enterovirus infections such as this study provides may assist with evaluating the need to research and develop treatments for infections caused by virulent human EV genotypes. PMID:23876671

  13. Brown adipose tissue improves whole-body glucose homeostasis and insulin sensitivity in humans

    Science.gov (United States)

    Brown adipose tissue (BAT) has attracted scientific interest as an antidiabetic tissue owing to its ability to dissipate energy as heat. Despite a plethora of data concerning the role of BAT in glucose metabolism in rodents, the role of BAT (if any) in glucose metabolism in humans remains unclear. T...

  14. Human gamma interferon production by cytotoxic T lymphocytes sensitized during hepatitis A virus infection

    International Nuclear Information System (INIS)

    Maier, K.; Gabriel, P.; Koscielniak, E.; Stierhof, Y.D.; Wiedmann, K.H.; Flehmig, B.; Vallbracht, A.

    1988-01-01

    The production of interferon (IFN) during a chromium-51 release assay with hepatitis A virus (HAV)-infected fibroblasts and autologous peripheral blood lymphocytes from patients with acute HAV infection was studied to determine whether IFN plays a role in immunopathogenesis of hepatitis A infection in humans. Skin fibroblasts of eight patients after acute HAV infection and from two control persons without history of current of past HAV infection were infected with HAV. Peripheral blood lymphocytes were collected at different times after the onset of icterus and tested in a chromium-51 release assay against autologous HAV-infected skin fibroblasts for their cytolytic and IFN-producing activity. The IFN produced during the assay was characterized and found to have the properties of human gamma IFN. Cytotoxicity and gamma IFN release were virus specific. The cell types responsible for both functions were characterized and found to be in the HLA-dependent T8 + lymphocyte subset. Considering that gamma IFN has an antiviral effect on persistent HAV infection in vitro and that it probably accounts for stimulation of HLA class I antigen expression on hepatocytes, these experimental results presented here demonstrate that human gamma IFN produced by HAV-specific T cells may participate in pathogenesis of hepatitis A infection in humans

  15. mTOR inhibition sensitizes human hepatocellular carcinoma cells to resminostat

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Xingang, E-mail: pengxinggang26@sina.com [Department of Emergency General Surgery, The Affiliated Hospital of Qingdao University, Qingdao (China); Zhang, Donghui, E-mail: zhangdonghuiyx@sina.com [Department of Infectious Disease, Linyi People’s Hospital, Linyi (China); Li, Zhengling, E-mail: lizhenglingzz@sina.com [Department of Nursing, Tengzhou Central People’s Hospital, Tengzhou (China); Fu, Meili, E-mail: fumeilidrlinyi@tom.com [Department of Infectious Disease, Linyi People’s Hospital, Linyi (China); Liu, Haiyan, E-mail: liuhaiyanlinyi5@sina.com [Department of Nursing, Linyi People’s Hospital, Linyi (China)

    2016-09-02

    Histone deacetylases (HDACs) hyper-activity in hepatocellular carcinoma (HCC) is often associated with patients’ poor prognosis. Our previous study has shown that resminostat, a novel HDAC inhibitor (HDACi), activated mitochondrial permeability transition pore (mPTP)-dependent apoptosis pathway in HCC cells. Here we explored the potential resminostat resistance factor by focusing on mammalian target of rapamycin (mTOR). We showed that AZD-2014, a novel mTOR kinase inhibitor, potentiated resminostat-induced cytotoxicity and proliferation inhibition in HCC cells. Molecularly, AZD-2014 enhanced resminostat-induced mPTP apoptosis pathway activation in HCC cells. Inhibition of this apoptosis pathway, by the caspase-9 specific inhibitor Ac-LEHD-CHO, the mPTP blockers (sanglifehrin A/cyclosporine A), or by shRNA-mediated knockdown of mPTP component cyclophilin-D (Cyp-D), significantly attenuated resminostat plus AZD-2014-induced cytotoxicity and apoptosis in HCC cells. Significantly, mTOR shRNA knockdown or kinase-dead mutation (Asp-2338-Ala) also sensitized HCC cells to resminostat, causing profound cytotoxicity and apoptosis induction. Together, these results suggest that mTOR could be a primary resistance factor of resminostat. Targeted inhibition of mTOR may thus significantly sensitize HCC cells to resminostat. - Highlights: • AZD-2014 potentiates resminostat’s cytotoxicity against HCC cells. • AZD-2014 facilitates resminostat-induced HCC cell apoptosis. • AZD-2014 augments resminostat-induced mitochondrial apoptosis pathway activation. • mTOR shRNA or kinase-dead mutation significantly sensitizes HCC cells to resminostat.

  16. Four days of simulated shift work reduces insulin sensitivity in humans.

    Science.gov (United States)

    Bescos, R; Boden, M J; Jackson, M L; Trewin, A J; Marin, E C; Levinger, I; Garnham, A; Hiam, D S; Falcao-Tebas, F; Conte, F; Owens, J A; Kennaway, D J; McConell, G K

    2018-06-01

    The aim of this study was to investigate the effects of 4 consecutive simulated night shifts on glucose homeostasis, mitochondrial function and central and peripheral rhythmicities compared with a simulated day shift schedule. Seventeen healthy adults (8M:9F) matched for sleep, physical activity and dietary/fat intake participated in this study (night shift work n = 9; day shift work n = 8). Glucose tolerance and insulin sensitivity before and after 4 nights of shift work were measured by an intravenous glucose tolerance test and a hyperinsulinaemic euglycaemic clamp respectively. Muscles biopsies were obtained to determine insulin signalling and mitochondrial function. Central and peripheral rhythmicities were assessed by measuring salivary melatonin and expression of circadian genes from hair samples respectively. Fasting plasma glucose increased (4.4 ± 0.1 vs. 4.6 ± 0.1 mmol L -1 ; P = .001) and insulin sensitivity decreased (25 ± 7%, P night shift, with no changes following the day shift. Night shift work had no effect on skeletal muscle protein expression (PGC1α, UCP3, TFAM and mitochondria Complex II-V) or insulin-stimulated pAkt Ser473, pTBC1D4Ser318 and pTBC1D4Thr642. Importantly, the metabolic changes after simulated night shifts occurred despite no changes in the timing of melatonin rhythmicity or hair follicle cell clock gene expression across the wake period (Per3, Per1, Nr1d1 and Nr1d2). Only 4 days of simulated night shift work in healthy adults is sufficient to reduce insulin sensitivity which would be expected to increase the risk of T2D. © 2018 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

  17. Human-related factors regulate the spatial ecology of domestic cats in sensitive areas for conservation.

    Directory of Open Access Journals (Sweden)

    Joaquim P Ferreira

    Full Text Available BACKGROUND: Domestic cats ranging freely in natural areas are a conservation concern due to competition, predation, disease transmission or hybridization with wildcats. In order to improve our ability to design effective control policies, we investigate the factors affecting their numbers and space use in natural areas of continental Europe. METHODOLOGY/PRINCIPAL FINDINGS: We describe the patterns of cat presence, abundance and space use and analyse the associated environmental and human constraints in a well-preserved Mediterranean natural area with small scattered local farms. We failed in detecting cats in areas away from human settlements (trapping effort above 4000 trap-nights, while we captured 30 individuals near inhabited farms. We identified 130 cats, all of them in farms still in use by people (30% of 128 farms. All cats were free-ranging and very wary of people. The main factor explaining the presence of cats was the presence of people, while the number of cats per farm was mostly affected by the occasional food provisioning with human refuse and the presence of people. The home ranges of eight radio tagged cats were centred at inhabited farms. Males went furthest away from the farms during the mating season (3.8 km on average, maximum 6.3 km, using inhabited farms as stepping-stones in their mating displacements (2.2 km of maximum inter-farm distance moved. In their daily movements, cats notably avoided entering in areas with high fox density. CONCLUSIONS: The presence, abundance and space use of cats were heavily dependent on human settlements. Any strategy aiming at reducing their impact in areas of conservation concern should aim at the presence of settlements and their spatial spread and avoid any access to human refuse. The movements of domestic cats would be limited in areas with large patches of natural vegetation providing good conditions for other carnivore mammals such as red foxes.

  18. Imaging of human breast tissue using polarization sensitive optical coherence tomography

    Science.gov (United States)

    Verma, Y.; Gautam, M.; Divakar Rao, K.; Swami, M. K.; Gupta, P. K.

    2011-12-01

    We report a study on the use of polarization sensitive optical coherence tomography (PSOCT) for discriminating malignant (invasive ductal carcinoma), benign (fibroadenoma) and normal (adipocytes) breast tissue sites. The results show that while conventional OCT, that utilizes only the intensity of light back-scattered from tissue microstructures, is able to discriminate breast tissues as normal (adipocytes) and abnormal (malignant and benign) tissues, PS-OCT helps in discriminating between malignant and benign tissue sites also. The estimated values of birefringence obtained from the PSOCT imaging show that benign breast tissue samples have significantly higher birefringence as compared to the malignant tissue samples.

  19. HLA-DR-specific suppressor cells after repeated allogeneic sensitizations of human lymphocytes in vitro

    International Nuclear Information System (INIS)

    Sasportes, M.; Fradelizi, D.; Dausset, J.

    1978-01-01

    In conclusion, DR-specific suppressor cells can be induced by repeated in vitro sensitizations. They were able to decrease a secondary proliferation, to suppress consistently, in a primary proliferative assay, when added as third cells (primed twice against a DR antigen [PLT II] and γ-irradiated), the response of unprimed cells towards stimulating cells, which share a DR specificity with the priming cell of the PLT II. The suppression follows the D part of the recombinant haplotype within an HLA-B/D recombinant family and is specific for the DR antigen used twice as stimulator for production of the PLT II

  20. Characterization of early events involved in human dendritic cell maturation induced by sensitizers: Cross talk between MAPK signalling pathways

    International Nuclear Information System (INIS)

    Trompezinski, Sandra; Migdal, Camille; Tailhardat, Magalie; Le Varlet, Beatrice; Courtellemont, Pascal; Haftek, Marek; Serres, Mireille

    2008-01-01

    Dendritic cells (DCs), efficient-antigen presenting cells play an important role in initiating and regulating immune responses. DC maturation following exposure to nickel or DNCB induced an up-regulation of phenotypic markers and inflammatory cytokine secretion. Early intracellular mechanisms involved in DC maturation required to be precise. To address this purpose, DCs derived from human monocytes were treated with sensitizers (nickel, DNCB or thimerosal) in comparison with an irritant (SDS). Our data confirming the up-regulation of CD86, CD54 and cytokine secretion (IL-8 and TNFα) induced by sensitizers but not by SDS, signalling transduction involved in DC maturation was investigated using these chemicals. Kinase activity measurement was assessed using two new sensitive procedures (Face TM and CBA) requiring few cells. SDS did not induce changes in signalling pathways whereas NiSO 4 , DNCB and thimerosal markedly activated p38 MAPK and JNK, in contrast Erk1/2 phosphorylation was completely inhibited by DNCB or thimerosal and only activated by nickel. A pre-treatment with p38 MAPK inhibitor (SB203580) suppressed Erk1/2 inhibition induced by DNCB or thimerosal demonstrating a direct interaction between p38 MAPK and Erk1/2. A pre-treatment with an antioxidant, N-acetyl-L-cysteine (NAC) markedly reduced Erk1/2 inhibition and p38 MAPK phosphorylation induced by DNCB and thimerosal, suggesting a direct activation of p38 MAPK via an oxidative stress and a regulation of MAPK signalling pathways depending on chemicals. Because of a high sensitivity of kinase activity measurements, these procedures will be suitable for weak or moderate sensitizer screening

  1. Impact of varying physical activity levels on airway sensitivity and bronchodilation in healthy humans.

    Science.gov (United States)

    Smith, Joshua R; Kurti, Stephanie P; Johnson, Ariel M; Kolmer, Sarah A; Harms, Craig

    2015-12-01

    The purpose of this study was to determine if the amount of physical activity influences airway sensitivity and bronchodilation in healthy subjects across a range of physical activity levels. Thirty healthy subjects (age, 21.9 ± 2.6 years; 13 men/17 women) with normal pulmonary function reported to the laboratory on 2 separate occasions where they were randomized to breathe either hypertonic saline (HS) (nebulized hypertonic saline (25%) for 20 min) or HS followed by 5 deep inspirations (DIs), which has been reported to bronchodilate the airways. Pulmonary function tests (PFTs) were performed prior to both conditions and following the HS breathing or 5 DIs. Moderate to vigorous physical activity (MVPA) level was measured via accelerometer worn for 7 days. Following the HS breathing, forced expiratory volume in 1 s (FEV1) and forced vital capacity (FVC) significantly decreased from baseline by -11.8% ± 8.4% and -9.3% ± 6.7%, respectively. A 2-segment linear model determined significant relationships between MVPA and percent change in FEV1 (r = 0.50) and FVC (r = 0.55). MVPA above ∼497 and ∼500 min/week for FEV1 and FVC, respectively, resulted in minor additional improvements (p > 0.05) in PFTs following the HS breathing. Following the DIs, FEV1 and FVC decreased (p 0.05) to MVPA. In conclusion, these data demonstrate that higher MVPA levels attenuated airway sensitivity but not bronchodilation in healthy subjects.

  2. A Modified LC/MS/MS Method with Enhanced Sensitivity for the Determination of Scopolamine in Human Plasma

    Science.gov (United States)

    Wang, Zuwei; Vaksman, Zalman; Putcha, Lakshmi

    2008-01-01

    Intranasal scopolamine is a choice drug for the treatment of motion sickness during space flight because of its quick onset of action, short half-life and favorable sideeffects profile. The dose administered usually ranges between 0.1 and 0.4 mg. Such small doses make it difficult to detect concentrations of scopolamine in biological fluids using existing sensitive LC/MS/MS method, especially when the biological sample volumes are limited. To measure scopolamine in human plasma to facilitate pharmacokinetic evaluation of the drug, we developed a sensitive LC/MS/MS method using 96 well micro elution plates for solid phase extraction (SPE) of scopolamine in human plasma. Human plasma (100-250 micro L) were loaded onto Waters Oasis HLB 96 well micro elution plate and eluted with 50 L of organic solvent without evaporation and reconstitution. HPLC separation of the eluted sample was performed using an Agilent Zorbax SB-CN column (50 x 2.1 mm) at a flow rate of 0.2 mL/min for 3 minutes. The mobile phase for separation was 80:20 (v/v) methanol: ammonium acetate (30 mM) in water. Concentrations of scopolamine were determined using a Micromass Quattro Micro(TM) mass spectrometer with electrospray ionization (ESI). ESI mass spectra were acquired in positive ion mode with multiple reaction monitoring for the determination of scopolamine m/z = 304.2 right arrow 138.1 and internal standard hyoscyamine m/z = 290.2 right arrow 124.1. The method is rapid, reproducible, specific and has the following parameters: scopolamine and the IS are eluted at about 1.1 and 1.7 min respectively. The linear range is 25-10000 pg/mL for scopolamine in human plasma with correlation coefficients greater than 0.99 and CV less than 0.5%. The intra-day and inter-day CVs are less than 15% for quality control samples with concentrations of 75,300, and 750 pg/mL of scopolamine in human plasma. SPE using 96 well micro elution plates allows rapid sample preparation and enhanced sensitivity for the LC

  3. Sensitivity of the human circadian pacemaker to nocturnal light: melatonin phase resetting and suppression

    Science.gov (United States)

    Zeitzer, J. M.; Dijk, D. J.; Kronauer, R.; Brown, E.; Czeisler, C.

    2000-01-01

    Ocular exposure to early morning room light can significantly advance the timing of the human circadian pacemaker. The resetting response to such light has a non-linear relationship to illuminance. The dose-response relationship of the human circadian pacemaker to late evening light of dim to moderate intensity has not been well established. Twenty-three healthy young male and female volunteers took part in a 9 day protocol in which a single experimental light exposure6.5 h in duration was given in the early biological night. The effects of the light exposure on the endogenous circadian phase of the melatonin rhythm and the acute effects of the light exposure on plasma melatonin concentration were calculated. We demonstrate that humans are highly responsive to the phase-delaying effects of light during the early biological night and that both the phase resetting response to light and the acute suppressive effects of light on plasma melatonin follow a logistic dose-response curve, as do many circadian responses to light in mammals. Contrary to expectations, we found that half of the maximal phase-delaying response achieved in response to a single episode of evening bright light ( approximately 9000 lux (lx)) can be obtained with just over 1 % of this light (dim room light of approximately 100 lx). The same held true for the acute suppressive effects of light on plasma melatonin concentrations. This indicates that even small changes in ordinary light exposure during the late evening hours can significantly affect both plasma melatonin concentrations and the entrained phase of the human circadian pacemaker.

  4. Response of sensitive human ataxia and resistant T-1 cell lines to accelerated heavy ions

    International Nuclear Information System (INIS)

    Tobias, C.A.; Blakely, E.A.; Chang, P.Y.; Lommel, L.; Roots, R.

    1983-07-01

    The radiation dose responses of fibroblast from a patient with Ataxia telangiectasis (AT-2SF) and an established line of human T-1 cells were studied. Nearly monoenergetic accelerated neon and argon ions were used at the Berkeley Bevalac with various residual range values. The LET of the particles varied from 30 keV/μm to over 1000 keV/μm. All Ataxia survival curves were exponential functions of the dose. Their radiosensitivity reached peak values at 100 to 200 keV/μm. Human T-1 cells have effective sublethal damage repair as has been evidenced by split dose experiments, and they are much more resistant to low LET than to high LET radiation. The repair-misrepair model has been used to interpret these results. We have obtained mathematical expressions that describe the cross sections and inactivation coefficients for both human cell lines as a function of the LET and the type of particle used. The results suggest either that high-LET particles induce a greater number of radiolesions per track or that heavy-ions at high LET induce lesions that kill cells more effectively and that are different from those produced at low LET. We assume that the lesions induced in T-1 and Ataxia cells are qualitatively similar and that each cell line attempts to repair these lesions. The result in most irradiated Ataxia cells, however, is either lethal misrepair or incomplete repair leading to cell death. 63 references, 10 figures, 1 table

  5. Identification of proteins sensitive to thermal stress in human neuroblastoma and glioma cell lines.

    Directory of Open Access Journals (Sweden)

    Guilian Xu

    Full Text Available Heat-shock is an acute insult to the mammalian proteome. The sudden elevation in temperature has far-reaching effects on protein metabolism, leads to a rapid inhibition of most protein synthesis, and the induction of protein chaperones. Using heat-shock in cells of neuronal (SH-SY5Y and glial (CCF-STTG1 lineage, in conjunction with detergent extraction and sedimentation followed by LC-MS/MS proteomic approaches, we sought to identify human proteins that lose solubility upon heat-shock. The two cell lines showed largely overlapping profiles of proteins detected by LC-MS/MS. We identified 58 proteins in detergent insoluble fractions as losing solubility in after heat shock; 10 were common between the 2 cell lines. A subset of the proteins identified by LC-MS/MS was validated by immunoblotting of similarly prepared fractions. Ultimately, we were able to definitively identify 3 proteins as putatively metastable neural proteins; FEN1, CDK1, and TDP-43. We also determined that after heat-shock these cells accumulate insoluble polyubiquitin chains largely linked via lysine 48 (K-48 residues. Collectively, this study identifies human neural proteins that lose solubility upon heat-shock. These proteins may represent components of the human proteome that are vulnerable to misfolding in settings of proteostasis stress.

  6. Human skeletal muscle ceramide content is not a major factor in muscle insulin sensitivity

    DEFF Research Database (Denmark)

    Skovbro, M; Baranowski, M; Skov-Jensen, C

    2008-01-01

    -hyperinsulinaemic clamp was performed for 120 and 90 min for step 1 and step 2, respectively. Muscle biopsies were obtained from vastus lateralis at baseline, and after steps 1 and 2. RESULTS: Glucose infusion rates increased in response to insulin infusion, and significant differences were present between groups (T2D......AIMS/HYPOTHESIS: In skeletal muscle, ceramides may be involved in the pathogenesis of insulin resistance through an attenuation of insulin signalling. This study investigated total skeletal muscle ceramide fatty acid content in participants exhibiting a wide range of insulin sensitivities. METHODS......: The middle-aged male participants (n=33) were matched for lean body mass and divided into four groups: type 2 diabetes (T2D, n=8), impaired glucose tolerance (IGT, n=9), healthy controls (CON, n=8) and endurance-trained (TR, n=8). A two step (28 and 80 mU m(-2) min(-1)) sequential euglycaemic...

  7. Rapid and sensitive multiplex single-tube nested PCR for the identification of five human Plasmodium species.

    Science.gov (United States)

    Saito, Takahiro; Kikuchi, Aoi; Kaneko, Akira; Isozumi, Rie; Teramoto, Isao; Kimura, Masatsugu; Hirasawa, Noriyasu; Hiratsuka, Masahiro

    2018-06-01

    Malaria is caused by five species of Plasmodium in humans. Microscopy is currently used for pathogen detection, requiring considerable training and technical expertise as the parasites are often difficult to differentiate morphologically. Rapid diagnostic tests are as reliable as microscopy and offer faster diagnoses but possess lower detection limits and are incapable of distinguishing among the parasitic species. To improve global health efforts towards malaria control, a rapid, sensitive, species-specific, and economically viable diagnostic method is needed. In this study, we designed a malaria diagnostic method involving a multiplex single-tube nested PCR targeting Plasmodium mitochondrial cytochrome c oxidase III and single-stranded tag hybridization chromatographic printed-array strip. The detection sensitivity was found to be at least 40 times higher than that of agarose gel electrophoresis with ethidium bromide. This system also enables the identification of both single- and mixed-species malaria infections. The assay was validated with 152 Kenyan samples; using nested PCR as the standard, the assay's sensitivity and specificity were 88.7% and 100.0%, respectively. The turnaround time required, from PCR preparation to signal detection, is 90min. Our method should improve the diagnostic speed, treatment efficacy, and control of malaria, in addition to facilitating surveillance within global malaria eradication programs. Copyright © 2018 Elsevier B.V. All rights reserved.

  8. TERRA Expression Levels Do Not Correlate With Telomere Length and Radiation Sensitivity in Human Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Alexandra eSmirnova

    2013-05-01

    Full Text Available Mammalian telomeres are transcribed into long non-coding telomeric RNA molecules (TERRA that seem to play a role in the maintenance of telomere stability. In human cells, CpG island promoters drive TERRA transcription and are regulated by methylation. It was suggested that the amount of TERRA may be related to telomere length. To test this hypothesis we measured telomere length and TERRA levels in single clones isolated from five human cell lines: HeLa (cervical carcinoma, BRC-230 (breast cancer, AKG and GK2 (gastric cancers and GM847 (SV40 immortalized skin fibroblasts. We observed great clonal heterogeneity both in TRF (Terminal Restriction Fragment length and in TERRA levels. However, these two parameters did not correlate with each other. Moreover, cell survival to γ-rays did not show a significant variation among the clones, suggesting that, in this cellular system, the intra-population variability in telomere length and TERRA levels does not influence sensitivity to ionizing radiation. This conclusion was supported by the observation that in a cell line in which telomeres were greatly elongated by the ectopic expression of telomerase, TERRA expression levels and radiation sensitivity were similar to the parental HeLa cell line.

  9. A Sensitive Sensor Cell Line for the Detection of Oxidative Stress Responses in Cultured Human Keratinocytes

    Directory of Open Access Journals (Sweden)

    Ute Hofmann

    2014-06-01

    Full Text Available In the progress of allergic and irritant contact dermatitis, chemicals that cause the generation of reactive oxygen species trigger a heat shock response in keratinocytes. In this study, an optical sensor cell line based on cultured human keratinocytes (HaCaT cells expressing green fluorescent protein (GFP under the control of the stress-inducible HSP70B’ promoter were constructed. Exposure of HaCaT sensor cells to 25 µM cadmium, a model substance for oxidative stress induction, provoked a 1.7-fold increase in total glutathione and a ~300-fold induction of transcript level of the gene coding for heat shock protein HSP70B’. An extract of Arnica montana flowers resulted in a strong induction of the HSP70B’ gene and a pronounced decrease of total glutathione in keratinocytes. The HSP70B’ promoter-based sensor cells conveniently detected cadmium-induced stress using GFP fluorescence as read-out with a limit of detection of 6 µM cadmium. In addition the sensor cells responded to exposure of cells to A. montana extract with induction of GFP fluorescence. Thus, the HaCaT sensor cells provide a means for the automated detection of the compromised redox status of keratinocytes as an early indicator of the development of human skin disorders and could be applied for the prediction of skin irritation in more complex in vitro 3D human skin models and in the development of micro-total analysis systems (µTAS that may be utilized in dermatology, toxicology, pharmacology and drug screenings.

  10. The sensitivity of the human breast to cancer induction by ionizing radiation

    International Nuclear Information System (INIS)

    Mole, R.H.

    1978-01-01

    Available evidence for the induction of cancer in the human breast by small doses of radiation is reviewed. A comparison is made of risk estimates for the frequency of breast cancer in excess of controls, per rad of ionizing radiation, resulting from multiple fluoroscopy, radiotherapy of non-malignant diseases of the breast, or the exposure of Japanese bomb survivors. The significance of the age at exposure is discussed, and consideration is given to the application of the evidence to practical problems in radiography, radiotherapy, screening by mammography, and radiological protection for occupational exposure. (U.K.)

  11. Sensitivity of the human breast to cancer induction by ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Mole, R H [Medical Research Council, Harwell (UK). Radiobiological Research Unit

    1978-06-01

    Available evidence for the induction of cancer in the human breast by small doses of radiation is reviewed. A comparison is made of risk estimates for the frequency of breast cancer in excess of controls, per rad of ionizing radiation, resulting from multiple fluoroscopy, radiotherapy of non-malignant diseases of the breast, or the exposure of Japanese bomb survivors. The significance of the age at exposure is discussed, and consideration is given to the application of the evidence to practical problems in radiography, radiotherapy, screening by mammography, and radiological protection for occupational exposure.

  12. Increased neutralization sensitivity and reduced replicative capacity of human immunodeficiency virus type 1 after short-term in vivo or in vitro passage through chimpanzees

    NARCIS (Netherlands)

    Beaumont, T.; Broersen, S.; van Nuenen, A.; Huisman, H. G.; de Roda Husman, A. M.; Heeney, J. L.; Schuitemaker, H.

    2000-01-01

    Development of disease is extremely rare in chimpanzees when inoculated with either T-cell-line-adapted neutralization-sensitive or primary human immunodeficiency virus type 1 (HIV-1), at first excluding a role for HIV-1 neutralization sensitivity in the clinical course of infection. Interestingly,

  13. Effects of extracellular plaque components on the chlorhexidine sensitivity of strains of Streptococcus mutans and human dental plaque

    International Nuclear Information System (INIS)

    Wolinsky, L.E.; Hume, W.R.

    1985-01-01

    An in vitro study was undertaken to determine the effects of sucrose-derived extracellular plaque components on the sensitivity of selected oral bacteria to chlorhexidine (CX). Cultures of Streptococcus mutans HS-6, OMZ-176, Ingbritt C, 6715-wt13, and pooled human plaque were grown in trypticase soy media with or without 1% sucrose. The sensitivity to CX of bacteria grown in each medium was determined by fixed-time exposure to CX and subsequent measurement of 3 H-thymidine uptake. One-hour exposure to CX at concentrations of 10(-4) M (0.01% w/v) or greater substantially inhibited subsequent cellular division among all the S. mutans strains and human plaque samples tested. An IC50 (the CX concentration which depressed 3 H-thymidine incorporation to 50% of control level) of close to 10(-4) M was noted for S. mutans strains HS-6, OMZ-176, and 6715-wt13 when grown in the presence of sucrose. The same strains grown in cultures without added sucrose showed about a ten-fold greater sensitivity to CX (IC50 close to 10(-5) M). A three-fold difference was noted for S. mutans Ingbritt C. Only a slight increase in the IC50 was noted for the plaque samples cultured in sucrose-containing media, but their threshold for depression of 3 H-thymidine uptake by CX was lower than that for the sucrose-free plaque samples. The study showed that extracellular products confer some protection against CX to the bacteria examined, and provided an explanation for the disparity between clinically-recommended concentrations for plaque suppression and data on in vitro susceptibility

  14. Peripheral erythrocytes decrease upon specific respiratory challenge with grass pollen allergen in sensitized mice and in human subjects.

    Directory of Open Access Journals (Sweden)

    Galateja Jordakieva

    Full Text Available BACKGROUND AND AIMS: Specific hyper-responsiveness towards an allergen and non-specific airway hyperreactivity both impair quality of life in patients with respiratory allergic diseases. We aimed to investigate cellular responses following specific and non-specific airway challenges locally and systemically in i sensitized BALB/c mice challenged with grass pollen allergen Phl p 5, and in ii grass pollen sensitized allergic rhinitis subjects undergoing specific airway challenge in the Vienna Challenge Chamber (VCC. METHODS AND RESULTS: BALB/c mice (n = 20 were intraperitoneally immunized with grass pollen allergen Phl p 5 and afterwards aerosol challenged with either the specific allergen Phl p 5 (n = 10 or the non-specific antigen ovalbumin (OVA (n = 10. A protocol for inducing allergic asthma as well as allergic rhinitis, according to the united airway concept, was used. Both groups of exposed mice showed significantly reduced physical activity after airway challenge. Specific airway challenge further resulted in goblet cell hyperplasia, enhanced mucous secretion, intrapulmonary leukocyte infiltration and lymphoid follicle formation, associated with significant expression of IL-4, IL-5 and IL-13 in splenocytes and also partially in lung tissue. Concerning circulating blood cell dynamics, we observed a significant drop of erythrocyte counts, hemoglobin and hematocrit levels in both mouse groups, challenged with allergen or OVA. A significant decrease in circulating erythrocytes and hematocrit levels after airway challenges with grass pollen allergen was also found in grass pollen sensitized human rhinitis subjects (n = 42 at the VCC. The effects on peripheral leukocyte counts in mice and humans however were opposed, possibly due to the different primary inflammation sites. CONCLUSION: Our data revealed that, besides significant leukocyte dynamics, particularly erythrocytes are involved in acute hypersensitivity reactions to respiratory allergens

  15. Novelty-Sensitive Dopaminergic Neurons in the Human Substantia Nigra Predict Success of Declarative Memory Formation.

    Science.gov (United States)

    Kamiński, Jan; Mamelak, Adam N; Birch, Kurtis; Mosher, Clayton P; Tagliati, Michele; Rutishauser, Ueli

    2018-04-12

    The encoding of information into long-term declarative memory is facilitated by dopamine. This process depends on hippocampal novelty signals, but it remains unknown how midbrain dopaminergic neurons are modulated by declarative-memory-based information. We recorded individual substantia nigra (SN) neurons and cortical field potentials in human patients performing a recognition memory task. We found that 25% of SN neurons were modulated by stimulus novelty. Extracellular waveform shape and anatomical location indicated that these memory-selective neurons were putatively dopaminergic. The responses of memory-selective neurons appeared 527 ms after stimulus onset, changed after a single trial, and were indicative of recognition accuracy. SN neurons phase locked to frontal cortical theta-frequency oscillations, and the extent of this coordination predicted successful memory formation. These data reveal that dopaminergic neurons in the human SN are modulated by memory signals and demonstrate a progression of information flow in the hippocampal-basal ganglia-frontal cortex loop for memory encoding. Copyright © 2018 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  16. Breed differences in dogs sensitivity to human points: a meta-analysis.

    Science.gov (United States)

    Dorey, Nicole R; Udell, Monique A R; Wynne, Clive D L

    2009-07-01

    The last decade has seen a substantial increase in research on the behavioral and cognitive abilities of pet dogs, Canis familiaris. The most commonly used experimental paradigm is the object-choice task in which a dog is given a choice of two containers and guided to the reinforced object by human pointing gestures. We review here studies of this type and attempt a meta-analysis of the available data. In the meta-analysis breeds of dogs were grouped into the eight categories of the American Kennel Club, and into four clusters identified by Parker and Ostrander [Parker, H.G., Ostrander, E.A., 2005. Canine genomics and genetics: running with the pack. PLoS Genet. 1, 507-513] on the basis of a genetic analysis. No differences in performance between breeds categorized in either fashion were identified. Rather, all dog breeds appear to be similarly and highly successful in following human points to locate desired food. We suggest this result could be due to the paucity of data available in published studies, and the restricted range of breeds tested.

  17. Optimization of PIXE-sensitivity for detection of Ti in thin human skin sections

    International Nuclear Information System (INIS)

    Pallon, Jan; Garmer, Mats; Auzelyte, Vaida; Elfman, Mikael; Kristiansson, Per; Malmqvist, Klas; Nilsson, Christer; Shariff, Asad; Wegden, Marie

    2005-01-01

    Modern sunscreens contain particles like TiO 2 having sizes of 25-70 nm and acting as a reflecting substance. For cosmetic reasons the particle size is minimized. Questions have been raised to what degree these nano particles penetrate the skin barrier, and how they do affect the human. The EU funded project 'Quality of skin as a barrier to ultra-fine particles' - NANODERM has started with the purpose to evaluate the possible risks of TiO 2 penetration into vital skin layers. The purpose of the work presented here was to find the optimal conditions for micro-PIXE analysis of Ti in thin skin sections. In the skin region where Ti is expected to be found, the naturally occurring major elements phosphorus, chlorine, sulphur and potassium have steep gradients and thus influence the X-ray background in a non-predictable manner. Based on experimental studies of Ti-exposed human skin sections using proton energies ranging from 1.8-2.55 MeV, the corresponding PIXE detection limits for Ti were calculated. The energy that was found to be the most favourable, 1.9 MeV, was then selected for future studies

  18. Human T-Cell Leukemia Virus I Tax Protein Sensitizes p53-Mutant Cells to DNA Damage

    Science.gov (United States)

    Mihaylova, Valia T.; Green, Allison M.; Khurgel, Moshe; Semmes, Oliver J.; Kupfer, Gary M.

    2018-01-01

    Mutations in p53 are a common cause of resistance of cancers to standard chemotherapy and, thus, treatment failure. Reports have shown that Tax, a human T-cell leukemia virus type I encoded protein that has been associated with genomic instability and perturbation of transcription and cell cycle, sensitizes HeLa cells to UV treatment. The extent to which Tax can sensitize cells and the mechanism by which it exerts its effect are unknown. In this study, we show that Tax sensitizes p53-mutant cells to a broad range of DNA-damaging agents, including mitomycin C, a bifunctional alkylator, etoposide, a topoisomerase II drug, and UV light, but not ionizing radiation, a double-strand break agent, or vinblastine, a tubulin poison. Tax caused hypersensitivity in all p53-deleted cell lines and several, but not all, mutant-expressed p53–containing cell lines, while unexpectedly being protective in p53 wild-type (wt) cells. The effect observed in p53-deleted lines could be reversed for this by transfection of wt p53. We also show that Tax activates a p53-independent proapoptotic program through decreased expression of the retinoblastoma protein and subsequent increased E2F1 expression. The expression of several proapoptotic proteins was also induced by Tax, including Puma and Noxa, culminating in a substantial increase in Bax dimerization. Our results show that Tax can sensitize p53-mutant cells to DNA damage while protecting p53 wt cells, a side benefit that might result in reduced toxicity in normal cells. Such studies hold the promise of a novel adjunctive therapy that could make cancer chemotherapy more effective. PMID:18559532

  19. Sensitivity to sodium arsenite in human melanoma cells depends upon susceptibility to arsenite-induced mitotic arrest

    International Nuclear Information System (INIS)

    McNeely, Samuel C.; Belshoff, Alex C.; Taylor, B. Frazier; Fan, Teresa W-M.; McCabe, Michael J.; Pinhas, Allan R.

    2008-01-01

    Arsenic induces clinical remission in patients with acute promyelocytic leukemia and has potential for treatment of other cancers. The current study examines factors influencing sensitivity to arsenic using human malignant melanoma cell lines. A375 and SK-Mel-2 cells were sensitive to clinically achievable concentrations of arsenite, whereas SK-Mel-3 and SK-Mel-28 cells required supratherapeutic levels for toxicity. Inhibition of glutathione synthesis, glutathione S-transferase (GST) activity, and multidrug resistance protein (MRP) transporter function attenuated arsenite resistance, consistent with studies suggesting that arsenite is extruded from the cell as a glutathione conjugate by MRP-1. However, MRP-1 was not overexpressed in resistant lines and GST-π was only slightly elevated. ICP-MS analysis indicated that arsenite-resistant SK-Mel-28 cells did not accumulate less arsenic than arsenite-sensitive A375 cells, suggesting that resistance was not attributable to reduced arsenic accumulation but rather to intrinsic properties of resistant cell lines. The mode of arsenite-induced cell death was apoptosis. Arsenite-induced apoptosis is associated with cell cycle alterations. Cell cycle analysis revealed arsenite-sensitive cells arrested in mitosis whereas arsenite-resistant cells did not, suggesting that induction of mitotic arrest occurs at lower intracellular arsenic concentrations. Higher intracellular arsenic levels induced cell cycle arrest in the S-phase and G 2 -phase in SK-Mel-3 and SK-Mel-28 cells, respectively. The lack of arsenite-induced mitotic arrest in resistant cell lines was associated with a weakened spindle checkpoint resulting from reduced expression of spindle checkpoint protein BUBR1. These data suggest that arsenite has potential for treatment of solid tumors but a functional spindle checkpoint is a prerequisite for a positive response to its clinical application

  20. Implied motion because of instability in Hokusai Manga activates the human motion-sensitive extrastriate visual cortex: an fMRI study of the impact of visual art.

    Science.gov (United States)

    Osaka, Naoyuki; Matsuyoshi, Daisuke; Ikeda, Takashi; Osaka, Mariko

    2010-03-10

    The recent development of cognitive neuroscience has invited inference about the neurosensory events underlying the experience of visual arts involving implied motion. We report functional magnetic resonance imaging study demonstrating activation of the human extrastriate motion-sensitive cortex by static images showing implied motion because of instability. We used static line-drawing cartoons of humans by Hokusai Katsushika (called 'Hokusai Manga'), an outstanding Japanese cartoonist as well as famous Ukiyoe artist. We found 'Hokusai Manga' with implied motion by depicting human bodies that are engaged in challenging tonic posture significantly activated the motion-sensitive visual cortex including MT+ in the human extrastriate cortex, while an illustration that does not imply motion, for either humans or objects, did not activate these areas under the same tasks. We conclude that motion-sensitive extrastriate cortex would be a critical region for perception of implied motion in instability.

  1. Initial slope of human tumor cell survival curves: its modification by the oxic cell sensitizer beta-arabinofuranosyladenine

    International Nuclear Information System (INIS)

    Chavaudra, N.; Halimi, M.; Parmentier, C.; Gaillard, N.; Grinfeld, S.; Malaise, E.P.

    1989-01-01

    The initial slope of the survival curve, which is a characteristic of each tumor cell line, varies with the histological group of the tumor. It is one of the factors on which clinical radioresponsiveness depends. The DNA dependant DNA polymerase inhibitor beta-ara A acts as an oxic cell sensitizer. This study was carried out on human tumor cell lines to look for a correlation between the degree of radiosensitization induced by beta-ara A and the radiosensitivity of a given cell line. Six human tumor cell lines with different radiosensitivities were used (the survival rate at 2 Gy and D ranged from 20 to 73% and from 1.2 to 3.2 Gy, respectively). beta-ara A had a major toxic effect on all cell lines but this varied greatly from one cell line to another and was concentration dependant; this toxic effect was taken into account when calculating the surviving fractions. For all cell lines, beta-ara A acted as an oxic radiosensitizer and the radiosensitization was concentration dependant. Analysis of the survival curves of the 6 cell lines using the linear quadratic model showed that concentrations of beta-ara A between 200 and 1000 microM induced an increase in the linear component while the quadratic component underwent no systematic change. The sensitizing enhancement ratio (SER) measured from the Ds ratios, varied greatly from one line to another. For example, at a concentration of 500 microM, the extreme values of Ds ratios were 1.5 and 2.6. The radiosensitization is greater, the higher the radiosensitivity of the cell line studied during exponential growth. The results do not favor the use of beta-ara A in the treatment of intrinsically radioresistant human tumors

  2. Initial slope of human tumor cell survival curves: its modification by the oxic cell sensitizer beta-arabinofuranosyladenine

    Energy Technology Data Exchange (ETDEWEB)

    Chavaudra, N.; Halimi, M.; Parmentier, C.; Gaillard, N.; Grinfeld, S.; Malaise, E.P.

    1989-05-01

    The initial slope of the survival curve, which is a characteristic of each tumor cell line, varies with the histological group of the tumor. It is one of the factors on which clinical radioresponsiveness depends. The DNA dependant DNA polymerase inhibitor beta-ara A acts as an oxic cell sensitizer. This study was carried out on human tumor cell lines to look for a correlation between the degree of radiosensitization induced by beta-ara A and the radiosensitivity of a given cell line. Six human tumor cell lines with different radiosensitivities were used (the survival rate at 2 Gy and D ranged from 20 to 73% and from 1.2 to 3.2 Gy, respectively). beta-ara A had a major toxic effect on all cell lines but this varied greatly from one cell line to another and was concentration dependant; this toxic effect was taken into account when calculating the surviving fractions. For all cell lines, beta-ara A acted as an oxic radiosensitizer and the radiosensitization was concentration dependant. Analysis of the survival curves of the 6 cell lines using the linear quadratic model showed that concentrations of beta-ara A between 200 and 1000 microM induced an increase in the linear component while the quadratic component underwent no systematic change. The sensitizing enhancement ratio (SER) measured from the Ds ratios, varied greatly from one line to another. For example, at a concentration of 500 microM, the extreme values of Ds ratios were 1.5 and 2.6. The radiosensitization is greater, the higher the radiosensitivity of the cell line studied during exponential growth. The results do not favor the use of beta-ara A in the treatment of intrinsically radioresistant human tumors.

  3. The effects of instructions on the sensitivity of negatively reinforced human behavior to extinction.

    Science.gov (United States)

    Alessandri, Jérôme; Cançado, Carlos R X

    2017-03-01

    The effects of instructions on the sensitivity of negatively reinforced (escape) behavior to extinction were studied. Initially, responding produced timeouts from pressing a force cell on a variable-ratio (VR) schedule, which was then discontinued (extinction). Based on extinction data, participants were distributed into two groups. Participants in the Persistence Group (for which response rates were low in extinction) were instructed that the experimenter expected them to continue responding in extinction after a second exposure to the VR schedule. Participants in the Extinction group (for which response rates were high in extinction) were instructed that the experimenter expected them to stop responding in extinction. Relative to the condition in which instructions were absent, extinction-response rates increased and decreased, respectively, for participants in the Persistence and Extinction groups. These results replicate and extend to negatively reinforced responding previous findings that showed behavioral control by instructions formulated as explicit experimenter demands or expectations. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Substrate Metabolism and Insulin Sensitivity During Fasting in Obese Human Subjects: Impact of GH Blockade.

    Science.gov (United States)

    Pedersen, Morten Høgild; Svart, Mads Vandsted; Lebeck, Janne; Bidlingmaier, Martin; Stødkilde-Jørgensen, Hans; Pedersen, Steen Bønløkke; Møller, Niels; Jessen, Niels; Jørgensen, Jens O L

    2017-04-01

    Insulin resistance and metabolic inflexibility are features of obesity and are amplified by fasting. Growth hormone (GH) secretion increases during fasting and GH causes insulin resistance. To study the metabolic effects of GH blockade during fasting in obese subjects. Nine obese males were studied thrice in a randomized design: (1) after an overnight fast (control), (2) after 72 hour fasting (fasting), and (3) after 72 hour fasting with GH blockade (pegvisomant) [fasting plus GH antagonist (GHA)]. Each study day consisted of a 4-hour basal period followed by a 2-hour hyperinsulinemic, euglycemic clamp combined with indirect calorimetry, assessment of glucose and palmitate turnover, and muscle and fat biopsies. GH levels increased with fasting (P fasting-induced reduction of serum insulin-like growth factor I was enhanced by GHA (P Fasting increased lipolysis and lipid oxidation independent of GHA, but fasting plus GHA caused a more pronounced suppression of lipid intermediates in response to hyperinsulinemic, euglycemic clamp. Fasting-induced insulin resistance was abrogated by GHA (P Fasting plus GHA also caused elevated glycerol levels and reduced levels of counterregulatory hormones. Fasting significantly reduced the expression of antilipolytic signals in adipose tissue independent of GHA. Suppression of GH activity during fasting in obese subjects reverses insulin resistance and amplifies insulin-stimulated suppression of lipid intermediates, indicating that GH is an important regulator of substrate metabolism, insulin sensitivity, and metabolic flexibility also in obese subjects. Copyright © 2017 by the Endocrine Society

  5. In vitro phagocytosis of methicillin resistant and methicillin sensitive staphylococcus aureus by human polymorphonuclear leucocytes

    International Nuclear Information System (INIS)

    Javed, N.; Tahir, R.; Abbas, A.

    2009-01-01

    Staphylococcus aureus is a gram positive bacterium that causes a number of diseases such as abscesses, infective endocarditis, septic arthritis, etc. It is acquiring resistance against many antibiotics like methicillin; therefore its control is becoming increasingly difficult. Peripheral blood phagocytes particularly polymorphonuclear leucocytes play an important role in the protective mechanisms against these organisms. Phagocytes interact with bacteria and phagocytose these microorganisms to kill them. Phenotypically different isolates of Staphylococcus aureus including methicillin resistant Staphylococcus aureus (MRSA) and methicillin sensitive Staphylococcus aureus (MSSA) were collected from various hospitals of Lahore, Pakistan. Fresh polymorphonuclaer leucocytes were obtained from healthy individuals by centrifugation using Ficol-Hypaque gradient combined with dextran sedimentation. Microbiological method was used for the determination of phagocytic index of phenotypic variants of Staphylococcus aureus. A significant difference was observed between the phagocytic index of both bacterial groups. MSSA group showed the Mean+-SD of 79.46%+-3.9 while MRSA group showed 72.35%+-2.5. Significant difference in phagocytic index indicates that it can be one of the mechanisms of MRSA to evade host immune system as compare to MSSA. (author)

  6. The drug sensitivity and transmission dynamics of human malaria on Nias Island, North Sumatra, Indonesia.

    Science.gov (United States)

    Fryauff, D J; Leksana, B; Masbar, S; Wiady, I; Sismadi, P; Susanti, A I; Nagesha, H S; Syafruddin; Atmosoedjono, S; Bangs, M J; Baird, J K

    2002-07-01

    . Night-biting mosquitoes were surveyed by human landing collections and tested for sporozoite infection. Among the five species of human-biting anophelines collected, Anopheles sundaicus was dominant (68%) and the only species found to be infective--two (1.2%) of 167 females being found carrying P. vivax sporozoites. The risk of malarial infection for humans on Nias was considered high because of the abundance of asymptomatic carriers, the reduced effectiveness of the available antimalarial drugs, and the biting and infection 'rates' of the local An. sundaicus.

  7. PKCδ regulates hepatic insulin sensitivity and hepatosteatosis in mice and humans

    DEFF Research Database (Denmark)

    Bezy, Olivier; Tran, Thien T; Pihlajamäki, Jussi

    2011-01-01

    C57BL/6J and 129S6/Sv (B6 and 129) mice differ dramatically in their susceptibility to developing diabetes in response to diet- or genetically induced insulin resistance. A major locus contributing to this difference has been mapped to a region on mouse chromosome 14 that contains the gene encoding...... tolerance, and reduced hepatosteatosis with aging. Conversely, mice with liver-specific overexpression of PKCδ developed hepatic insulin resistance characterized by decreased insulin signaling, enhanced lipogenic gene expression, and hepatosteatosis. Therefore, changes in the expression and regulation...... of PKCδ between strains of mice and in obese humans play an important role in the genetic risk of hepatic insulin resistance, glucose intolerance, and hepatosteatosis; and thus PKCδ may be a potential target in the treatment of metabolic syndrome....

  8. Glycolipid-Dependent, Protease Sensitive Internalization of Pseudomonas aeruginosa Into Cultured Human Respiratory Epithelial Cells

    Science.gov (United States)

    Emam, Aufaugh; Carter, William G; Lingwood, Clifford

    2010-01-01

    Internalization of PAK strain Pseudomonas aeruginosa into human respiratory epithelial cell lines and HeLa cervical cancer cells in vitro was readily demonstrable via a gentamycin protection assay. Depletion of target cell glycosphingolipids (GSLs) using a glucosyl ceramide synthase inhibitor, P4, completely prevented P. aeruginosa internalization. In contrast, P4 treatment had no effect on the internalization of Salmonella typhimurium into HeLa cells. Internalized P. aeruginosa were within membrane vacuoles, often containing microvesicles, between the bacterium and the limiting membrane. P. aeruginosa internalization was markedly enhanced by target cell pretreatment with the exogenous GSL, deacetyl gangliotetraosyl ceramide (Gg4). Gg4 binds the lipid raft marker, GM1 ganglioside. Target cell pretreatment with TLCK, but not other (serine) protease inhibitors, prevented both P. aeruginosa host cell binding and internalization. NFkB inhibition also prevented internalization. A GSL-containing lipid-raft model of P. aeruginosa host cell binding/internalization is proposed PMID:21270937

  9. Human thiopurine methyltransferase pharmacogenetics: effect of phenotype on sensitivity of cultured lymphocytes to 6-mercaptopurine

    International Nuclear Information System (INIS)

    Van Loon, J.; Weinshilboum, R.

    1986-01-01

    Thiopurine methyltransferase (EC 2.1.1.67, TPMT) catalyzes the S-methylation of thiopurine drugs such as 6-mercaptopurine (6-MP). TPMT activity in human lymphocytes and other tissues is controlled by a common genetic polymorphism. These experiments were designed to study the relationship between TPMT phenotype and the effect of 6-MP on 3 H-thymidine ( 3 H-TdR) incorporation into phytohemaglutinin (PHA) stimulated human peripheral blood lymphocytes. Lymphocytes were obtained from the blood of nine subjects, three subjects with each TPMT phenotype. 6-MP dose response curves were performed at optimal (10 μg/ml) and suboptimal (1 μg/ml) concentrations of PHA. ED50 values for 6-MP with lymphocytes from subjects who genetically lacked TPMT activity were higher than ED50 values for lymphocytes from subjects with genetically intermediate or high TPMT activity. However, ED50 values decreased as level of stimulation increased. Therefore, the effects of 6-MP were studied at a series of PHA concentrations that ranged from 0.1 μg/ml to 10 μg/ml. Lymphocytes from subjects who lacked TPMT activity had significantly higher K/sub ii/ values (1.37 +/- 0.340 μM; mean +/- SEM) for inhibition of 3 H-TdR incorporation by 6-MP than did lymphocytes from subjects with intermediate or high TPMT activity (0.529 +/- 0.068 μM and 0.327 +/- 0.064 μM, respectively, P < .05 for both comparisons)

  10. Human thiopurine methyltransferase pharmacogenetics: effect of phenotype on sensitivity of cultured lymphocytes to 6-mercaptopurine

    Energy Technology Data Exchange (ETDEWEB)

    Van Loon, J.; Weinshilboum, R.

    1986-03-05

    Thiopurine methyltransferase (EC 2.1.1.67, TPMT) catalyzes the S-methylation of thiopurine drugs such as 6-mercaptopurine (6-MP). TPMT activity in human lymphocytes and other tissues is controlled by a common genetic polymorphism. These experiments were designed to study the relationship between TPMT phenotype and the effect of 6-MP on /sup 3/H-thymidine (/sup 3/H-TdR) incorporation into phytohemaglutinin (PHA) stimulated human peripheral blood lymphocytes. Lymphocytes were obtained from the blood of nine subjects, three subjects with each TPMT phenotype. 6-MP dose response curves were performed at optimal (10 ..mu..g/ml) and suboptimal (1 ..mu..g/ml) concentrations of PHA. ED50 values for 6-MP with lymphocytes from subjects who genetically lacked TPMT activity were higher than ED50 values for lymphocytes from subjects with genetically intermediate or high TPMT activity. However, ED50 values decreased as level of stimulation increased. Therefore, the effects of 6-MP were studied at a series of PHA concentrations that ranged from 0.1 ..mu..g/ml to 10 ..mu..g/ml. Lymphocytes from subjects who lacked TPMT activity had significantly higher K/sub ii/ values (1.37 +/- 0.340 ..mu..M; mean +/- SEM) for inhibition of /sup 3/H-TdR incorporation by 6-MP than did lymphocytes from subjects with intermediate or high TPMT activity (0.529 +/- 0.068 ..mu..M and 0.327 +/- 0.064 ..mu..M, respectively, P < .05 for both comparisons).

  11. Sensitivity-encoded (SENSE) proton echo-planar spectroscopic imaging (PEPSI) in the human brain.

    Science.gov (United States)

    Lin, Fa-Hsuan; Tsai, Shang-Yueh; Otazo, Ricardo; Caprihan, Arvind; Wald, Lawrence L; Belliveau, John W; Posse, Stefan

    2007-02-01

    Magnetic resonance spectroscopic imaging (MRSI) provides spatially resolved metabolite information that is invaluable for both neuroscience studies and clinical applications. However, lengthy data acquisition times, which are a result of time-consuming phase encoding, represent a major challenge for MRSI. Fast MRSI pulse sequences that use echo-planar readout gradients, such as proton echo-planar spectroscopic imaging (PEPSI), are capable of fast spectral-spatial encoding and thus enable acceleration of image acquisition times. Combining PEPSI with recent advances in parallel MRI utilizing RF coil arrays can further accelerate MRSI data acquisition. Here we investigate the feasibility of ultrafast spectroscopic imaging at high field (3T and 4T) by combining PEPSI with sensitivity-encoded (SENSE) MRI using eight-channel head coil arrays. We show that the acquisition of single-average SENSE-PEPSI data at a short TE (15 ms) can be accelerated to 32 s or less, depending on the field strength, to obtain metabolic images of choline (Cho), creatine (Cre), N-acetyl-aspartate (NAA), and J-coupled metabolites (e.g., glutamate (Glu) and inositol (Ino)) with acceptable spectral quality and localization. The experimentally measured reductions in signal-to-noise ratio (SNR) and Cramer-Rao lower bounds (CRLBs) of metabolite resonances were well explained by both the g-factor and reduced measurement times. Thus, this technology is a promising means of reducing the scan times of 3D acquisitions and time-resolved 2D measurements. Copyright (c) 2007 Wiley-Liss, Inc.

  12. Reduction of Orc6 expression sensitizes human colon cancer cells to 5-fluorouracil and cisplatin.

    Directory of Open Access Journals (Sweden)

    Elaine J Gavin

    Full Text Available Previous studies from our group have shown that the expression levels of Orc6 were highly elevated in colorectal cancer patient specimens and the induction of Orc6 was associated with 5-fluorouracil (5-FU treatment. The goal of this study was to investigate the molecular and cellular impact of Orc6 in colon cancer. In this study, we use HCT116 (wt-p53 and HCT116 (null-p53 colon cancer cell lines as a model system to investigate the impact of Orc6 on cell proliferation, chemosensitivity and pathways involved with Orc6. We demonstrated that the down regulation of Orc6 sensitizes colon cancer cells to both 5-FU and cisplatin (cis-pt treatment. Decreased Orc6 expression in HCT-116 (wt-p53 cells by RNA interference triggered cell cycle arrest at G1 phase. Prolonged inhibition of Orc6 expression resulted in multinucleated cells in HCT-116 (wt-p53 cell line. Western immunoblot analysis showed that down regulation of Orc6 induced p21 expression in HCT-116 (wt-p53 cells. The induction of p21 was mediated by increased level of phosphorylated p53 at ser-15. By contrast, there is no elevated expression of p21 in HCT-116 (null-p53 cells. Orc6 down regulation also increased the expression of DNA damaging repair protein GADD45beta and reduced the expression level of JNK1. Orc6 may be a potential novel target for future anti cancer therapeutic development in colon cancer.

  13. Suppression of NRF2–ARE activity sensitizes chemotherapeutic agent-induced cytotoxicity in human acute monocytic leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Hui [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Institute of Disease Control and Prevention, Academy of Military Medical Sciences, Beijing (China); Wang, Huihui [School of Public Health, China Medical University, 77 Puhe Road, Shenyang North New Area, Shenyang (China); Xue, Peng [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Key Laboratory of the Public Health Safety, Ministry of Education, School of Public Health, Fudan University, Shanghai (China); Hou, Yongyong [School of Public Health, China Medical University, 77 Puhe Road, Shenyang North New Area, Shenyang (China); Dong, Jian [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Institute of Biology and Medicine, Wuhan University of Science and Technology, Wuhan (China); Zhou, Tong [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Qu, Weidong [Key Laboratory of the Public Health Safety, Ministry of Education, School of Public Health, Fudan University, Shanghai (China); Peng, Shuangqing [Institute of Disease Control and Prevention, Academy of Military Medical Sciences, Beijing (China); Li, Jin; Carmichael, Paul L. [Unilever, Safety & Environmental Assurance Centre, Colworth Science Park, Sharnbrook, Bedfordshire MK44 1LQ (United Kingdom); Nelson, Bud; Clewell, Rebecca; Zhang, Qiang; Andersen, Melvin E. [The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States); Pi, Jingbo, E-mail: jpi@mail.cmu.edu.cn [School of Public Health, China Medical University, 77 Puhe Road, Shenyang North New Area, Shenyang (China); The Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC (United States)

    2016-02-01

    Nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of the antioxidant response element (ARE)-dependent transcription, plays a pivotal role in chemical detoxification in normal and tumor cells. Consistent with previous findings that NRF2–ARE contributes to chemotherapeutic resistance of cancer cells, we found that stable knockdown of NRF2 by lentiviral shRNA in human acute monocytic leukemia (AML) THP-1 cells enhanced the cytotoxicity of several chemotherapeutic agents, including arsenic trioxide (As{sub 2}O{sub 3}), etoposide and doxorubicin. Using an ARE-luciferase reporter expressed in several human and mouse cells, we identified a set of compounds, including isonicotinic acid amides, isoniazid and ethionamide, that inhibited NRF2–ARE activity. Treatment of THP-1 cells with ethionamide, for instance, significantly reduced mRNA expression of multiple ARE-driven genes under either basal or As{sub 2}O{sub 3}-challenged conditions. As determined by cell viability and cell cycle, suppression of NRF2–ARE by ethionamide also significantly enhanced susceptibility of THP-1 and U937 cells to As{sub 2}O{sub 3}-induced cytotoxicity. In THP-1 cells, the sensitizing effect of ethionamide on As{sub 2}O{sub 3}-induced cytotoxicity was highly dependent on NRF2. To our knowledge, the present study is the first to demonstrate that ethionamide suppresses NRF2–ARE signaling and disrupts the transcriptional network of the antioxidant response in AML cells, leading to sensitization to chemotherapeutic agents. - Highlights: • Identification of novel inhibitors of ARE-dependent transcription • Suppression of NRF2–ARE sensitizes THP-1 cells to chemotherapy. • Ethionamide suppresses ARE-dependent transcriptional activity. • Ethionamide and isoniazid increase the cytotoxicity of As{sub 2}O{sub 3} in AML cells. • Sensitization of THP-1 cells to As{sub 2}O{sub 3} toxicity by ethionamide is NRF2-dependent.

  14. Immunoregulatory adherent cells in human tuberculosis: radiation-sensitive antigen-specific suppression by monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kleinhenz, M.E.; Ellner, J.J.

    1985-07-01

    In human tuberculosis, adherent mononuclear cells (AMC) selectively depress in vitro responses to the mycobacterial antigen tuberculin purified protein derivative (PPD). The phenotype of this antigen-specific adherent suppressor cell was characterized by examining the functional activity of adherent cells after selective depletion of sheep erythrocyte-rosetting T cells or OKM1-reactive monocytes. Adherent cell suppression was studied in the (/sup 3/H)thymidine-incorporation microculture assay by using T cells rigorously depleted of T cells with surface receptors for the Fc portion of IgG (T gamma cells) as antigen-responsive cells. PPD-induced (/sup 3/H)thymidine incorporation by these non gamma T cells was uniformly reduced (mean, 42% +/- 10% (SD)) when autologous AMC were added to non gamma T cells at a ratio of 1:2. Antigen-specific suppression by AMC was not altered by depletion of sheep erythrocyte-rosetting T cells or treatment with indomethacin. However, AMC treated with OKM1 and complement or gamma irradiation (1,500 rads) no longer suppressed tuberculin responses in vitro. These studies identify the antigen-specific adherent suppressor cell in tuberculosis as an OKM1-reactive, non-erythrocyte-rosetting monocyte. The radiosensitivity of this monocyte immunoregulatory function may facilitate its further definition.

  15. Immunoregulatory adherent cells in human tuberculosis: radiation-sensitive antigen-specific suppression by monocytes

    International Nuclear Information System (INIS)

    Kleinhenz, M.E.; Ellner, J.J.

    1985-01-01

    In human tuberculosis, adherent mononuclear cells (AMC) selectively depress in vitro responses to the mycobacterial antigen tuberculin purified protein derivative (PPD). The phenotype of this antigen-specific adherent suppressor cell was characterized by examining the functional activity of adherent cells after selective depletion of sheep erythrocyte-rosetting T cells or OKM1-reactive monocytes. Adherent cell suppression was studied in the [ 3 H]thymidine-incorporation microculture assay by using T cells rigorously depleted of T cells with surface receptors for the Fc portion of IgG (T gamma cells) as antigen-responsive cells. PPD-induced [ 3 H]thymidine incorporation by these non gamma T cells was uniformly reduced (mean, 42% +/- 10% [SD]) when autologous AMC were added to non gamma T cells at a ratio of 1:2. Antigen-specific suppression by AMC was not altered by depletion of sheep erythrocyte-rosetting T cells or treatment with indomethacin. However, AMC treated with OKM1 and complement or gamma irradiation (1,500 rads) no longer suppressed tuberculin responses in vitro. These studies identify the antigen-specific adherent suppressor cell in tuberculosis as an OKM1-reactive, non-erythrocyte-rosetting monocyte. The radiosensitivity of this monocyte immunoregulatory function may facilitate its further definition

  16. Sensitivity analysis of a validated subject-specific finite element model of the human craniofacial skeleton.

    Science.gov (United States)

    Szwedowski, T D; Fialkov, J; Whyne, C M

    2011-01-01

    Developing a more complete understanding of the mechanical response of the craniofacial skeleton (CFS) to physiological loads is fundamental to improving treatment for traumatic injuries, reconstruction due to neoplasia, and deformities. Characterization of the biomechanics of the CFS is challenging due to its highly complex structure and heterogeneity, motivating the utilization of experimentally validated computational models. As such, the objective of this study was to develop, experimentally validate, and parametrically analyse a patient-specific finite element (FE) model of the CFS to elucidate a better understanding of the factors that are of intrinsic importance to the skeletal structural behaviour of the human CFS. An FE model of a cadaveric craniofacial skeleton was created from subject-specific computed tomography data. The model was validated based on bone strain measurements taken under simulated physiological-like loading through the masseter and temporalis muscles (which are responsible for the majority of craniofacial physiologic loading due to mastication). The baseline subject-specific model using locally defined cortical bone thicknesses produced the strongest correlation to the experimental data (r2 = 0.73). Large effects on strain patterns arising from small parametric changes in cortical thickness suggest that the very thin bony structures present in the CFS are crucial to characterizing the local load distribution in the CFS accurately.

  17. Analysis of human knee osteoarthritic cartilage using polarization sensitive second harmonic generation microscopy

    Science.gov (United States)

    Kumar, Rajesh; Grønhaug, Kirsten M.; Romijn, Elisabeth I.; Drogset, Jon O.; Lilledahl, Magnus B.

    2014-05-01

    Osteoarthritis is one of the most prevalent joint diseases in the world. Although the cause of osteoarthritis is not exactly clear, the disease results in a degradation of the quality of the articular cartilage including collagen and other extracellular matrix components. We have investigated alterations in the structure of collagen fibers in the cartilage tissue of the human knee using mulitphoton microscopy. Due to inherent high nonlinear susceptibility, ordered collagen fibers present in the cartilage tissue matrix produces strong second harmonic generation (SHG) signals. Significant morphological differences are found in different Osteoarthritic grades of cartilage by SHG microscopy. Based on the polarization analysis of the SHG signal, we find that a few locations of hyaline cartilage (mainly type II collagen) is being replaced by fibrocartilage (mainly type I cartilage), in agreement with earlier literature. To locate the different types and quantify the alteration in the structure of collagen fiber, we employ polarization-SHG microscopic analysis, also referred to as _-tensor imaging. The image analysis of p-SHG image obtained by excitation polarization measurements would represent different tissue constituents with different numerical values at pixel level resolution.

  18. Human Sarcoma growth is sensitive to small-molecule mediated AXIN stabilization.

    Directory of Open Access Journals (Sweden)

    Alessandra De Robertis

    Full Text Available Sarcomas are mesenchymal tumors showing high molecular heterogeneity, reflected at the histological level by the existence of more than fifty different subtypes. Genetic and epigenetic evidences link aberrant activation of the Wnt signaling to growth and progression of human sarcomas. This phenomenon, mainly accomplished by autocrine loop activity, is sustained by gene amplification, over-expression of Wnt ligands and co-receptors or epigenetic silencing of endogenous Wnt antagonists. We previously showed that pharmacological inhibition of Wnt signaling mediated by Axin stabilization produced in vitro and in vivo antitumor activity in glioblastoma tumors. Here, we report that targeting different sarcoma cell lines with the Wnt inhibitor/Axin stabilizer SEN461 produces a less transformed phenotype, as supported by modulation of anchorage-independent growth in vitro. At the molecular level, SEN461 treatment enhanced the stability of the scaffold protein Axin1, a key negative regulator of the Wnt signaling with tumor suppressor function, resulting in downstream effects coherent with inhibition of canonical Wnt signaling. Genetic phenocopy of small molecule Axin stabilization, through Axin1 over-expression, coherently resulted in strong impairment of soft-agar growth. Importantly, sarcoma growth inhibition through pharmacological Axin stabilization was also observed in a xenograft model in vivo in female CD-1 nude mice. Our findings suggest the usefulness of Wnt inhibitors with Axin stabilization activity as a potentialyl clinical relevant strategy for certain types of sarcomas.

  19. Radiation sensitization by CAPE on human HeLa cells of cervical cancer

    International Nuclear Information System (INIS)

    Wang Xiaoqiang; Cao Jianping; Fan Saijun; Zun Wei; Huang Xiaofei; Liu Yang; Chen Xialin; Gong Xiaomei; Peng Xiaomei; Zeng Jing

    2009-01-01

    Objective: To study the radiosensitizing effect of caffic acid phenethyl ester (CAPE) on human cervical cancer HeLa cells. Methods: MTT assay was used to measure the relation between the inhibition effect and CAPE concentrations by CAPE with different concentrations on HeLa cells for 24 hours. HeLa cells were divided into the control and experimental groups, both of which were given 0, 2, 4, 6 and 8 Gy of 60Co γ-irradiation, respectively. The cell clones were counted. Meanwhile HeLa cells were divided into the control, CAPE, irradiation and combination groups. Flow cytometric analysis was adopted to detect the changes of cell cycle distribution induced by CAPE. Results: The inhibition rate of CAPE acting on Hela cells increased with concentrations (F=126. 49 ∼ 3654.88, P 0 ) (1.45 and 1.82 Gy) and the quasi-threshold dose (D q ) (1.89 and 3.21 Gy) of HeLa cells in experimental group decreased comparing with control group, SER was 1.26. Compared with the sole irradiation group, cells in G 2 /M phase of the CAPE group and the sole irradiation group increased (P 2 /M arrest and may be related to the inhibition of the sub-lethal damage repair. (authors)

  20. Radiation induced bystander effects in modification of cellular radio-sensitivity in human cancer cells

    International Nuclear Information System (INIS)

    Pandey, B.N.

    2012-01-01

    Radiation-induced Bystander Effect is manifestation of radiation effects in non-irradiated cells in the population. The phenomenon may have significant implication in risk of radiation induced cancer incidence and outcome of cancer radiotherapy. To understand the bystander interaction in tumor cells, we have studied secretion of diffusible factors from control and irradiated tumor cells of different origin. Our results showed a good correlation between magnitude of secretion of diffusible factors and survival of tumor cells. These diffusible factors are shown to affect proliferation and survival of tumor cells involving regulation of kinases and genes/proteins involved in apoptotic machinery. Our experiments using pharmacological inhibitors showed involvement of activating transcription factor 2 (ATF-2) signaling in survival of tumor cells after treatment with diffusible factors. These factors seem to be involved in exerting radio-resistance in tumor cells. Furthermore, in proton microbeam irradiation studies showed induction of double strand break measured as gH2AX foci in human lung carcinoma cells, which was found to propagate to bystander tumor cells during post-irradiation incubation. Implication of these observations in outcome of cancer radiotherapy scenario would be discussed. (author)

  1. Modification of the radiation sensitivity of human tumour cells by a bis-benzimidazole derivative

    Energy Technology Data Exchange (ETDEWEB)

    Smith, P J; Anderson, C O [Medical Research Council, Cambridge (UK)

    1984-10-01

    A comparison was made of the ability of either X-radiation or a DNA-specific ligand (the vital bis-benzimidazole dye; Hoechst 33342) to induce: cell killing, inhibition of de novo DNA synthesis, DNA strand breakage and the delay of cell division in human colon adenocarcinoma cells in vitro. Unlike radiation-induced cell killing, ligand-induced cytotoxicity appeared to be positively correlated with the extent of inhibition of de novo DNA synthesis-a feature consistent with the persistent binding of ligand molecules to nuclear DNA. Ligand-induced DNA strand-breaks disappeared slowly although ligand-treated cells retained apparently normal capacities to repair discrete radiogenic DNA strand-breaks. Pre-treatment of cells with Hoechst 33342 resulted in a dose-modifying enhancement of radiation resistance not associated with altered dosimetry for strand-break induction. However, radioresistance was accompanied by the protracted retention of cells in the G/sub 2/ phase of the cell cycle. We suggest that the results provide direct evidence that the retention of cells in G/sub 2/ phase is a sparing phenomenon and is triggered by the responses of chromatin domains to the presence of DNA damage.

  2. HepG2 human hepatocarcinomas cells sensitization by endogenous porphyrins

    Science.gov (United States)

    Vonarx-Coinsmann, Veronique; Foultier, Marie-Therese; de Brito, Leonor X.; Morlet, Laurent; Patrice, Thierry

    1995-03-01

    We assessed the ability of the human hepatocarcinoma cell line HepG2 to synthesize PpIX in vitro from exogenous ALA and analyzed ALA-induced toxicity and phototoxicity on this cell line. ALA induced a slight dose-dependent dark toxicity, with 79 and 66% cell survival respectively for ALA 50 and 100 mg/ml after 3-h incubation. Whereas the same treatment followed by laser irradiation (l equals 632 nm, 25 J/sq cm) induced dose-dependent phototoxicity, with 54 and 19% cell survival 24 h after PDT. Whatever the incubation time with ALA, a 3-h delay before light exposure was found optimal to reach a maximal phototoxicity. Photoproducts induced by porphyrin light irradiation absorbed light in the red spectral region at longer wavelengths than did the original porphyrins. The possible enhancement of PDT effects after ALA HepG2 cell incubation was investigated by irradiating cells successively with red light (l equals 632 nm) and light (l equals 650 nm). Total fluence was kept constant at 25 J/sq cm. Phototoxicity was lower when cells were irradiated for increased periods of l equals 650 nm light than with l equals 632 nm light alone. Any photoproducts involved had either a short life or were poorly photoreactive. HepG2 cells, synthesizing enzymes and precursors of endogenous porphyrin synthesis, represent a good in vitro model for experiments using ALA-PpIX-PDT.

  3. Variation in sensitizing effect of caffeine in human tumour cell lines after γ-irradiation

    International Nuclear Information System (INIS)

    Valenzuela, M.T.; Almodovar, M.R. de; Mateos, S.; McMillan, T.J.

    2000-01-01

    We have investigated whether the protective role of the G2 checkpoint has increasing importance when the p53-dependent G1 checkpoint is inactivated. We have studied the differential effect of caffeine by clonogenic assays and flow cytometry in three human tumour cell lines with different functionality of p53 protein. The radiosensitizing effect of caffeine (2 mM) expressed itself as a significant decrease in surviving fraction at 2 Gy and a significant increase in α-values in RT112 and TE671, both with non-functional p53. However, no radiosensitizing effect was seen in cells with a normal p53 function (MCF-7 BUS). Two millimoles of caffeine also caused important changes in the cell cycle progression after irradiation. MCF-7 BUS showed a G1 arrest after irradiation and an early G2 arrest but those cells that reached the second G2 did not arrest significantly. In contrast, TE671 exhibited radiosensitization by caffeine, no G1 arrest, a G2 arrest in those cells irradiated in G2, no significant accumulation in the second G2 but an overall delay in release from the first cell cycle, which could be abrogated by caffeine. RT112 was similar to TE671 except that the emphasis in a G2 arrest was shifted from the block in cells irradiated in G2 to those irradiated at other cell cycle phases. The data presented confirm that p53 status can be a significant determinant of the efficacy of caffeine as radiosensitizer in these tumour cell lines, and document the importance of the G2 checkpoint in this effect. (author)

  4. Monaural and binaural contributions to interaural-level-difference sensitivity in human auditory cortex.

    Science.gov (United States)

    Stecker, G Christopher; McLaughlin, Susan A; Higgins, Nathan C

    2015-10-15

    Whole-brain functional magnetic resonance imaging was used to measure blood-oxygenation-level-dependent (BOLD) responses in human auditory cortex (AC) to sounds with intensity varying independently in the left and right ears. Echoplanar images were acquired at 3 Tesla with sparse image acquisition once per 12-second block of sound stimulation. Combinations of binaural intensity and stimulus presentation rate were varied between blocks, and selected to allow measurement of response-intensity functions in three configurations: monaural 55-85 dB SPL, binaural 55-85 dB SPL with intensity equal in both ears, and binaural with average binaural level of 70 dB SPL and interaural level differences (ILD) ranging ±30 dB (i.e., favoring the left or right ear). Comparison of response functions equated for contralateral intensity revealed that BOLD-response magnitudes (1) generally increased with contralateral intensity, consistent with positive drive of the BOLD response by the contralateral ear, (2) were larger for contralateral monaural stimulation than for binaural stimulation, consistent with negative effects (e.g., inhibition) of ipsilateral input, which were strongest in the left hemisphere, and (3) also increased with ipsilateral intensity when contralateral input was weak, consistent with additional, positive, effects of ipsilateral stimulation. Hemispheric asymmetries in the spatial extent and overall magnitude of BOLD responses were generally consistent with previous studies demonstrating greater bilaterality of responses in the right hemisphere and stricter contralaterality in the left hemisphere. Finally, comparison of responses to fast (40/s) and slow (5/s) stimulus presentation rates revealed significant rate-dependent adaptation of the BOLD response that varied across ILD values. Copyright © 2015. Published by Elsevier Inc.

  5. Weather and place-based human behavior: recreational preferences and sensitivity

    Science.gov (United States)

    de Freitas, C. R.

    2015-01-01

    This study examines the links between biometeorological variables and the behavior of beach recreationists along with their rating of overall weather conditions. To identify and describe significance of on-site atmospheric conditions, two separate forms of response are examined. The first is sensory perception of the immediate atmospheric surround expressed verbally, which was the subject of earlier work. In the research reported here, on-site observations of behavior that reflect the effects of weather and climate are examined. By employing, independently, separate indicators of on-site experience, the reliability of each is examined and interpreted and apparent threshold conditions verified. The study site is King's Beach located on the coast of Queensland, Australia. On-site observations of atmospheric variables and beach user behavior are made for the daylight hours of 45 days spread over a 12-month period. The results show that behavioral data provide reliable and meaningful indications of the significance of the atmospheric environment for leisure. Atmospheric conditions within the zone of acceptability are those that the beach users can readily cope with or modify by a range of minor behavioral adjustments. Optimal weather conditions appear to be those requiring no specific behavioral adjustment. Attendance levels reflect only the outer limits of acceptability of the meteorological environment, while duration of visit enables calibration of levels of approval in so far as it reflects rating of on-site weather within a broad zone of tolerance. In a broad theoretical sense, the results add to an understanding of the relationship between weather and human behavior. This information is potentially useful in effective tourism management and planning.

  6. Beyond classic ecological assessment: The use of functional indices to indicate fish assemblages sensitivity to human disturbance in estuaries.

    Science.gov (United States)

    Teichert, Nils; Lepage, Mario; Lobry, Jérémy

    2018-05-22

    Assessing ecological health of aquatic ecosystems is crucial in the current context of biodiversity loss to guide and prioritize management actions. Although several fish-based indices were developed to assess the ecological status of estuarine ecosystems, they do not provide guidance on the causal responses of communities to disturbances. The functional trait-based approach provides an understanding of how human disturbance affects the composition of biological and ecological traits in assemblages, as well as their consequences for ecosystem functioning. Here, we evaluate the responses of fish assemblages to human disturbance in 30 French estuaries using several taxonomic and functional indices (e.g. diversity, evenness or redundancy). We tested whether these indices can provide additional information on the human impacts and health of assemblages that are not reflected by the ecological indicator (fish-based index ELFI). Results indicated that high values of local human disturbances were associated to a decrease in fish abundance, decrease in species richness and reduced functional redundancy, whereas taxonomic and functional evenness increased. In contrast, the functional richness remained stable suggesting that the functional traits of species removed by stressors were maintained by more tolerant species. Indeed, we found that the local disturbances mainly resulted in a decrease in the proportions of small benthic species feeding on macro-invertebrates, which were dominant in the studied estuaries. Some functional alterations were detected by the fish-based index, but the decline of functional redundancy was not reflected, highlighting a serious concern for management. Indeed, the abrupt collapse of functional redundancy in response to local disturbances can decrease the ability of assemblages to maintain certain species traits in the face of future environmental disturbance, including climate change. From a management perspective, the application of such

  7. Three-site sandwich radioimmunoassay with monoclonal antibodies for a sensitive determination of human alpha-fetoprotein

    International Nuclear Information System (INIS)

    Nomura, M.; Imai, M.; Takahashi, K.; Kumakura, T.; Tachibana, K.; Aoyagi, S.; Usuda, S.; Nakamura, T.; Miyakawa, Y.; Mayumi, M.

    1983-01-01

    Utilizing monoclonal antibodies against human alpha-fetoprotein, 3 distinct antigenic determinants were identified. These antigenic determinants, provisionally designated a, b and c, were arranged in such a manner that the binding of one determinant with the corresponding antibody did not inhibit, or only barely inhibited the binding of antibodies directed to the other 2 determinants. Monoclonal antibodies with 3 different specificities were, therefore, applied to develop a sandwich-type solid-phase radioimmunoassay of the antigen in which wells were coated with anti-a, and radiolabeled anti-b together with radiolabeled anti-c was employed to detect the bound antigen. The 3-site sandwich radioimmunoassay involving 3 different determinants gave a higher sensitivity than 2-site assays in which only anti-b or anti-c was employed as a radiolabeled reagent, because the radioactivity of the 2 labeled antibodies was added on the antigen bound to immobilized anti-a. (Auth.)

  8. Effects of melatonin on prepulse inhibition, habituation and sensitization of the human startle reflex in healthy volunteers

    DEFF Research Database (Denmark)

    Lehtinen, Emilia K; Ucar, Ebru; Glenthøj, Birte Y

    2014-01-01

    Prepulse inhibition of the startle reflex (PPI) is an operational measure of sensorimotor gating, which is demonstrated to be impaired in patients with schizophrenia. In addition, a disruption of the circadian rhythm together with blunted melatonin secretion is regularly found in patients...... with schizophrenia and it is theorized that these may contribute to their attentional deficits. The aim of this study was to assess the effects of acute melatonin on healthy human sensorimotor gating. Twenty-one healthy male volunteers were administered melatonin or placebo after which their levels of PPI were...... assessed. Melatonin significantly reduced startle magnitude and ratings of alertness, but did not influence PPI, nor sensitization and habituation. However, when taking baseline scores in consideration, melatonin significantly increased PPI in low scoring individuals while significantly decreasing...

  9. Sensitivity of human peripheral lymphocyte chromosomes to various X-ray doses and subsequent storage in Plexiglass or glass containers

    International Nuclear Information System (INIS)

    Ivanov, B.; Bulanova, M.; Geogieva, I.

    1979-01-01

    A study was performed to determine whether chromosomal aberrations produced in vitro by various X-ray doses in human lymphocytes were affected by post-irradiation storage of the blood in plastic or glass containers. Following X-ray doses of up to 400 R, the yields of cells with aberrations and the incidence of dicentrics, rings, interstitial deletions, symmetrical changes and chromosome fragments increased with dose. After storage of the irradiated lymphocytes in either Plexiglass or glass, the values for exchange aberrations, deletions and aberrant cells were compared. The only statistically significant difference was a slight increase in the percentage of aberrant cells stored in the plastic containers at the 400 R dose level. It was concluded that plastics appear to have a sensitizing effect on the genetic structure of the peripheral lymphocyte and thus the use of this material to store blood in biological dosimetry studies should be discouraged. (U.K.)

  10. Role of capsaicin- and heat-sensitive afferents in stimulation of acupoint-induced pain and analgesia in humans.

    Science.gov (United States)

    Lei, Jing; Ye, Gang; Wu, Jiang-Tao; Pertovaara, Antti; You, Hao-Jun

    2017-09-01

    We investigated role of capsaicin-sensitive afferents within and without the areas of Zusanli (ST36)/Shangjuxu (ST37) acupoints along the stomach (ST) meridian in the perception and modulation of pain assessed by visual analog scale of pain and its distribution rated by subjects, pressure pain threshold (PPT), and heat pain threshold (HPT) in humans. Compared with the treatment of non-acupoint area, capsaicin (100µg/50µl) administered into either ST36 or ST37 acupoint caused the strongest pain intensity and the most extensive pain distribution, followed by rapid onset, bilateral, long-lasting secondary mechanical hyperalgesia and slower onset secondary heat hypoalgesia (1day after the capsaicin treatment). Between treatments of different acupoints, capsaicin administrated into the ST36 acupoint exhibited the stronger pain intensity and more widespread pain distribution compared with the treatment of ST37 acupoint. A period of 30- to 45-min, but not 15-min, 43°C heating-needle stimulation applied to the ST36 acupoint significantly enhanced the HPT, and had no effect on PPT. Upon trapezius muscle pain elicited by the i.m. injection of 5.8% saline, pre-emptive treatment of the contralateral ST36 acupoint with 43°C heating-needle stimulation alleviated the ongoing muscle pain, reduced painful area, and reversed the decrease in HPT. It is suggested that (1) pain elicited from the acupoint and non-acupoint areas differs significantly, which are supposed to be dependent on the different distributions and contributions of capsaicin-sensitive afferents. (2) Non-painful heat stimulation is a valid approach in prevention of ongoing muscle pain with associated post-effects of peripheral and central sensitization. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

  11. SiRNA-mediated IGF-1R inhibition sensitizes human colon cancer SW480 cells to radiation

    International Nuclear Information System (INIS)

    Yavari, Kamal; Taghikhani, Mohammad; Mesbah-Namin, Seyed A.; Maragheh, Mohammad Ghannadi; Babaei, Mohammad Hosein; Arfaee, Ali Jabbary; Madani, Hossein; Mirzaei, Hamid Reza

    2010-01-01

    Purpose. Insulin like growth factor receptor 1 (IGF-1R) is well-documented to play a key role in radiation response and tumor radiosensitivity, thus offering an attractive clinic drug target to enhance tumor sensitivity to anti-cancer radiotherapy. Material and methods. Human colon carcinoma SW480 cells were transfected with the specific small interference RNA (siRNA) expression vector (pkD-shRNA-IGF-1R-V2) designed to target IGF-1R mRNA. The expression of IGF-1R mRNA and its protein among the transfected and untransfected cells were detected by semi-quantitative RT-PCR and ELISA assay. The changes in cell radiosensitivity were examined by MTT assay. Results. Transfection of mammalian expression vector pkD containing IGF-1R siRNA was shown to reduce IGF-1R mRNA levels by up to 95%. ELISA assay detected a similar inhibition of IGF-1R protein levels in cells transfected with IGF-1R siRNA. SW480 cells transfected with the expression vector for siRNA significantly rendered cells more sensitive to radiation and the highest radiation enhancement ratio was 2.02 ± 0.08. Conclusion. These data provide the first evidence that specific siRNA fragment (pkD-shRNA-IGF-1R-V2) targeting human IGF-1R mRNA is able to enhance colon cancer radiosensitivity. Also results indicated that, combining IGF-1R siRNA and radiation significantly enhances antitumor efficacy compared with either modality alone

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

    Science.gov (United States)

    Ghosal, Abhisek; Sekar, Thillai V.

    2014-01-01

    Biotin is essential for the normal function of pancreatic beta cells. These cells obtain biotin from their surroundings via transport across their cell membrane. Little is known about the uptake mechanism involved, how it is regulated, and how it is affected by internal and external factors. We addressed these issues using the mouse-derived pancreatic beta-TC-6 cells and freshly isolated mouse and human primary pancreatic beta cells as models. The results showed biotin uptake by pancreatic beta-TC-6 cells occurs via a Na+-dependent, carrier-mediated process, that is sensitive to desthiobiotin, as well as to pantothenic acid and lipoate; the process is also saturable as a function of concentration (apparent Km = 22.24 ± 5.5 μM). These cells express the sodium-dependent multivitamin transporter (SMVT), whose knockdown (with doxycycline-inducible shRNA) led to a sever inhibition in biotin uptake. Similarly, uptake of biotin by mouse and human primary pancreatic islets is Na+-dependent and carrier-mediated, and both cell types express SMVT. Biotin uptake by pancreatic beta-TC-6 cells is also adaptively regulated (via transcriptional mechanism) by extracellular substrate level. Chronic treatment of pancreatic beta-TC-6 cells with bacterial lipopolysaccharides (LPS) leads to inhibition in biotin uptake. This inhibition is mediated via a Toll-Like receptor 4-mediated process and involves a decrease in membrane expression of SMVT. These findings show, for the first time, that pancreatic beta cells/islets take up biotin via a specific and regulated carrier-mediated process, and that the process is sensitive to the effect of LPS. PMID:24904078

  13. Long-term exposure to abnormal glucose levels alters drug metabolism pathways and insulin sensitivity in primary human hepatocytes

    Science.gov (United States)

    Davidson, Matthew D.; Ballinger, Kimberly R.; Khetani, Salman R.

    2016-06-01

    Hyperglycemia in type 2 diabetes mellitus has been linked to non-alcoholic fatty liver disease, which can progress to inflammation, fibrosis/cirrhosis, and hepatocellular carcinoma. Understanding how chronic hyperglycemia affects primary human hepatocytes (PHHs) can facilitate the development of therapeutics for these diseases. Conversely, elucidating the effects of hypoglycemia on PHHs may provide insights into how the liver adapts to fasting, adverse diabetes drug reactions, and cancer. In contrast to declining PHH monocultures, micropatterned co-cultures (MPCCs) of PHHs and 3T3-J2 murine embryonic fibroblasts maintain insulin-sensitive glucose metabolism for several weeks. Here, we exposed MPCCs to hypo-, normo- and hyperglycemic culture media for ~3 weeks. While albumin and urea secretion were not affected by glucose level, hypoglycemic MPCCs upregulated CYP3A4 enzyme activity as compared to other glycemic states. In contrast, hyperglycemic MPCCs displayed significant hepatic lipid accumulation in the presence of insulin, while also showing decreased sensitivity to insulin-mediated inhibition of glucose output relative to a normoglycemic control. In conclusion, we show for the first time that PHHs exposed to hypo- and hyperglycemia can remain highly functional, but display increased CYP3A4 activity and selective insulin resistance, respectively. In the future, MPCCs under glycemic states can aid in novel drug discovery and mechanistic investigations.

  14. Use of γ-H2AX Foci Assay on Human Peripheral Blood Lymphocytes as Sensitive Biomarker of Exposure

    International Nuclear Information System (INIS)

    Gajski, G.; Garaj-Vrhovac, V.; Geric, M.; Filipic, M.; Nunic, J.; Straser, A.; Zegura, B.

    2013-01-01

    In modern medicine, it is impossible to imagine diagnostics and treatments without equipment that emit radiation (X-ray, CT, PET, etc.). At the same time there is a need to minimize the amount of radiation that the patient will gain during such medical examination. In that manner ALARA (As Low As Reasonably Achievable) principle and dosimetry are the bases of assuring patients safety. The induction of gamma phosphorylated H2AX histone is newly developed tool in biodosimetry, which is more sensitive for the detection of radiation caused DNA damage than currently used micronucleus and comet assay. Gamma phosphorylation of H2AX histone is a consequence of DNA double strand breaks and its role is to trigger the DNA repair mechanisms. In this study, we tested the effect of 2 and 4 Gy X-rays on human peripheral blood lymphocytes from two healthy volunteers using γ-H2AX foci assay. The FITC signal from labelled antibodies was monitored using flow cytometry and clearly demonstrated the difference in control samples and irradiated samples. There was also the difference between the exposed blood samples from the two volunteers. The results of present study reveal new sensitive method that is capable of detecting changes in DNA when exposed to different doses of radiation, and thus potentially optimizing the ALARA principle.(author)

  15. Development of a polymerase chain reaction applicable to rapid and sensitive detection of Clonorchis sinensis eggs in human stool samples

    Science.gov (United States)

    Cho, Pyo Yun; Na, Byoung-Kuk; Mi Choi, Kyung; Kim, Jin Su; Cho, Shin-Hyeong; Lee, Won-Ja; Lim, Sung-Bin; Cha, Seok Ho; Park, Yun-Kyu; Pak, Jhang Ho; Lee, Hyeong-Woo; Hong, Sung-Jong; Kim, Tong-Soo

    2013-01-01

    Microscopic examination of eggs of parasitic helminths in stool samples has been the most widely used classical diagnostic method for infections, but tiny and low numbers of eggs in stool samples often hamper diagnosis of helminthic infections with classical microscopic examination. Moreover, it is also difficult to differentiate parasite eggs by the classical method, if they have similar morphological characteristics. In this study, we developed a rapid and sensitive polymerase chain reaction (PCR)-based molecular diagnostic method for detection of Clonorchis sinensis eggs in stool samples. Nine primers were designed based on the long-terminal repeat (LTR) of C. sinensis retrotransposon1 (CsRn1) gene, and seven PCR primer sets were paired. Polymerase chain reaction with each primer pair produced specific amplicons for C. sinensis, but not for other trematodes including Metagonimus yokogawai and Paragonimus westermani. Particularly, three primer sets were able to detect 10 C. sinensis eggs and were applicable to amplify specific amplicons from DNA samples purified from stool of C. sinensis-infected patients. This PCR method could be useful for diagnosis of C. sinensis infections in human stool samples with a high level of specificity and sensitivity. PMID:23916334

  16. Age-Associated Impairments in Mitochondrial ADP Sensitivity Contribute to Redox Stress in Senescent Human Skeletal Muscle

    Directory of Open Access Journals (Sweden)

    Graham P. Holloway

    2018-03-01

    Full Text Available Summary: It remains unknown if mitochondrial bioenergetics are altered with aging in humans. We established an in vitro method to simultaneously determine mitochondrial respiration and H2O2 emission in skeletal muscle tissue across a range of biologically relevant ADP concentrations. Using this approach, we provide evidence that, although the capacity for mitochondrial H2O2 emission is not increased with aging, mitochondrial ADP sensitivity is impaired. This resulted in an increase in mitochondrial H2O2 and the fraction of electron leak to H2O2, in the presence of virtually all ADP concentrations examined. Moreover, although prolonged resistance training in older individuals increased muscle mass, strength, and maximal mitochondrial respiration, exercise training did not alter H2O2 emission rates in the presence of ADP, the fraction of electron leak to H2O2, or the redox state of the muscle. These data establish that a reduction in mitochondrial ADP sensitivity increases mitochondrial H2O2 emission and contributes to age-associated redox stress. : Holloway et al. show that an inability of ADP to decrease mitochondrial reactive oxygen species emission contributes to redox stress in skeletal muscle tissue of older individuals and that this process is not recovered following prolonged resistance-type exercise training, despite the general benefits of resistance training for muscle health. Keywords: mitochondria, aging, muscle, ROS, H2O2, ADP, respiration, bioenergetics, exercise, resistance training

  17. Construction and expression of secreting type human TRAIL gene vector mediated by hypoxia/radiation double sensitive promoter

    International Nuclear Information System (INIS)

    Yang Yanming; Jia Xiaojing; Qu Yaqin; Li Yanbo

    2009-01-01

    Objective: To construct secreting type human TRAIL (shTRAIL) gene vector pcDNA3.1-HRE/Egr1-shTRAIL mediated by hypoxia/radiation double sensitive promoter, and observe the effect of hypoxia and radiation on shTRAIL. Methods: HRE upper and lower strands were gotten by chemical synthesis, double strands HRE was gotten by PCR; pMD19T-Egr1 was digested by Sac I and Hind III, then Egr1 was obtained, pshuttle-shTRAIL was digested by Kpn I and BamH I, then shTRAIL was obtained; HRE/Egr1 double sensitive promoter mediated shTRAIL expression vector pcDNA3.1-HRE/Egr1-shTRAIL was constructed by gene recombination technique, it was identified correctly by enzyme digestion, PCR and sequencing. A549 cells were divided into normal, hypoxia (0.1%), irradiation (6 Gy) and hypoxia + irradiation groups. Results: After enzyme digestion by BamH I and Sma I, the fragments which lengths were 1284 bp and 4 998 bp, 2 292 bp and 3 990 bp were obtained; the vector was amplified by PCR with Egr1 and shTRAIL primer, the products which lengthens were 469 bp and 820 bp were obtained; pcDNA3.1-HRE/Egr1-shTRAIL was sequenced, the result was same to designed, this demonstrated that the construction was right. The vectors were transfected into A549 cells of adenocarcinoma of lung, the expression levels of shTRAIL mRNA and protein were increased after treated with hypoxia and radiation, it had statistically significant differences compared with normal group (P<0.05), and when they were combinated, the effect was more obvious. Conclusion: Secreting type human TRAIL gene vector pcDNA3.1-HRE/Egr1-shTRAIL mediated by hypoxia/radiation double sensitive promoter is constructed successfully, and hypoxia and radiation could increase the expression of TRAIL, and they have synergetic effect. (authors)

  18. Regions of mid-level human visual cortex sensitive to the global coherence of local image patches.

    Science.gov (United States)

    Mannion, Damien J; Kersten, Daniel J; Olman, Cheryl A

    2014-08-01

    The global structural arrangement and spatial layout of the visual environment must be derived from the integration of local signals represented in the lower tiers of the visual system. This interaction between the spatially local and global properties of visual stimulation underlies many of our visual capacities, and how this is achieved in the brain is a central question for visual and cognitive neuroscience. Here, we examine the sensitivity of regions of the posterior human brain to the global coordination of spatially displaced naturalistic image patches. We presented observers with image patches in two circular apertures to the left and right of central fixation, with the patches drawn from either the same (coherent condition) or different (noncoherent condition) extended image. Using fMRI at 7T (n = 5), we find that global coherence affected signal amplitude in regions of dorsal mid-level cortex. Furthermore, we find that extensive regions of mid-level visual cortex contained information in their local activity pattern that could discriminate coherent and noncoherent stimuli. These findings indicate that the global coordination of local naturalistic image information has important consequences for the processing in human mid-level visual cortex.

  19. Small protease sensitive oligomers of PrPSc in distinct human prions determine conversion rate of PrP(C.

    Directory of Open Access Journals (Sweden)

    Chae Kim

    Full Text Available The mammalian prions replicate by converting cellular prion protein (PrP(C into pathogenic conformational isoform (PrP(Sc. Variations in prions, which cause different disease phenotypes, are referred to as strains. The mechanism of high-fidelity replication of prion strains in the absence of nucleic acid remains unsolved. We investigated the impact of different conformational characteristics of PrP(Sc on conversion of PrP(C in vitro using PrP(Sc seeds from the most frequent human prion disease worldwide, the Creutzfeldt-Jakob disease (sCJD. The conversion potency of a broad spectrum of distinct sCJD prions was governed by the level, conformation, and stability of small oligomers of the protease-sensitive (s PrP(Sc. The smallest most potent prions present in sCJD brains were composed only of∼20 monomers of PrP(Sc. The tight correlation between conversion potency of small oligomers of human sPrP(Sc observed in vitro and duration of the disease suggests that sPrP(Sc conformers are an important determinant of prion strain characteristics that control the progression rate of the disease.

  20. Antioxidants keep the potentially probiotic but highly oxygen-sensitive human gut bacterium Faecalibacterium prausnitzii alive at ambient air.

    Directory of Open Access Journals (Sweden)

    M Tanweer Khan

    Full Text Available The beneficial human gut microbe Faecalibacterium prausnitzii is a 'probiotic of the future' since it produces high amounts of butyrate and anti-inflammatory compounds. However, this bacterium is highly oxygen-senstive, making it notoriously difficult to cultivate and preserve. This has so far precluded its clinical application in the treatment of patients with inflammatory bowel diseases. The present studies were therefore aimed at developing a strategy to keep F. prausnitzii alive at ambient air. Our previous research showed that F. prausnitzii can survive in moderately oxygenized environments like the gut mucosa by transfer of electrons to oxygen. For this purpose, the bacterium exploits extracellular antioxidants, such as riboflavin and cysteine, that are abundantly present in the gut. We therefore tested to what extent these antioxidants can sustain the viability of F. prausnitzii at ambient air. The present results show that cysteine can facilitate the survival of F. prausnitzii upon exposure to air, and that this effect is significantly enhanced the by addition of riboflavin and the cryoprotectant inulin. The highly oxygen-sensitive gut bacterium F. prausnitzii can be kept alive at ambient air for 24 h when formulated with the antioxidants cysteine and riboflavin plus the cryoprotectant inulin. Improved formulations were obtained by addition of the bulking agents corn starch and wheat bran. Our present findings pave the way towards the biomedical exploitation of F. prausnitzii in redox-based therapeutics for treatment of dysbiosis-related inflammatory disorders of the human gut.

  1. High-sensitivity simultaneous liquid chromatography-tandem mass spectrometry assay of ethinyl estradiol and levonorgestrel in human plasma

    Institute of Scientific and Technical Information of China (English)

    Abhishek Gandhi; Swati Guttikar; Priti Trivedi

    2015-01-01

    A sensitive and simultaneous liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of ethinyl estradiol and levonorgestrel. The analytes were extracted with methyl-tert-butyl ether: n-hexane (50:50, v/v) solvent mixture, followed by dansyl derivatization. The chromatographic separation was performed on a Kinetex C18 (50 mm × 4.6 mm, 2.6μm) column with a mobile phase of 0.1% (v/v) formic acid in water and acetonitrile in gradient composition. The mass transitions were monitored in electrospray positive ionization mode. The assay exhibited a linear range of 0.100-20.0 ng/mL for levonorgestrel and 4.00-500 pg/mL for ethinyl estradiol in human plasma. A run time of 9.0 min for each sample made it possible to analyze a throughput of more than 100 samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic and bioequivalence studies.

  2. Rapid, simple and highly sensitive LC-ESI-MS/MS method for the quantification of tamsulosin in human plasma.

    Science.gov (United States)

    Ramakrishna, N V S; Vishwottam, K N; Manoj, S; Koteshwara, M; Wishu, S; Varma, D P

    2005-12-01

    A simple, rapid, sensitive and specific liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of tamsulosin (I), a highly selective alpha1-adrenoceptor antagonist used for the treatment of patients with symptomatic benign prostatic hyperplasia. The analyte and internal standard, mosapride (II) were extracted by liquid-liquid extraction with diethyl ether-dichloromethane (70:30, v/v) using a Glas-Col Multi-Pulse Vortexer. The chromatographic separation was performed on a reverse phase Waters symmetry C18 column with a mobile phase of 0.03% formic acid-acetonitrile (30:70, v/v). The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 409.1 solidus in circle 228.1 and m/z 422.3 solidus in circle 198.3 were used to measure I and II, respectively. The assay exhibited a linear dynamic range of 0.1-50.0 ng/mL for tamsulosin in human plasma. The lower limit of quantitation was 100 pg/mL with a relative standard deviation of less than 10%. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. A run time of 2.0 min for each sample made it possible to analyze a throughput of more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. (c) 2005 John Wiley & Sons, Ltd.

  3. Docetaxel modulates the delayed rectifier potassium current (IK) and ATP-sensitive potassium current (IKATP) in human breast cancer cells.

    Science.gov (United States)

    Sun, Tao; Song, Zhi-Guo; Jiang, Da-Qing; Nie, Hong-Guang; Han, Dong-Yun

    2015-04-01

    Ion channel expression and activity may be affected during tumor development and cancer growth. Activation of potassium (K(+)) channels in human breast cancer cells is reported to be involved in cell cycle progression. In this study, we investigated the effects of docetaxel on the delayed rectifier potassium current (I K) and the ATP-sensitive potassium current (I KATP) in two human breast cancer cell lines, MCF-7 and MDA-MB-435S, using the whole-cell patch-clamp technique. Our results show that docetaxel inhibited the I K and I KATP in both cell lines in a dose-dependent manner. Compared with the control at a potential of +60 mV, treatment with docetaxel at doses of 0.1, 1, 5, and 10 µM significantly decreased the I K in MCF-7 cells by 16.1 ± 3.5, 30.2 ± 5.2, 42.5 ± 4.3, and 46.4 ± 9% (n = 5, P < 0.05), respectively and also decreased the I KATP at +50 mV. Similar results were observed in MDA-MB-435S cells. The G-V curves showed no significant changes after treatment of either MCF-7 or MDA-MB-435S cells with 10 μM docetaxel. The datas indicate that the possible mechanisms of I K and I KATP inhibition by docetaxel may be responsible for its effect on the proliferation of human breast cancer cells.

  4. Monocrotophos induces the expression and activity of xenobiotic metabolizing enzymes in pre-sensitized cultured human brain cells.

    Directory of Open Access Journals (Sweden)

    Vinay K Tripathi

    Full Text Available The expression and metabolic profile of cytochrome P450s (CYPs is largely missing in human brain due to non-availability of brain tissue. We attempted to address the issue by using human brain neuronal (SH-SY5Y and glial (U373-MG cells. The expression and activity of CYP1A1, 2B6 and 2E1 were carried out in the cells exposed to CYP inducers viz., 3-methylcholanthrene (3-MC, cyclophosphamide (CPA, ethanol and known neurotoxicant- monocrotophos (MCP, a widely used organophosphorous pesticide. Both the cells show significant induction in the expression and CYP-specific activity against classical inducers and MCP. The induction level of CYPs was comparatively lower in MCP exposed cells than cells exposed to classical inducers. Pre-exposure (12 h of cells to classical inducers significantly added the MCP induced CYPs expression and activity. The findings were concurrent with protein ligand docking studies, which show a significant modulatory capacity of MCP by strong interaction with CYP regulators-CAR, PXR and AHR. Similarly, the known CYP inducers- 3-MC, CPA and ethanol have also shown significantly high docking scores with all the three studied CYP regulators. The expression of CYPs in neuronal and glial cells has suggested their possible association with the endogenous physiology of the brain. The findings also suggest the xenobiotic metabolizing capabilities of these cells against MCP, if received a pre-sensitization to trigger the xenobiotic metabolizing machinery. MCP induced CYP-specific activity in neuronal cells could help in explaining its effect on neurotransmission, as these CYPs are known to involve in the synthesis/transport of the neurotransmitters. The induction of CYPs in glial cells is also of significance as these cells are thought to be involved in protecting the neurons from environmental insults and safeguard them from toxicity. The data provide better understanding of the metabolizing capability of the human brain cells against

  5. A mechanics approach to the study of pressure sensitive adhesives and human skin for transdermal drug delivery applications

    Science.gov (United States)

    Taub, Marc Barry

    Transdermal drug delivery is an alternative approach to the systemic delivery of pharmaceuticals where drugs are administered through the skin and absorbed percutaneously. This method of delivery offers several advantages over more traditional routes; most notably, the avoidance of the fast-pass metabolism of the liver and gut, the ability to offer controlled release rates, and the possibility for novel devices. Pressure sensitive adhesives (PSAs) are used to bond transdermal drug delivery devices to the skin because of their good initial and long-term adhesion, clean removability, and skin and drug compatibility. However, an understanding of the mechanics of adhesion to the dermal layer, together with quantitative and reproducible test methods for measuring adhesion, have been lacking. This study utilizes a mechanics-based approach to quantify the interfacial adhesion of PSAs bonded to selected substrates, including human dermal tissue. The delamination of PSA layers is associated with cavitation in the PSA followed by the formation of an extensive cohesive zone behind the debond tip. A quantitative metrology was developed to assess the adhesion and delamination of PSAs, such that it could be possible to easily distinguish between the adhesive characteristics of different PSA compositions and to provide a quantitative basis from which the reliability of adhesive layers bonded to substrates could be studied. A mechanics-based model was also developed to predict debonding in terms of the relevant energy dissipation mechanisms active during this process. As failure of transdermal devices may occur cohesively within the PSA layer, adhesively at the interface between the PSA and the skin, or cohesively between the corneocytes that comprise the outermost layer of the skin, it was also necessary to explore the mechanical and fracture properties of human skin. The out-of-plane delamination of corneocytes was studied by determining the strain energy release rate during

  6. Pharmacologic requirements for obtaining sensitization of human tumor cells in vitro to combined 5-fluorouracil or ftorafur and x rays

    International Nuclear Information System (INIS)

    Byfield, J.E.; Calabro-Jones, P.; Klisak, I.; Kulhanian, F.

    1982-01-01

    The combined effects of X ray and 5-fluorouracil (5-FU) in tissue culture have been studied using two human adenocarcinoma lines (HeLa and HT-29 cells). Both showed similar sensitivities to 5-FU, HeLa cells appearing somewhat more resistant to higher concentrations. Combined treatment of both cell types with 5-FU and X rays led to a time-dependent enhancement of cell killing (''radiosensitization''). Only post-radiation incubation had any effect, prior exposure to 5-FU being strictly additive. Enhanced cell killing by combined 5-FU and X rays could not be explained by either the infliction of additional acute damage in the immediate post-radiation period or an inhibitory effect of 5-FU on the repair of sub-lethal X ray injury. Rather, the enhanced cytotoxicity proved to be dependent on a damage expressed in time periods exceeding the duration of a cell doubling time in vitro. Overall, the data equally suggest that X rays may sensitize the cells to 5-FU. The enhanced cell killing is maximized if the cells are continuously exposed to 5-FU for 48 hours following the X ray exposure. These results indicate that clinical treatment regimens might be useful in evaluating 5-FU infusional scheduling in accordance with these unique requirements, which are not met by conventional bolus 5-FU and X ray therapy fractionation regimens. Ftorafur, a drug proposed to act as a slow release form of 5-FU, was found to show limited cytotoxic potential in vitro and did not significantly enhance cell killing after X ray exposure

  7. Cys34-cysteinylated human serum albumin is a sensitive plasma marker in oxidative stress-related chronic diseases.

    Directory of Open Access Journals (Sweden)

    Kohei Nagumo

    Full Text Available The degree of oxidized cysteine (Cys 34 in human serum albumin (HSA, as determined by high performance liquid chromatography (HPLC, is correlated with oxidative stress related pathological conditions. In order to further characterize the oxidation of Cys34-HSA at the molecular level and to develop a suitable analytical method for a rapid and sensitive clinical laboratory analysis, the use of electrospray ionization time-of-flight mass spectrometer (ESI-TOFMS was evaluated. A marked increase in the cysteinylation of Cys34 occurs in chronic liver and kidney diseases and diabetes mellitus. A significant positive correlation was observed between the Cys-Cys34-HSA fraction of plasma samples obtained from 229 patients, as determined by ESI-TOFMS, and the degree of oxidized Cys34-HSA determined by HPLC. The Cys-Cys34-HSA fraction was significantly increased with the progression of liver cirrhosis, and was reduced by branched chain amino acids (BCAA treatment. The changes in the Cys-Cys34-HSA fraction were significantly correlated with the alternations of the plasma levels of advanced oxidized protein products, an oxidative stress marker for proteins. The binding ability of endogenous substances (bilirubin and tryptophan and drugs (warfarin and diazepam to HSA purified from chronic liver disease patients were significantly suppressed but significantly improved by BCAA supplementation. Interestingly, the changes in this physiological function of HSA in chronic liver disease were correlated with the Cys-Cys34-HSA fraction. In conclusion, ESI-TOFMS is a suitable high throughput method for the rapid and sensitive quantification of Cys-Cys34-HSA in a large number of samples for evaluating oxidative stress related chronic disease progression or in response to a treatment.

  8. Regulation of the oxidative balance with coenzyme Q10 sensitizes human glioblastoma cells to radiation and temozolomide.

    Science.gov (United States)

    Frontiñán-Rubio, Javier; Santiago-Mora, Raquel María; Nieva-Velasco, Consuelo María; Ferrín, Gustavo; Martínez-González, Alicia; Gómez, María Victoria; Moreno, María; Ariza, Julia; Lozano, Eva; Arjona-Gutiérrez, Jacinto; Gil-Agudo, Antonio; De la Mata, Manuel; Pesic, Milica; Peinado, Juan Ramón; Villalba, José M; Pérez-Romasanta, Luis; Pérez-García, Víctor M; Alcaín, Francisco J; Durán-Prado, Mario

    2018-05-18

    To investigate how the modulation of the oxidative balance affects cytotoxic therapies in glioblastoma, in vitro. Human glioblastoma U251 and T98 cells and normal astrocytes C8D1A were loaded with coenzyme Q10 (CoQ). Mitochondrial superoxide ion (O 2 - ) and H 2 O 2 were measured by fluorescence microscopy. OXPHOS performance was assessed in U251 cells with an oxytherm Clark-type electrode. Radio- and chemotherapy cytotoxicity was assessed by immunostaining of γH2AX (24 h), annexin V and nuclei morphology, at short (72 h) and long (15 d) time. Hif-1α, SOD1, SOD2 and NQO1 were determined by immunolabeling. Catalase activity was measured by classic enzymatic assay. Glutathione levels and total antioxidant capacity were quantified using commercial kits. CoQ did not affect oxygen consumption but reduced the level of O 2 - and H 2 O 2 while shifted to a pro-oxidant cell status mainly due to a decrease in catalase activity and SOD2 level. Hif-1α was dampened, echoed by a decrease lactate and several key metabolites involved in glutathione synthesis. CoQ-treated cells were twofold more sensitive than control to radiation-induced DNA damage and apoptosis in short and long-term clonogenic assays, potentiating TMZ-induced cytotoxicity, without affecting non-transformed astrocytes. CoQ acts as sensitizer for cytotoxic therapies, disarming GBM cells, but not normal astrocytes, against further pro-oxidant injuries, being potentially useful in clinical practice for this fatal pathology. Copyright © 2018 Elsevier B.V. All rights reserved.

  9. Radioecological sensitivity

    International Nuclear Information System (INIS)

    Howard, Brenda J.; Strand, Per; Assimakopoulos, Panayotis

    2003-01-01

    After the release of radionuclide into the environment it is important to be able to readily identify major routes of radiation exposure, the most highly exposed individuals or populations and the geographical areas of most concern. Radioecological sensitivity can be broadly defined as the extent to which an ecosystem contributes to an enhanced radiation exposure to Man and biota. Radioecological sensitivity analysis integrates current knowledge on pathways, spatially attributes the underlying processes determining transfer and thereby identifies the most radioecologically sensitive areas leading to high radiation exposure. This identifies where high exposure may occur and why. A framework for the estimation of radioecological sensitivity with respect to humans is proposed and the various indicators by which it can be considered have been identified. These are (1) aggregated transfer coefficients (Tag), (2) action (and critical) loads, (3) fluxes and (4) individual exposure of humans. The importance of spatial and temporal consideration of all these outputs is emphasized. Information on the extent of radionuclide transfer and exposure to humans at different spatial scales is needed to reflect the spatial differences which can occur. Single values for large areas, such as countries, can often mask large variation within the country. Similarly, the relative importance of different pathways can change with time and therefore assessments of radiological sensitivity are needed over different time periods after contamination. Radioecological sensitivity analysis can be used in radiation protection, nuclear safety and emergency preparedness when there is a need to identify areas that have the potential of being of particular concern from a risk perspective. Prior identification of radioecologically sensitive areas and exposed individuals improve the focus of emergency preparedness and planning, and contribute to environmental impact assessment for future facilities. The

  10. The implications of non-linear biological oscillations on human electrophysiology for electrohypersensitivity (EHS) and multiple chemical sensitivity (MCS).

    Science.gov (United States)

    Sage, Cindy

    2015-01-01

    maintenance; and resilience can be compromised. Electrohypersensitivity can be caused by successive assaults on human bioelectrochemical dynamics from exogenous electromagnetic fields (EMF) and RFR or a single acute exposure. Once sensitized, further exposures are widely reported to cause reactivity to lower and lower intensities of EMF/RFR, at which point thousand-fold lower levels can cause adverse health impacts to the electrosensitive person. Electrohypersensitivity (EHS) can be a precursor to, or linked with, multiple chemical sensitivity (MCS) based on reports of individuals who first develop one condition, then rapidly develop the other. Similarity of chemical biomarkers is seen in both conditions [histamines, markers of oxidative stress, auto-antibodies, heat shock protein (HSP), melatonin markers and leakage of the blood-brain barrier]. Low intensity pulsed microwave activation of voltage-gated calcium channels (VGCCs) is postulated as a mechanism of action for non-thermal health effects.

  11. Neuronal markers are expressed in human gliomas and NSE knockdown sensitizes glioblastoma cells to radiotherapy and temozolomide

    International Nuclear Information System (INIS)

    Yan, Tao; Skaftnesmo, Kai Ove; Leiss, Lina; Sleire, Linda; Wang, Jian; Li, Xingang; Enger, Per Øyvind

    2011-01-01

    Expression of neuronal elements has been identified in various glial tumors, and glioblastomas (GBMs) with neuronal differentiation patterns have reportedly been associated with longer survival. However, the neuronal class III β-tubulin has been linked to increasing malignancy in astrocytomas. Thus, the significance of neuronal markers in gliomas is not established. The expressions of class III β-tubulin, neurofilament protein (NFP), microtubule-associated protein 2 (MAP2) and neuron-specific enolase (NSE) were investigated in five GBM cell lines and two GBM biopsies with immunocytochemistry and Western blot. Moreover, the expression levels were quantified by real-time qPCR under different culture conditions. Following NSE siRNA treatment we used Electric cell-substrate impedance sensing (ECIS) to monitor cell growth and migration and MTS assays to study viability after irradiation and temozolomide treatment. Finally, we quantitated NSE expression in a series of human glioma biopsies with immunohistochemistry using a morphometry software, and collected survival data for the corresponding patients. The biopsies were then grouped according to expression in two halves which were compared by survival analysis. Immunocytochemistry and Western blotting showed that all markers except NFP were expressed both in GBM cell lines and biopsies. Notably, qPCR demonstrated that NSE was upregulated in cellular stress conditions, such as serum-starvation and hypoxia, while we found no uniform pattern for the other markers. NSE knockdown reduced the migration of glioma cells, sensitized them to hypoxia, radio- and chemotherapy. Furthermore, we found that GBM patients in the group with the highest NSE expression lived significantly shorter than patients in the low-expression group. Neuronal markers are aberrantly expressed in human GBMs, and NSE is consistently upregulated in different cellular stress conditions. Knockdown of NSE reduces the migration of GBM cells and sensitizes

  12. Influence of promoter/enhancer region haplotypes on MGMT transcriptional regulation: a potential biomarker for human sensitivity to alkylating agents.

    Science.gov (United States)

    Xu, Meixiang; Nekhayeva, Ilona; Cross, Courtney E; Rondelli, Catherine M; Wickliffe, Jeffrey K; Abdel-Rahman, Sherif Z

    2014-03-01

    The O6-methylguanine-DNA methyltransferase gene (MGMT) encodes the direct reversal DNA repair protein that removes alkyl adducts from the O6 position of guanine. Several single-nucleotide polymorphisms (SNPs) exist in the MGMT promoter/enhancer (P/E) region. However, the haplotype structure encompassing these SNPs and their functional/biological significance are currently unknown. We hypothesized that MGMT P/E haplotypes, rather than individual SNPs, alter MGMT transcription and can thus alter human sensitivity to alkylating agents. To identify the haplotype structure encompassing the MGMT P/E region SNPs, we sequenced 104 DNA samples from healthy individuals and inferred the haplotypes using the data generated. We identified eight SNPs in this region, namely T7C (rs180989103), T135G (rs1711646), G290A (rs61859810), C485A (rs1625649), C575A (rs113813075), G666A (rs34180180), C777A (rs34138162) and C1099T (rs16906252). Phylogenetics and Sequence Evolution analysis predicted 21 potential haplotypes that encompass these SNPs ranging in frequencies from 0.000048 to 0.39. Of these, 10 were identified in our study population as 20 paired haplotype combinations. To determine the functional significance of these haplotypes, luciferase reporter constructs representing these haplotypes were transfected into glioblastoma cells and their effect on MGMT promoter activity was determined. Compared with the most common (reference) haplotype 1, seven haplotypes significantly upregulated MGMT promoter activity (18-119% increase; P alkylating agents.

  13. Correlation of collagen synthesis with polarization-sensitive optical coherence tomography imaging of in vitro human atherosclerosis

    Science.gov (United States)

    Kuo, Wen-Chuan; Shyu, Jeou-Jong; Chou, Nai-Kuan; Lai, Chih-Ming; Tien, En-Kuang; Huang, Huan-Jang; Chou, Chien; Jan, Gwo-Jen

    2005-04-01

    Atherosclerosis is unquestionably the leading cause of morbidity and mortality in developed countries. In the mean time, the worldwide importance of acute vascular syndromes is increasing. Because collagen fiber is a critical component of atherosclerotic lesions; it constitutes up to 60% of the total atherosclerotic plaque protein. The uncontrolled collagen accumulation leads to arterial stenosis, whereas excessive collagen breakdown weakens plaques thereby making them prone to rupture finally. Thus, in this study, we present the first application, to our knowledge, of using polarization-sensitive optical coherence tomography (PS-OCT) in human atherosclerosis. We demonstrate this technique for imaging of intensity, birefringence, and fast-axis orientation simultaneously in atherosclerotic plaques. This in vitro study suggests that the birefringence change in plaque is due to the prominent deposition of collagen according to the correlation of PS-OCT images with histological counterpart. Moreover, we can acquire quantitative criteria based on the change of polarization of incident beam to estimate whether the collagen synthesized is "too much" or "not enough". Thus by combining of high resolution intensity imaging and birefringence detection makes PS-OCT could be a potentially powerful tool for early assessment of atherosclerosis appearance and the prediction of plaque rupture in clinic.

  14. miR-320 enhances the sensitivity of human colon cancer cells to chemoradiotherapy in vitro by targeting FOXM1

    Energy Technology Data Exchange (ETDEWEB)

    Wan, Lu-Ying; Deng, Jun; Xiang, Xiao-Jun; Zhang, Ling; Yu, Feng; Chen, Jun; Sun, Zhe; Feng, Miao; Xiong, Jian-Ping, E-mail: jpxiong@medmail.com.cn

    2015-02-06

    Highlights: • miR-320 plays a significant role in chemoresistance. • This role might be attribute to targeting FOXM1. • The Wnt/β-catenin pathway also involves in this chemotherapy sensitivity. - Abstract: miR-320 expression level is found to be down-regulated in human colon cancer. To date, however, its underlying mechanisms in the chemo-resistance remain largely unknown. In this study, we demonstrated that ectopic expression of miR-320 led to inhibit HCT-116 cell proliferation, invasion and hypersensitivity to 5-Fu and Oxaliplatin. Also, knockdown of miR-320 reversed these effects in HT-29 cells. Furthermore, we identified an oncogene, FOXM1, as a direct target of miR-320. In addition, miR-320 could inactive the activity of Wnt/β-catenin pathway. Finally, we found that miR-320 and FOXM1 protein had a negative correlation in colon cancer tissues and adjacent normal tissues. These findings implied that miR-320–FOXM1 axis may overcome chemo-resistance of colon cancer cells and provide a new therapeutic target for the treatment of colon cancer.

  15. Sensitive spectrofluorimetric determination of tizanidine in pharmaceutical preparations, human plasma and urine through derivatization with dansyl chloride.

    Science.gov (United States)

    Ulu, Sevgi Tatar

    2012-01-01

    A sensitive spectrofluorimetric method was developed for the determination of tizanidine in human plasma, urine and pharmaceutical preparations. The method is based on reaction of tizanidine with 1-dimethylaminonaphthalene-5-sulphonyl chloride (dansyl chloride) in an alkaline medium to form a highly fluorescent derivative that was measured at 511 nm after excitation at 383 nm. The different experimental parameters affecting the fluorescence intensity of tizanidine was carefully studied and optimized. The fluorescence-concentration plots were rectilinear over the ranges 50-500 and 20-300 ng/mL for plasma and urine, respectively, detection limits of 1.81 and 0.54 ng/mL and quantification limits of 5.43 and 1.62 ng/mL for plasma and urine, respectively. The method presents good performance in terms of linearity, detection and quantification limits, precision, accuracy and specificity. The proposed method was successfully applied for the determination of tizanidine in pharmaceutical preparations. The results obtained were compared with a reference method, using t- and F-tests. Copyright © 2011 John Wiley & Sons, Ltd.

  16. Synthesis of high generation thermo-sensitive dendrimers for extraction of rivaroxaban from human fluid and pharmaceutic samples.

    Science.gov (United States)

    Parham, Negin; Panahi, Homayon Ahmad; Feizbakhsh, Alireza; Moniri, Elham

    2018-04-13

    In this present study, poly (N-isopropylacrylamide) as a thermo-sensitive agent was grafted onto magnetic nanoparticles, then ethylenediamine and methylmethacrylate were used to synthesize the first generation of poly amidoamine (PAMAM) dendrimers successively and the process continued alternatively until the ten generations of dendrimers. The synthesized nanocomposite was investigated using Fourier transform infrared spectrometry, thermalgravimetry analysis, X-ray diffractometry, elemental analysis and vibrating-sample magnetometer. The particle size and morphology were characterized using dynamic light scattering, field emission scanning electron microscopy and transmission electron microscopy. Batch experiments were conducted to investigate the parameters affecting adsorption and desorption of rivaroxaban by synthesized nanocomposite. The maximum sorption of rivaroxaban by the synthesized nanocomposite was obtained at pH of 8. The resulting grafted magnetic nanoparticle dendrimers were applied for extraction of rivaroxaban from biologic human liquids and medicinal samples. The specifications of rivaroxaban sorbed by a magnetic nanoparticle dendrimer showed good accessibility and high capacity of the active sites within the dendrimers. Urine and drug matrix extraction recoveries of more than 92.5 and 99.8 were obtained, respectively. Copyright © 2018 Elsevier B.V. All rights reserved.

  17. Determination of ABO blood grouping from human oral squamous epithelium by the highly sensitive immunohistochemical staining method EnVision+.

    Science.gov (United States)

    Noda, Hiroshi; Yokota, Makoto; Tatsumi, Shinji; Sugiyama, Shizuyuki

    2002-03-01

    Using the highly sensitive immunohistochemical staining method EnVision+, which employs a dextran polymer reagent for the secondary antibody, the detection of the ABH antigens was attempted in the oral squamous epithelium. This new technique uses monoclonal antibody as a primary antibody and it takes about three hours for staining. The time is much shorter than conventional absorption-elution testing or absorption-inhibition testing for the determination of ABO blood grouping. Secretor saliva samples were stained at strong intensity by the antibody, which corresponded to its blood group and anti-H. On the one hand, nonsecretor saliva samples were stained at strong intensity only by the antibody that corresponded to its blood group, and at weak intensity only by anti-H. Since human oral squamous epithelium antigens were stained specifically by this method, we can examine the ABO blood group of saliva samples and perform cytodiagnosis at the same time. Our research suggested that the EnVision+ Method is a useful technique for ABO blood grouping of saliva in forensic cases.

  18. Effect of antisense c-raf-1 on tumorigenicity and radiation sensitivity of a human squamous carcinoma

    International Nuclear Information System (INIS)

    Kasid, U.; Pfeifer, A.; Brennan, T.; Beckett, M.; Weichselbaum, R.R.; Dritschilo, A.; Mark, G.E.

    1989-01-01

    Antisense RNA-mediated inhibition of gene expression was used to investigate the biological function of the c-raf-1 gene in a radiation-resistant human squamous carcinoma cell line, SQ-20B. S1 nuclease protection assays revealed that transfection of full-length raf complementary DNA in the antisense orientation (AS) leads to a specific reduction (greater than tenfold) of steady-state levels of the endogenous c-raf-1 sense (S) transcript in SQ-20B cells. In nude mice, the malignant potential of SQ-20B cells transfected with raf (S) was significantly increased relative to that of SQ-20B cells transfected with raf (AS). SQ-20B cells containing transfected raf (S) maintained a radiation-resistant phenotype as compared to those cells harboring the AS version, which appeared to have enhanced radiation sensitivity. These data indicate that the reduced expression of endogenous c-raf-1 is sufficient to modulate the tumorigenicity and the radiation-resistant phenotype of SQ-20B cells, thus implicating c-raf-1 in a pathway important to the genesis of this type of cancer

  19. A sensitive radioimmunoassay of atrial natriuretic peptide in human plasma, using a tracer with an immobilized glycouril agent

    International Nuclear Information System (INIS)

    Rosmalen, F.M.A.; Tan, A.C.I.T.L.; Benraad, T.J.

    1987-01-01

    A highly specific and sensitive radioimmunoassay (RIA) for alpha-human atrial natriuretic peptide (hANP[1-28]) in plasma was developed. The assay used a [ 125 I]monoiodotyrosyl-hANP[1-28] tracer, prepared with an immobilized glycouril agent (Protag) and purified by high pressure liquid chromatography (HPLC), and a highly specific antiserum raised against hANP[1-28], coupled to keyhole limpet haemocyanin, in sheep. Plasma was extracted using C-18 Seppak cartridges. A good parallelism was found after dilution prior to extraction of plasma of patients with congestive heart failure (CHF) or of plasma of healthy subjects. Recovery of hANP[1-28] added to plasma was 96%. The limit of detection was 0.8 pg/tube, intra- and inter-assay variation were 9 and 12%, respectively. Mean plasma ANP values in 25 normal persons with a normal salt intake was 26.0 ± 15.5 (± SD) pg/ml. Plasma levels of 18 subjects (7 normals, 11 CHF) were measured using four different antisera after the extraction step. High correlations were found between the values obtained with these four antisera. (Auth.)

  20. An ultra-sensitive monoclonal antibody-based enzyme-linked immunosobent assay for dibutyl phthalate in human urinary

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Lifang [Institute of Pharmacology, Toxicology and Biochemical Pharmaceutics, College of Pharmaceutical Science, Zhejiang University, Hangzhou 310058 (China); Lei, Yajing [Hangzhou EPIE Bio-detection Technology Limited, Hangzhou 310051 (China); Zhang, Dai; Ahmed, Shabbir [Institute of Pharmacology, Toxicology and Biochemical Pharmaceutics, College of Pharmaceutical Science, Zhejiang University, Hangzhou 310058 (China); Chen, Shuqing, E-mail: chenshuqing@zju.edu.cn [Institute of Pharmacology, Toxicology and Biochemical Pharmaceutics, College of Pharmaceutical Science, Zhejiang University, Hangzhou 310058 (China)

    2016-01-15

    Dibutyl phthalate (DBP) has been extensively used as a plasticizer in many daily products, which is highly toxic to human, notably affecting the reproductive and developmental function. As the previous method is expensive, time-consuming, low sensitivity and just focused on the environment. Present study was aimed to establish an ultra-sensitive and simple method based on good quality monoclonal antibody, applying to evaluate excretion level of DBP in urine samples of Chinese population directly. A monoclonal antibody was generated and characterized after fusion of myeloma cells with spleen cells isolated from BALB/c mouse. The mouse was previously immunized using a specially designed amino derivative of DBP conjugated with bovine serum albumin (BSA) as immunogen. Cross-reactivity values of the monoclonal antibody against DBP, di-isobutyl phthalate (DIBP) were observed 100% and 1.25%, while for dimethyl phthalate (DMP), butyl benzyl phthalate (BBP) and didecyl phthalate (DDP) the values were < 0.06%. The standard curve was constructed at 0–50 ng mL{sup −1} and good linearity (R{sup 2} = 0.994) was achieved. The observed IC{sub 50} (7.34 ng mL{sup −1}) and LOD (0.06 ng mL{sup −1}) values was improved 1000-fold to polyclonal antibody and 5-fold to other monoclonal antibodies. A total 1246 urine samples were analyzed and the detection frequency of DBP was observed 72.87% by ic-ELISA. The 95th percentile and mean concentration of DBP were 12.07 and 3.00 ng mL{sup −1}. Acceptable recovery rates of DBP were 97.8–114.3% and coefficients variation 5.93–11.09%. The concentrations of DBP in females were found significantly higher (p < 0.05) than males. Similarly, the DBP in middle aged and low educated individuals was found higher (p < 0.001) than the others. Considering the adverse health effects, DBP internal exposure in the Chinese population should be reduced. The ic-ELISA method has been proved as a cost effective, specific, and highly sensitive screening

  1. Sulfite-induced protein radical formation in LPS aerosol-challenged mice: Implications for sulfite sensitivity in human lung disease

    Directory of Open Access Journals (Sweden)

    Ashutosh Kumar

    2018-05-01

    Full Text Available Exposure to (bisulfite (HSO3– and sulfite (SO32– has been shown to induce a wide range of adverse reactions in sensitive individuals. Studies have shown that peroxidase-catalyzed oxidation of (bisulfite leads to formation of several reactive free radicals, such as sulfur trioxide anion (.SO3–, peroxymonosulfate (–O3SOO., and especially the sulfate (SO4. – anion radicals. One such peroxidase in neutrophils is myeloperoxidase (MPO, which has been shown to form protein radicals. Although formation of (bisulfite-derived protein radicals is documented in isolated neutrophils, its involvement and role in in vivo inflammatory processes, has not been demonstrated. Therefore, we aimed to investigate (bisulfite-derived protein radical formation and its mechanism in LPS aerosol-challenged mice, a model of non-atopic asthma. Using immuno-spin trapping to detect protein radical formation, we show that, in the presence of (bisulfite, neutrophils present in bronchoalveolar lavage and in the lung parenchyma exhibit, MPO-catalyzed oxidation of MPO to a protein radical. The absence of radical formation in LPS-challenged MPO- or NADPH oxidase-knockout mice indicates that sulfite-derived radical formation is dependent on both MPO and NADPH oxidase activity. In addition to its oxidation by the MPO-catalyzed pathway, (bisulfite is efficiently detoxified to sulfate by the sulfite oxidase (SOX pathway, which forms sulfate in a two-electron oxidation reaction. Since SOX activity in rodents is much higher than in humans, to better model sulfite toxicity in humans, we induced SOX deficiency in mice by feeding them a low molybdenum diet with tungstate. We found that mice treated with the SOX deficiency diet prior to exposure to (bisulfite had much higher protein radical formation than mice with normal SOX activity. Altogether, these results demonstrate the role of MPO and NADPH oxidase in (bisulfite-derived protein radical formation and show the involvement of

  2. Comparative DNA microarray analysis of human monocyte derived dendritic cells and MUTZ-3 cells exposed to the moderate skin sensitizer cinnamaldehyde

    International Nuclear Information System (INIS)

    Python, Francois; Goebel, Carsten; Aeby, Pierre

    2009-01-01

    The number of studies involved in the development of in vitro skin sensitization tests has increased since the adoption of the EU 7th amendment to the cosmetics directive proposing to ban animal testing for cosmetic ingredients by 2013. Several studies have recently demonstrated that sensitizers induce a relevant up-regulation of activation markers such as CD86, CD54, IL-8 or IL-1β in human myeloid cell lines (e.g., U937, MUTZ-3, THP-1) or in human peripheral blood monocyte-derived dendritic cells (PBMDCs). The present study aimed at the identification of new dendritic cell activation markers in order to further improve the in vitro evaluation of the sensitizing potential of chemicals. We have compared the gene expression profiles of PBMDCs and the human cell line MUTZ-3 after a 24-h exposure to the moderate sensitizer cinnamaldehyde. A list of 80 genes modulated in both cell types was obtained and a set of candidate marker genes was selected for further analysis. Cells were exposed to selected sensitizers and non-sensitizers for 24 h and gene expression was analyzed by quantitative real-time reverse transcriptase-polymerase chain reaction. Results indicated that PIR, TRIM16 and two Nrf2-regulated genes, CES1 and NQO1, are modulated by most sensitizers. Up-regulation of these genes could also be observed in our recently published DC-activation test with U937 cells. Due to their role in DC activation, these new genes may help to further refine the in vitro approaches for the screening of the sensitizing properties of a chemical.

  3. An in vitro method for detecting chemical sensitization using human reconstructed skin models and its applicability to cosmetic, pharmaceutical, and medical device safety testing.

    Science.gov (United States)

    McKim, James M; Keller, Donald J; Gorski, Joel R

    2012-12-01

    Chemical sensitization is a serious condition caused by small reactive molecules and is characterized by a delayed type hypersensitivity known as allergic contact dermatitis (ACD). Contact with these molecules via dermal exposure represent a significant concern for chemical manufacturers. Recent legislation in the EU has created the need to develop non-animal alternative methods for many routine safety studies including sensitization. Although most of the alternative research has focused on pure chemicals that possess reasonable solubility properties, it is important for any successful in vitro method to have the ability to test compounds with low aqueous solubility. This is especially true for the medical device industry where device extracts must be prepared in both polar and non-polar vehicles in order to evaluate chemical sensitization. The aim of this research was to demonstrate the functionality and applicability of the human reconstituted skin models (MatTek Epiderm(®) and SkinEthic RHE) as a test system for the evaluation of chemical sensitization and its potential use for medical device testing. In addition, the development of the human 3D skin model should allow the in vitro sensitization assay to be used for finished product testing in the personal care, cosmetics, and pharmaceutical industries. This approach combines solubility, chemical reactivity, cytotoxicity, and activation of the Nrf2/ARE expression pathway to identify and categorize chemical sensitizers. Known chemical sensitizers representing extreme/strong-, moderate-, weak-, and non-sensitizing potency categories were first evaluated in the skin models at six exposure concentrations ranging from 0.1 to 2500 µM for 24 h. The expression of eight Nrf2/ARE, one AhR/XRE and two Nrf1/MRE controlled gene were measured by qRT-PCR. The fold-induction at each exposure concentration was combined with reactivity and cytotoxicity data to determine the sensitization potential. The results demonstrated that

  4. A sensitive and selective liquid chromatography/tandem mass spectrometry method for quantitative analysis of efavirenz in human plasma.

    Directory of Open Access Journals (Sweden)

    Praveen Srivastava

    Full Text Available A selective and a highly sensitive method for the determination of the non-nucleoside reverse transcriptase inhibitor (NNRTI, efavirenz, in human plasma has been developed and fully validated based on high performance liquid chromatography tandem mass spectrometry (LC-MS/MS. Sample preparation involved protein precipitation followed by one to one dilution with water. The analyte, efavirenz was separated by high performance liquid chromatography and detected with tandem mass spectrometry in negative ionization mode with multiple reaction monitoring. Efavirenz and ¹³C₆-efavirenz (Internal Standard, respectively, were detected via the following MRM transitions: m/z 314.20243.90 and m/z 320.20249.90. A gradient program was used to elute the analytes using 0.1% formic acid in water and 0.1% formic acid in acetonitrile as mobile phase solvents, at a flow-rate of 0.3 mL/min. The total run time was 5 min and the retention times for the internal standard (¹³C₆-efavirenz and efavirenz was approximately 2.6 min. The calibration curves showed linearity (coefficient of regression, r>0.99 over the concentration range of 1.0-2,500 ng/mL. The intraday precision based on the standard deviation of replicates of lower limit of quantification (LLOQ was 9.24% and for quality control (QC samples ranged from 2.41% to 6.42% and with accuracy from 112% and 100-111% for LLOQ and QC samples. The inter day precision was 12.3% and 3.03-9.18% for LLOQ and quality controls samples, and the accuracy was 108% and 95.2-108% for LLOQ and QC samples. Stability studies showed that efavirenz was stable during the expected conditions for sample preparation and storage. The lower limit of quantification for efavirenz was 1 ng/mL. The analytical method showed excellent sensitivity, precision, and accuracy. This method is robust and is being successfully applied for therapeutic drug monitoring and pharmacokinetic studies in HIV-infected patients.

  5. Adherence performances of pressure sensitive adhesives on a model viscoelastic synthetic film: a tool for the understanding of adhesion on the human skin.

    Science.gov (United States)

    Renvoise, Julien; Burlot, Delphine; Marin, Gérard; Derail, Christophe

    2009-02-23

    This work deals with the rheological behavior and adherence properties of pressure sensitive adhesive formulations dedicated to medical applications. We have developed a specific viscoelastic substrate which mimics adhesion on human skin to measure the adherence properties of PSAs when they are stuck on the human skin. By comparing peeling results of PSAs, dedicated to medical applications, stuck on human skin and on this viscoelastic substrate we show that this substrate, based on a blend of natural proteins, presents a better representation of the interactions occurring at the skin/adhesive interface than conventional substrates used for peel test (i.e. glass and steel).

  6. Prediction of the contact sensitizing potential of chemicals using analysis of gene expression changes in human THP-1 monocytes.

    Science.gov (United States)

    Arkusz, Joanna; Stępnik, Maciej; Sobala, Wojciech; Dastych, Jarosław

    2010-11-10

    The aim of this study was to find differentially regulated genes in THP-1 monocytic cells exposed to sensitizers and nonsensitizers and to investigate if such genes could be reliable markers for an in vitro predictive method for the identification of skin sensitizing chemicals. Changes in expression of 35 genes in the THP-1 cell line following treatment with chemicals of different sensitizing potential (from nonsensitizers to extreme sensitizers) were assessed using real-time PCR. Verification of 13 candidate genes by testing a large number of chemicals (an additional 22 sensitizers and 8 nonsensitizers) revealed that prediction of contact sensitization potential was possible based on evaluation of changes in three genes: IL8, HMOX1 and PAIMP1. In total, changes in expression of these genes allowed correct detection of sensitization potential of 21 out of 27 (78%) test sensitizers. The gene expression levels inside potency groups varied and did not allow estimation of sensitization potency of test chemicals. Results of this study indicate that evaluation of changes in expression of proposed biomarkers in THP-1 cells could be a valuable model for preliminary screening of chemicals to discriminate an appreciable majority of sensitizers from nonsensitizers. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  7. HPW-RX40 restores anoikis sensitivity of human breast cancer cells by inhibiting integrin/FAK signaling

    Energy Technology Data Exchange (ETDEWEB)

    Chen, I-Hua; Shih, Hsin-Chu [Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Hsieh, Pei-Wen [Graduate Institute of Natural Products, School of Traditional Chinese Medicine, and Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan (China); Chang, Fang-Rong [Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Cancer Center, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan (China); Wu, Yang-Chang, E-mail: yachwu@mail.cmu.edu [School of Pharmacy, College of Pharmacy, China Medical University, Taichung, Taiwan (China); Wu, Chin-Chung, E-mail: ccwu@kmu.edu.tw [Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Department of Marine Biotechnology and Resources, National Sun Yat-sen University, Kaohsiung 80708, Taiwan (China); Research Center for Natural Products and Drug Development, Kaohsiung Medical University, Kaohsiung, Taiwan (China)

    2015-12-01

    Anoikis is defined as apoptosis, which is induced by inappropriate cell–matrix interactions. Cancer cells with anoikis resistance tend to undergo metastasis, and this phenomenon has been reported to be associated with integrin and FAK activity. HPW-RX40 is a derivative of 3,4-methylenedioxy-β-nitrostyrene, which is known to prevent platelet aggregation by inhibition of integrin. In the present study, we investigated the effect of HPW-RX40 on an anoikis-resistant human breast cancer cell line MDA-MB-231. HPW-RX40 inhibited cell aggregation and induced cell death in suspending MDA-MB-231 cells, but had only little effect on the monolayer growth of adherent cells. Analysis of caspase activation and poly (ADP-ribose) polymerase (PARP) cleavage confirmed anoikis in HPW-RX40-treated suspending cancer cells. HPW-RX40 also affected the Bcl-2 family proteins in detached cancer cells. Furthermore, HPW-RX40 inhibited detachment-induced activation of FAK and the downstream phosphorylation of Src and paxillin, but did not affect this pathway in adherent cancer cells. We also found that the expression and activation of β1 integrin in MDA-MB-231 cells were reduced by HPW-RX40. The combination of HPW-RX40 with an EGFR inhibitor led to enhanced anoikis and inhibition of the FAK pathway in breast cancer cells. Taken together, our results suggest that HPW-RX40 restores the anoikis sensitivity in the metastatic breast cancer cells by inhibiting integrin and subsequent FAK activation, and reveal a potential strategy for prevention of tumor metastasis. - Highlights: • The β-nitrostyrene derivative, HPW-RX40, induces anoikis in human breast cancer cells. • HPW-RX40 inhibits the integrin/FAK signaling pathway. • The combination of HPW-RX40 with an EGFR inhibitor leads to enhanced anoikis. • HPW-RX40 may have a potential to prevent the spread of metastatic breast cancer.

  8. HPW-RX40 restores anoikis sensitivity of human breast cancer cells by inhibiting integrin/FAK signaling

    International Nuclear Information System (INIS)

    Chen, I-Hua; Shih, Hsin-Chu; Hsieh, Pei-Wen; Chang, Fang-Rong; Wu, Yang-Chang; Wu, Chin-Chung

    2015-01-01

    Anoikis is defined as apoptosis, which is induced by inappropriate cell–matrix interactions. Cancer cells with anoikis resistance tend to undergo metastasis, and this phenomenon has been reported to be associated with integrin and FAK activity. HPW-RX40 is a derivative of 3,4-methylenedioxy-β-nitrostyrene, which is known to prevent platelet aggregation by inhibition of integrin. In the present study, we investigated the effect of HPW-RX40 on an anoikis-resistant human breast cancer cell line MDA-MB-231. HPW-RX40 inhibited cell aggregation and induced cell death in suspending MDA-MB-231 cells, but had only little effect on the monolayer growth of adherent cells. Analysis of caspase activation and poly (ADP-ribose) polymerase (PARP) cleavage confirmed anoikis in HPW-RX40-treated suspending cancer cells. HPW-RX40 also affected the Bcl-2 family proteins in detached cancer cells. Furthermore, HPW-RX40 inhibited detachment-induced activation of FAK and the downstream phosphorylation of Src and paxillin, but did not affect this pathway in adherent cancer cells. We also found that the expression and activation of β1 integrin in MDA-MB-231 cells were reduced by HPW-RX40. The combination of HPW-RX40 with an EGFR inhibitor led to enhanced anoikis and inhibition of the FAK pathway in breast cancer cells. Taken together, our results suggest that HPW-RX40 restores the anoikis sensitivity in the metastatic breast cancer cells by inhibiting integrin and subsequent FAK activation, and reveal a potential strategy for prevention of tumor metastasis. - Highlights: • The β-nitrostyrene derivative, HPW-RX40, induces anoikis in human breast cancer cells. • HPW-RX40 inhibits the integrin/FAK signaling pathway. • The combination of HPW-RX40 with an EGFR inhibitor leads to enhanced anoikis. • HPW-RX40 may have a potential to prevent the spread of metastatic breast cancer.

  9. Simple and sensitive high performance liquid chromatographic method for the simultaneous quantitation of the phenylalanine in human plasma

    Directory of Open Access Journals (Sweden)

    Hossein Danafar

    2015-09-01

    Full Text Available Phenylalanine (Phe is the most reliable indicator for the diagnosis of phenylketonuria (PKU. The purpose of this study is to establish a reliable and quick method for the assignment of Phe in peripheral capillary blood from newborns and children by high performance liquid chromatography with ultraviolet detection (HPLC-UV. PKU is an inborn error of metabolism characterized by the inability of the body to use Phe. A rapid and sensitive high performance liquid chromatographic (HPLC method has been developed for determination of Phe in plasma. The method uses a protein precipitation step with sulfosalicilic acid for sample preparation by separation on a Nova-pack C18 column using sodium acetate buffer and acetonitrile (94: 6 v/v adjusted to pH 6.5 with glacial acetic acid. The eluted peaks detected by a UV detector was set at wavelength of 215 nm. The method was validated in the range of Phe concentrations from 0.1 to 20 µg/ml. The limits of detection (LOD and quantitation (LOQ of the method were 0.05 and 0.1 µg/ml, respectively. The average drug recovery from plasma was 88.60 percent throughout the linear concentration range., with the average within-run and between-run accuracy values of 103.3 and 115.350, respectively. The method is quick, easy, very steady and precise for the screen, assignment, and evaluation of Phe in human plasma by HPLC, which is particularly a useful way for screening and diagnosis of PKU and monitoring of a diet therapy.

  10. Trichostatin A (TSA) sensitizes the human prostatic cancer cell line DU145 to death receptor ligands treatment.

    Science.gov (United States)

    Taghiyev, Agshin F; Guseva, Natalya V; Sturm, Mary T; Rokhlin, Oskar W; Cohen, Michael B

    2005-04-01

    The human prostatic carcinoma cell line DU145 has previously been found to be resistant to treatment with TNF-family ligands. However, TRAIL, TNF-alpha and anti-Fas antibodies (Ab) treatment in combination with the histone deacetylase inhibitor Trichostatin A (TSA) converted the phenotype of DU145 from resistant to sensitive. TSA induced 15% cell death but simultaneous treatment with TRAIL, TNF-alpha and anti-Fas Ab resulted in 55%, 70% and 40% cell death, respectively. Simultaneous treatment did not increase the level of TSA-induced histone acetylation, but induced the release of acetylated histones from chromatin into the cytosol. This release was caspase dependent since it was abrogated by Z-VAD-fmk. In addition, treatment with TSA induced caspase-9 activation and resulted in the release of cytochrome c and Smac/DIABLO from mitochondria. To further investigate the role of caspase-9 in TSA-mediated apoptosis we used two different approaches: (1) cells were pretreated with the caspase-9 inhibitor Z-LEHD-fmk, and (2) cells were transfected with a dominant-negative form of caspase-9. Both approaches gave similar results: cells became resistant to treatment with TSA. These data indicate that TSA mediates its effect via the mitochondrial pathway. This was confirmed by examining DU145 overexpressing Bcl-2. These transfectants were resistant to TSA treatment. Taken together, our data shows that only simultaneous treatment with TNF-family ligands and TSA in DU145 resulted in caspase activity sufficient to induce apoptosis. The combination of TSA and TNF-family ligands could potentially be the basis for the treatment of prostate cancer.

  11. Sensitization of recombinant human tumor necrosis factor-related apoptosis-inducing ligand-resistant malignant melanomas by quercetin.

    Science.gov (United States)

    Turner, Katherine A; Manouchehri, Jasmine M; Kalafatis, Michael

    2018-03-28

    Malignant melanoma is the most commonly diagnosed skin cancer associated with a high rate of metastasis. Low-stage melanoma is easily treated, but metastatic malignant melanoma is an extremely treatment-resistant malignancy with low survival rates. The application of recombinant human tumor necrosis factor-related apoptosis-inducing ligand (rhTRAIL) for the treatment of metastatic malignant melanoma holds considerable promise because of its selective proapoptotic activity towards cancer cells and not nontransformed cells. Unfortunately, the clinical utilization of rhTRAIL has been terminated due to the resistance of many cancer cells to undergo apoptosis in response to rhTRAIL. However, rhTRAIL-resistance can be abrogated through the cotreatment with compounds derived from 'Mother Nature' such as quercetin that can modulate cellular components responsible for rhTRAIL-resistance. Here, we show that rhTRAIL-resistant malignant melanomas are sensitized by quercetin. Quercetin action is manifested by the upregulation of rhTRAIL-binding receptors DR4 and DR5 on the surface of cancer cells and by increased rate of the proteasome-mediated degradation of the antiapoptotic protein FLIP. Our data provide for a new efficient and nontoxic treatment of malignant melanoma.This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives License 4.0 (CCBY-NC-ND), where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal. http://creativecommons.org/licenses/by-nc-nd/4.0/.

  12. Comparative sensitivity to environmental variation and human disturbance of Asian tapirs (Tapirus indicus) and other wild ungulates in Thailand.

    Science.gov (United States)

    Lynam, Antony J; Tantipisanuh, Naruemon; Chutipong, Wanlop; Ngoprasert, Dusit; Baker, Megan C; Cutter, Passanan; Gale, George; Kitamura, Shumpei; Steinmetz, Robert; Sukmasuang, Ronglarp; Thunhikorn, Somying

    2012-12-01

    Southeast Asia's tropical forests suffer the highest rates of deforestation and disturbance of any on Earth, with poorly understood impacts on native fauna. Asian tapirs (Tapirus indicus) are among the least studied of the large mammals in these forests. Using records from 9 camera trap surveys in 7 of the largest (>1000 km(2) ) protected area complexes, we assessed the influence of environmental variation and human-induced disturbance on tapir occurrence. Tapirs were detected at 13% of locations sampled, significantly associated with evergreen forest (P tapir presence 87% of the time. According to this model, tapir occurrence was positively influenced by annual rainfall and proximity to the forest edge. However, tapirs may not avoid edges but instead prefer wetter evergreen forest, a habitat type that tended to occur further from the forest edge at higher elevations in our particular study sites (P tapirs showed a range of differing responses. Tapirs are expected to be less sensitive to disturbance because they are not targets for hunting and trade, and are almost entirely active at night, so avoid peak traffic periods in parks. Tapir populations in Thailand may be more stable than in other parts of their global range because rates of forest loss have decreased >40% over the past 20 years. We recommend surveys to fill gaps in the understanding of the status in lesser-known protected areas, research to better understand the fine-scale environmental influences on behavior and habitats of tapirs, and other forest ungulates, and continued legal status for tapirs in the highest category of protection. © 2012 Wiley Publishing Asia Pty Ltd, ISZS and IOZ/CAS.

  13. Semi-Metric Topology of the Human Connectome: Sensitivity and Specificity to Autism and Major Depressive Disorder.

    Directory of Open Access Journals (Sweden)

    Tiago Simas

    Full Text Available The human functional connectome is a graphical representation, consisting of nodes connected by edges, of the inter-relationships of blood oxygenation-level dependent (BOLD time-series measured by MRI from regions encompassing the cerebral cortices and, often, the cerebellum. Semi-metric analysis of the weighted, undirected connectome distinguishes an edge as either direct (metric, such that there is no alternative path that is accumulatively stronger, or indirect (semi-metric, where one or more alternative paths exist that have greater strength than the direct edge. The sensitivity and specificity of this method of analysis is illustrated by two case-control analyses with independent, matched groups of adolescents with autism spectrum conditions (ASC and major depressive disorder (MDD.Significance differences in the global percentage of semi-metric edges was observed in both groups, with increases in ASC and decreases in MDD relative to controls. Furthermore, MDD was associated with regional differences in left frontal and temporal lobes, the right limbic system and cerebellum. In contrast, ASC had a broadly increased percentage of semi-metric edges with a more generalised distribution of effects and some areas of reduction. In summary, MDD was characterised by localised, large reductions in the percentage of semi-metric edges, whilst ASC is characterised by more generalised, subtle increases. These differences were corroborated in greater detail by inspection of the semi-metric backbone for each group; that is, the sub-graph of semi-metric edges present in >90% of participants, and by nodal degree differences in the semi-metric connectome.These encouraging results, in what we believe is the first application of semi-metric analysis to neuroimaging data, raise confidence in the methodology as potentially capable of detection and characterisation of a range of neurodevelopmental and psychiatric disorders.

  14. The Role of the Co-Chaperone, CHIP, in Androgen Independent Prostate Cancer

    Science.gov (United States)

    2008-02-01

    C4-2 there was no evidence of lysosomal congregration, the hallmark of autophagy, rather vaculolization of cytoplasm and disintergration of cellular...RNA polymerase promoter at the 5′ end. Second-strand synthesis was followed by cRNA production incorporating a biotinylated base. Hybridization to

  15. Role of Androgen Receptor in Growth of Androgen Independent Prostate Cancer

    National Research Council Canada - National Science Library

    Chen, Charlie

    2003-01-01

    ...) overexpression is the only consistent change in the progression of prostate cancer. In the last grand period, I confirmed by western blot analysis that androgen receptor protein is higher in HR than HS tumors...

  16. Suppression of gastric acid increases the risk of developing immunoglobulin E-mediated drug hypersensitivity: human diclofenac sensitization and a murine sensitization model.

    Science.gov (United States)

    Riemer, A B; Gruber, S; Pali-Schöll, I; Kinaciyan, T; Untersmayr, E; Jensen-Jarolim, E

    2010-03-01

    Hypersensitivity reactions towards non-steroidal anti-inflammatory drugs (NSAID) are common, although true allergies are detectable only in a subgroup of patients. The current study was prompted by a case observation, where a patient experienced generalized urticaria following his second course of diclofenac and proton pump inhibitor medication, and was found to have diclofenac-specific IgE. During recent years, our group has been investigating the importance of gastric digestion in the development of food allergies, demonstrating anti-acid medication as a risk factor for sensitization against food proteins. Here, we aimed to investigate whether the mechanism of food allergy induction described can also be causative in NSAID allergy, using diclofenac as a paradigm. We subjected BALB/c mice to several oral immunization regimens modelled after the patient's medication intake. Diclofenac was applied with or without gastric acid suppression, in various doses, alone or covalently coupled to albumin, a protein abundant in gastric juices. Immune responses were assessed on the antibody level, and functionally examined by in vitro and in vivo crosslinking assays. Only mice receiving albumin-coupled diclofenac under gastric acid suppression developed anti-diclofenac IgG1 and IgE, whereas no immune responses were induced by the drug alone or without gastric acid suppression. Antibody induction was dose dependent with the group receiving the higher dose of the drug showing sustained anti-diclofenac titres. The antibodies induced triggered basophil degranulation in vitro and positive skin tests in vivo. Gastric acid suppression was found to be a causative mechanism in the induction of IgE-mediated diclofenac allergy.

  17. Ionizing radiation and nitric oxide donor sensitize Fas-induced apoptosis via up-regulation of Fas in human cervical cancer cells

    International Nuclear Information System (INIS)

    Park, In Chul; Woo, Sang Hyeok; Park, Myung Jin; Lee, Hyung Chahn; Lee Su Jae; Hong, Young Joon; Lee, Seung Hoon; Hong, Seok II; Rhee, Chang Hun

    2004-01-01

    Fas/CD95/Apo1 is a transmembrane receptor known to trigger apoptotic cell death in several cell types. In the present study, we showed that ionizing radiation (IR) and NO donor, S-nitroso-N-acetyl-penicillamine (SNAP), sensitized Fas-induced apoptotic cell death of HeLa human cervical cancers. Suboptimal dose of IR and SNAP up-regulated cell-surface Fas antigen, detected by FACScan using FITC-anti-Fas antibody. When combined with IR or SNAP, agonistic anti-Fas antibody CH-11 resulted in marked enhancement of apoptosis. This sensitization was completely abrogated by anti-Fas neutralizing antibody ZB4. During the IR and SNAP sensitized Fas-induced apoptosis, mitochondria permeabilization, cytochrome c release, and DNA fragmentation were detected. Furthermore, combined treatment of IR and SNAP additively up-regulated the surface Fas protein expression and sensitized Fas-induced apoptosis. Our finding demonstrate that sensitization of HeLa cervical cells to Fas-mediated apoptosis by IR and NO donor is most likely due to the up-regulation of Fas expression and also provides a means with which to sensitize tumors to the killing effects of cancer therapy via the Fas receptor

  18. Improved physiologically based pharmacokinetic model for oral exposures to chromium in mice, rats, and humans to address temporal variation and sensitive populations.

    Science.gov (United States)

    Kirman, C R; Suh, M; Proctor, D M; Hays, S M

    2017-06-15

    A physiologically based pharmacokinetic (PBPK) model for hexavalent chromium [Cr(VI)] in mice, rats, and humans developed previously (Kirman et al., 2012, 2013), was updated to reflect an improved understanding of the toxicokinetics of the gastrointestinal tract following oral exposures. Improvements were made to: (1) the reduction model, which describes the pH-dependent reduction of Cr(VI) to Cr(III) in the gastrointestinal tract under both fasted and fed states; (2) drinking water pattern simulations, to better describe dosimetry in rodents under the conditions of the NTP cancer bioassay; and (3) parameterize the model to characterize potentially sensitive human populations. Important species differences, sources of non-linear toxicokinetics, and human variation are identified and discussed within the context of human health risk assessment. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  19. A Sensitivity Study of Human Errors in Optimizing Surveillance Test Interval (STI) and Allowed Outage Time (AOT) of Standby Safety System

    International Nuclear Information System (INIS)

    Chung, Dae Wook; Shin, Won Ky; You, Young Woo; Yang, Hui Chang

    1998-01-01

    In most cases, the surveillance test intervals (STIs), allowed outage times (AOTS) and testing strategies of safety components in nuclear power plant are prescribed in plant technical specifications. And, in general, it is required that standby safety system shall be redundant (i.e., composed of multiple components) and these components are tested by either staggered test strategy or sequential test strategy. In this study, a linear model is presented to incorporate the effects of human errors associated with test into the evaluation of unavailability. The average unavailabilities of 1/4, 2/4 redundant systems are computed considering human error and testing strategy. The adverse effects of test on system unavailability, such as component wear and test-induced transient have been modelled. The final outcome of this study would be the optimized human error domain from 3-D human error sensitivity analysis by selecting finely classified segment. The results of sensitivity analysis show that the STI and AOT can be optimized provided human error probability is maintained within allowable range. (authors)

  20. Improved physiologically based pharmacokinetic model for oral exposures to chromium in mice, rats, and humans to address temporal variation and sensitive populations

    Energy Technology Data Exchange (ETDEWEB)

    Kirman, C.R., E-mail: ckirman@summittoxicology.com [Summit Toxicology, PO Box 3209, Bozeman, MT 59715 (United States); Suh, M.; Proctor, D.M. [ToxStrategies, Mission Viejo, CA (United States); Hays, S.M. [Summit Toxicology, PO Box 3209, Bozeman, MT 59715 (United States)

    2017-06-15

    A physiologically based pharmacokinetic (PBPK) model for hexavalent chromium [Cr(VI)] in mice, rats, and humans developed previously (Kirman et al., 2012, 2013), was updated to reflect an improved understanding of the toxicokinetics of the gastrointestinal tract following oral exposures. Improvements were made to: (1) the reduction model, which describes the pH-dependent reduction of Cr(VI) to Cr(III) in the gastrointestinal tract under both fasted and fed states; (2) drinking water pattern simulations, to better describe dosimetry in rodents under the conditions of the NTP cancer bioassay; and (3) parameterize the model to characterize potentially sensitive human populations. Important species differences, sources of non-linear toxicokinetics, and human variation are identified and discussed within the context of human health risk assessment. - Highlights: • An improved version of the PBPK model for Cr(VI) toxicokinetics was developed. • The model incorporates data collected to fill important data gaps. • Model predictions for specific age groups and sensitive subpopulations are provided. • Implications to human health risk assessment are discussed.

  1. Interferon-gamma and tumor necrosis factor-alpha sensitize primarily resistant human endometrial stromal cells to Fas-mediated apoptosis

    DEFF Research Database (Denmark)

    Fluhr, Herbert; Krenzer, Stefanie; Stein, Gerburg M

    2007-01-01

    The subtle interaction between the implanting embryo and the maternal endometrium plays a pivotal role during the process of implantation. Human endometrial stromal cells (ESCs) express Fas and the implanting trophoblast cells secrete Fas ligand (FASLG, FasL), suggesting a possible role for Fas......-mediated signaling during early implantation. Here we show that ESCs are primarily resistant to Fas-mediated apoptosis independently of their state of hormonal differentiation. Pre-treatment of ESCs with interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha sensitizes them to become apoptotic upon stimulation...... of Fas by an agonistic anti-Fas antibody. Incubation of ESCs with the early embryonic signal human chorionic gonadotropin (hCG, CGB) does not influence their reaction to Fas stimulation. The sensitizing effect of IFN-gamma and TNF-alpha was accompanied by a significant upregulation of Fas and FLICE...

  2. Using detergent to enhance detection sensitivity of African trypanosomes in human CSF and blood by loop-mediated isothermal amplification (LAMP).

    Science.gov (United States)

    Grab, Dennis J; Nikolskaia, Olga V; Inoue, Noboru; Thekisoe, Oriel M M; Morrison, Liam J; Gibson, Wendy; Dumler, J Stephen

    2011-08-01

    The loop-mediated isothermal amplification (LAMP) assay, with its advantages of simplicity, rapidity and cost effectiveness, has evolved as one of the most sensitive and specific methods for the detection of a broad range of pathogenic microorganisms including African trypanosomes. While many LAMP-based assays are sufficiently sensitive to detect DNA well below the amount present in a single parasite, the detection limit of the assay is restricted by the number of parasites present in the volume of sample assayed; i.e. 1 per µL or 10(3) per mL. We hypothesized that clinical sensitivities that mimic analytical limits based on parasite DNA could be approached or even obtained by simply adding detergent to the samples prior to LAMP assay. For proof of principle we used two different LAMP assays capable of detecting 0.1 fg genomic DNA (0.001 parasite). The assay was tested on dilution series of intact bloodstream form Trypanosoma brucei rhodesiense in human cerebrospinal fluid (CSF) or blood with or without the addition of the detergent Triton X-100 and 60 min incubation at ambient temperature. With human CSF and in the absence of detergent, the LAMP detection limit for live intact parasites using 1 µL of CSF as the source of template was at best 10(3) parasites/mL. Remarkably, detergent enhanced LAMP assay reaches sensitivity about 100 to 1000-fold lower; i.e. 10 to 1 parasite/mL. Similar detergent-mediated increases in LAMP assay analytical sensitivity were also found using DNA extracted from filter paper cards containing blood pretreated with detergent before card spotting or blood samples spotted on detergent pretreated cards. This simple procedure for the enhanced detection of live African trypanosomes in biological fluids by LAMP paves the way for the adaptation of LAMP for the economical and sensitive diagnosis of other protozoan parasites and microorganisms that cause diseases that plague the developing world.

  3. Inhibition of WNT signaling reduces differentiation and induces sensitivity to doxorubicin in human malignant neuroblastoma SH-SY5Y cells.

    Science.gov (United States)

    Suebsoonthron, Junjira; Jaroonwitchawan, Thiranut; Yamabhai, Montarop; Noisa, Parinya

    2017-06-01

    Neuroblastoma is one of the most common cancers in infancy, arising from the neuroblasts during embryonic development. This cancer is difficult to treat and resistance to chemotherapy is often found; therefore, clinical trials of novel therapeutic approaches, such as targeted-cancer signaling, could be an alternative for a better treatment. WNT signaling plays significant roles in the survival, proliferation, and differentiation of human neuroblastoma. In this report, WNT signaling of a malignant human neuroblastoma cell line, SH-SY5Y cells, was inhibited by XAV939, a specific inhibitor of the Tankyrase enzyme. XAV939 treatment led to the reduction of β-catenin within the cells, confirming its inhibitory effect of WNT. The inhibition of WNT signaling by XAV939 did not affect cell morphology, survival, and proliferation; however, the differentiation and sensitivity to anticancer drugs of human neuroblastoma cells were altered. The treatment of XAV939 resulted in the downregulation of mature neuronal markers, including β-tubulin III, PHOX2A, and PHOX2B, whereas neural progenitor markers (PAX6, TFAP2α, and SLUG) were upregulated. In addition, the combination of XAV939 significantly enhanced the sensitivity of SH-SY5Y and IMR-32 cells to doxorubicin in both 2D and 3D culture systems. Microarray gene expression profiling suggested numbers of candidate target genes of WNT inhibition by XAV939, in particular, p21, p53, ubiquitin C, ZBED8, MDM2, CASP3, and FZD1, and this explained the enhanced sensitivity of SH-SY5Y cells to doxorubicin. Altogether, these results proposed that the altered differentiation of human malignant neuroblastoma cells by inhibiting WNT signaling sensitized the cells to anticancer drugs. This approach could thus serve as an effective treatment option for aggressive brain malignancy.

  4. Quantification of rifampicin in human plasma and cerebrospinal fluid by a highly sensitive and rapid liquid chromatographic–tandem mass spectrometric method

    OpenAIRE

    Srivastava, Abhishek; Waterhouse, David; Ardrey, Alison; Ward, Stephen A.

    2012-01-01

    A highly sensitive and rapid liquid chromatography tandem mass spectrometry (LC–MS/MS) method has been developed to measure the levels of the antitubercular drug rifampicin (RIF) in human plasma and cerebrospinal fluid (CSF). The analyte and internal standard (IS) were isolated from plasma and CSF by a simple organic solvent based precipitation of proteins followed by centrifugation. Detection was carried out by electrospray positive ionization mass spectrometry in the multiple-reaction monit...

  5. Depletion of Human DNA in Spiked Clinical Specimens for Improvement of Sensitivity of Pathogen Detection by Next-Generation Sequencing

    OpenAIRE

    Hasan, Mohammad R.; Rawat, Arun; Tang, Patrick; Jithesh, Puthen V.; Thomas, Eva; Tan, Rusung; Tilley, Peter

    2016-01-01

    Next-generation sequencing (NGS) technology has shown promise for the detection of human pathogens from clinical samples. However, one of the major obstacles to the use of NGS in diagnostic microbiology is the low ratio of pathogen DNA to human DNA in most clinical specimens. In this study, we aimed to develop a specimen-processing protocol to remove human DNA and enrich specimens for bacterial and viral DNA for shotgun metagenomic sequencing. Cerebrospinal fluid (CSF) and nasopharyngeal aspi...

  6. Minimization of nonspecific binding to improve the sensitivity of a magnetic immunoradiometric assay (IRMA) for human thyrotropin (hTSH)

    International Nuclear Information System (INIS)

    Peroni, C.N.; Ribela, M.T.C.P.; Bartolini, P.

    1996-01-01

    An IRMA of hTSH, based on magnetic solid phase separation, was studied especially in terms of its nonspecific bindings (B 0 ). These were identified as a product of the interaction between radioiodinated anti-hTSH monoclonal antibody ( 125 I-mAB) and the uncoupled magnetizable cellulose particle (matrix). The negative effects of B 0 on the assay performance were minimized and practically eliminated, in the optimized system, with tracer storage at 4 deg. C, repurification and pre-incubation with the same matrix, serum addition during incubation and solid phase saturation with milk proteins. These findings were used in order to reproducibly decrease non specific binding to values 60 /B 0 ) into values of 300-500. This way, hTSH IRMAs were obtained with functional sensitivities of about 0.05 mlU/L and analytical sensitivities of the order of 0.02 mlU/L, which represent an approximate 10-fold increase in sensitivity when compared with non-optimized system. A more optimistic sensitivity calculation, based on Rodbard's definition, provided values down to 0.008 mlU/L. Such sensitivities, moreover, were obtained in a very reproducible way and all over the useful tracer life. (author). 10 refs, 1 fig., 8 tabs

  7. Skin Sensitive Difference of Human Body Sections under Clothing--Multiple Analysis of Skin Surface Temperature Changes

    Institute of Scientific and Technical Information of China (English)

    李俊; 吴海燕; 张渭源

    2003-01-01

    A new researching method on clothing comfort perception is developed.By it the skin surface temperature changes and subjective psychological perception of human body sections stimulated by the same cold stimulation are studied.With the multiple comparison analysis method the changing laws of skin temperature of main human body sections is obtained.

  8. Evaluation of selected biomarkers for the detection of chemical sensitization in human skin: a comparative study applying THP-1, MUTZ-3 and primary dendritic cells in culture.

    Science.gov (United States)

    Hitzler, Manuel; Bergert, Antje; Luch, Andreas; Peiser, Matthias

    2013-09-01

    Dendritic cells (DCs) exhibit the unique capacity to induce T cell differentiation and proliferation, two processes that are crucially involved in allergic reactions. By combining the exclusive potential of DCs as the only professional antigen-presenting cells of the human body with the well known handling advantages of cell lines, cell-based alternative methods aimed at detecting chemical sensitization in vitro commonly apply DC-like cells derived from myeloid cell lines. Here, we present the new biomarkers programmed death-ligand 1 (PD-L1), DC immunoreceptor (DCIR), IL-16, and neutrophil-activating protein-2 (NAP-2), all of which have been detectable in primary human DCs upon exposure to chemical contact allergens. To evaluate the applicability of DC-like cells in the prediction of a chemical's sensitization potential, the expression of cell surface PD-L1 and DCIR was analyzed. In contrast to primary DCs, only minor subpopulations of MUTZ-3 and THP-1 cells presented PD-L1 or DCIR at their surface. After exposure to increasing concentrations of nickel and cinnamic aldehyde, the expression level of PD-L1 and DCIR revealed much stronger affected on monocyte-derived DCs (MoDCs) or Langerhans cells (MoLCs) when compared to THP-1 and MUTZ-3 cells. Applying protein profiler arrays we further identified the soluble factors NAP-2, IL-16, IL-8 and MIP-1α as sensitive biomarkers showing the capacity to discriminate sensitizing from non-sensitizing chemicals or irritants. An allergen-specific release of IL-8 and MIP-1α could be detected in the supernatants of MoDCs and MoLCs and also in MUTZ-3 and THP-1 cells, though at much lower levels. On the protein and transcriptional level, NAP-2 and IL-16 indicated sensitizers most sensitively and specifically in MoDCs. Altogether, we have proven the reciprocal regulated surface molecules PD-L1 and DCIR and the soluble factors MIP-1α, NAP-2 and IL-16 as reliable biomarkers for chemical sensitization. We further show that primary

  9. Need for a gender-sensitive human security framework: results of a quantitative study of human security and sexual violence in Djohong District, Cameroon.

    Science.gov (United States)

    Parmar, Parveen Kaur; Agrawal, Pooja; Goyal, Ravi; Scott, Jennifer; Greenough, P Gregg

    2014-01-01

    Human security shifts traditional concepts of security from interstate conflict and the absence of war to the security of the individual. Broad definitions of human security include livelihoods and food security, health, psychosocial well-being, enjoyment of civil and political rights and freedom from oppression, and personal safety, in addition to absence of conflict. In March 2010, we undertook a population-based health and livelihood study of female refugees from conflict-affected Central African Republic living in Djohong District, Cameroon and their female counterparts within the Cameroonian host community. Embedded within the survey instrument were indicators of human security derived from the Leaning-Arie model that defined three domains of psychosocial stability suggesting individuals and communities are most stable when their core attachments to home, community and the future are intact. While the female refugee human security outcomes describe a population successfully assimilated and thriving in their new environments based on these three domains, the ability of human security indicators to predict the presence or absence of lifetime and six-month sexual violence was inadequate. Using receiver operating characteristic (ROC) analysis, the study demonstrates that common human security indicators do not uncover either lifetime or recent prevalence of sexual violence. These data suggest that current gender-blind approaches of describing human security are missing serious threats to the safety of one half of the population and that efforts to develop robust human security indicators should include those that specifically measure violence against women.

  10. Cultural variation is part of human nature : Literary universals, context-sensitivity, and "shakespeare in the bush".

    Science.gov (United States)

    Sugiyama, Michelle Scalise

    2003-12-01

    In 1966, Laura Bohannan wrote her classic essay challenging the supposition that great literary works speak to universal human concerns and conditions and, by extension, that human nature is the same everywhere. Her evidence: the Tiv of West Africa interpret Hamlet differently from Westerners. While Bohannan's essay implies that cognitive universality and cultural variation are mutually exclusive phenomena, adaptationist theory suggests otherwise. Adaptive problems ("the human condition") and cognitive adaptations ("human nature") are constant across cultures. What differs between cultures is habitat: owing to environmental variation, the means and information relevant to solving adaptive problems differ from place to place. Thus, we find differences between cultures not because human minds differ in design but largely because human habitats differ in resources and history. On this view, we would expect world literature to express both human universals and cultural particularities. Specifically, we should expect to find literary universality at the macro level (e.g., adaptive problems, cognitive adaptations) and literary variation at the micro level (e.g., local solutions to adaptive problems).

  11. Drug and radiation sensitivity measurements of successful primary monolayer culturing of human tumor cells using cell-adhesive matrix and supplemented medium

    International Nuclear Information System (INIS)

    Baker, F.L.; Spitzer, G.; Ajani, J.A.

    1986-01-01

    The limitations of the agar suspension culture method for primary culturing of human tumor cells prompted development of a monolayer system optimized for cell adhesion and growth. This method grew 83% of fresh human tumor cell biopsy specimens, cultured and not contaminated, from a heterogeneous group of 396 tumors including lung cancer (93 of 114, 82%); melanoma (54 of 72, 75%); sarcoma (46 of 59, 78%); breast cancer (35 of 39, 90%); ovarian cancer (16 of 21, 76%); and a miscellaneous group consisting of gastrointestinal, genitourinary, mesothelioma, and unknown primaries (78 of 91, 86%). Cell growth was characterized morphologically with Papanicolaoustained coverslip cultures and cytogenetically with Giemsastained metaphase spreads. Morphological features such as nuclear pleomorphism, chromatin condensation, basophilic cytoplasm, and melanin pigmentation were routinely seen. Aneuploid metaphases were seen in 90% of evaluable cultures, with 15 of 28 showing 70% or more aneuploid metaphases. Colony-forming efficiency ranged between 0.01 and 1% of viable tumor cells, with a median efficiency of 0.2%. This culture system uses a low inoculum of 25,000 viable cells per well which permitted chemosensitivity testing of nine drugs at four doses in duplicate from 2.2 X 10(6) viable tumor cells and radiation sensitivity testing at five doses in quadruplicate from 0.6 X 10(6) cells. Cultures were analyzed for survival by computerized image analysis of crystal violet-stained cells. Drug sensitivity studies showed variability in sensitivity and in survival curve shape with exponential cell killing for cisplatin, Adriamycin, and etoposide, and shouldered survival curves for 5-fluorouracil frequently seen. Radiation sensitivity studies also showed variability in both sensitivity and survival curve shape. Many cultures showed exponential cell killing, although others had shouldered survival curves

  12. Development of a sensitive ELISA for the quantification of human tumour necrosis factor-alpha using 4 polyclonal antibodies.

    NARCIS (Netherlands)

    Grebenchtchikov, N.J.; Ven-Jongekrijg, J. van der; Pesman, G.J.; Geurts-Moespot, A.; Meer, J.W.M. van der; Sweep, C.G.J.

    2005-01-01

    Despite the availability of many assays to measure concentrations of tumour necrosis factor alpha (TNF-alpha) in body fluids, these assays often lack specificity or sensitivity and are often of questionable reliability, resulting in inconsistent results. Therefore, we have developed an ELISA that is

  13. Role of IgG4 in histamine release from human basophil leucocytes. I. Sensitization of cells from normal donors

    DEFF Research Database (Denmark)

    Poulsen, L K; Stahl Skov, P; Mosbech, H

    1988-01-01

    Several conflicting reports on the ability of IgG4 to mediate type I allergic reactions have appeared lately. We have developed a model system for testing this possibility, using passive sensitization of basophil leucocytes from normal individuals. At first, the system was optimized with regard t...

  14. Investigation of some parameters influencing the sensitivity of human tooth enamel to gamma radiation using electron paramagnetic resonance

    International Nuclear Information System (INIS)

    El-Faramawy, N.

    2008-01-01

    Electron paramagnetic resonance (EPR) has been successfully used as a physical technique for gamma radiation dose reconstruction using calcified tissues. To minimize potential discrepancies between EPR readings in future studies, the effects of cavity response factor, tooth position and donor gender on the estimated gamma radiation dose were studied. It was found that the EPR response per sample mass used for assessment of doses in teeth outside of the 70-100 mg range should be corrected by a factor which is a function of the sample mass. In the EPR measurements, the difference in sensitivity of different tooth positions to γ-radiation was taken into consideration. It was determined that among all the premolars and molars tooth positions, the relative standard deviation of sensitivity was 6.5%, with the wisdom teeth and the first molars having the highest and lowest sensitivity to γ-radiation, respectively. The current results reveal no effect of the donor gender on the sensitivity to γ-radiation. (author)

  15. Sensitive Media

    Directory of Open Access Journals (Sweden)

    Malinowska Anna

    2017-12-01

    Full Text Available The paper engages with what we refer to as “sensitive media,” a concept associated with developments in the overall media environment, our relationships with media devices, and the quality of the media themselves. Those developments point to the increasing emotionality of the media world and its infrastructures. Mapping the trajectories of technological development and impact that the newer media exert on human condition, our analysis touches upon various forms of emergent affect, emotion, and feeling in order to trace the histories and motivations of the sensitization of “the media things” as well as the redefinition of our affective and emotional experiences through technologies that themselves “feel.”

  16. Modelling ecological and human exposure to POPs in Venice lagoon - Part II: Quantitative uncertainty and sensitivity analysis in coupled exposure models.

    Science.gov (United States)

    Radomyski, Artur; Giubilato, Elisa; Ciffroy, Philippe; Critto, Andrea; Brochot, Céline; Marcomini, Antonio

    2016-11-01

    The study is focused on applying uncertainty and sensitivity analysis to support the application and evaluation of large exposure models where a significant number of parameters and complex exposure scenarios might be involved. The recently developed MERLIN-Expo exposure modelling tool was applied to probabilistically assess the ecological and human exposure to PCB 126 and 2,3,7,8-TCDD in the Venice lagoon (Italy). The 'Phytoplankton', 'Aquatic Invertebrate', 'Fish', 'Human intake' and PBPK models available in MERLIN-Expo library were integrated to create a specific food web to dynamically simulate bioaccumulation in various aquatic species and in the human body over individual lifetimes from 1932 until 1998. MERLIN-Expo is a high tier exposure modelling tool allowing propagation of uncertainty on the model predictions through Monte Carlo simulation. Uncertainty in model output can be further apportioned between parameters by applying built-in sensitivity analysis tools. In this study, uncertainty has been extensively addressed in the distribution functions to describe the data input and the effect on model results by applying sensitivity analysis techniques (screening Morris method, regression analysis, and variance-based method EFAST). In the exposure scenario developed for the Lagoon of Venice, the concentrations of 2,3,7,8-TCDD and PCB 126 in human blood turned out to be mainly influenced by a combination of parameters (half-lives of the chemicals, body weight variability, lipid fraction, food assimilation efficiency), physiological processes (uptake/elimination rates), environmental exposure concentrations (sediment, water, food) and eating behaviours (amount of food eaten). In conclusion, this case study demonstrated feasibility of MERLIN-Expo to be successfully employed in integrated, high tier exposure assessment. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Sensitivity of human cells expressing low-fidelity or weak-catalytic-activity variants of DNA polymerase ζ to genotoxic stresses.

    Science.gov (United States)

    Suzuki, Tetsuya; Grúz, Petr; Honma, Masamitsu; Adachi, Noritaka; Nohmi, Takehiko

    2016-09-01

    Translesion DNA polymerases (TLS pols) play critical roles in defense mechanisms against genotoxic agents. The defects or mutations of TLS pols are predicted to result in hypersensitivity of cells to environmental mutagens. In this study, human cells expressing DNA polymerase ζ (Pol ζ) variants with low fidelity or weak catalytic activity have been established with Nalm-6-MSH+ cells and their sensitivity to mutagenicity and cytotoxicity of benzo[a]pyrene diol epoxide (BPDE) and ultraviolet-C light (UV-C) was examined. The low-fidelity mutants were engineered by knocking-in DNA sequences that direct changes of leucine 2618 to either phenylalanine (L2618F) or methionine (L2618M) of Pol ζ. The weak-catalytic-activity mutants were generated by knocking-in DNA sequences that direct changes of either tyrosine 2779 to phenylalanine (Y2779F) or aspartate 2781 to asparagine (D2781N). In addition, a +1 frameshift mutation, i.e., CCC to CCCC, was introduced in the coding region of the TK1 gene to measure the mutant frequencies. Doubling time and spontaneous TK mutant frequencies of the established cell lines were similar to those of the wild-type cells. The low-fidelity mutants displayed, however, higher sensitivity to the mutagenicity of BPDE and UV-C than the wild-type cells although their cytotoxic sensitivity was not changed. In contrast, the weak-catalytic-activity mutants were more sensitive to the cytotoxicity of BPDE and UV-C than the wild-type cells, and displayed much higher sensitivity to the clastogenicity of BPDE than the wild-type cells in an in vitro micronucleus assay. These results indicate that human Pol ζ is involved in TLS across DNA lesions induced by BPDE and UV-C and also that the TLS plays important roles in induction of mutations, clastogenicity and in cellular survival of the damaged human cells. Similarities and differences in in vivo roles of yeast and human Pol ζ in genome integrity are discussed. Copyright © 2016 Elsevier B.V. All rights

  18. Gold nanoparticle labeling with tyramide signal amplification for highly sensitive detection of alpha fetoprotein in human serum by ICP-MS.

    Science.gov (United States)

    Li, Xiaoting; Chen, Beibei; He, Man; Xiao, Guangyang; Hu, Bin

    2018-01-01

    In this work, we developed an immunoassay based on tyramide signal amplification (TSA) and gold nanoparticles (Au NPs) labeling for highly sensitive detection of alpha fetoprotein (AFP) by inductively coupled plasma mass spectrometry (ICP-MS). AFP was captured by anti-AFP1 coating on the 96-well plate and labeled by anti-AFP2-horseradish peroxidase (HRP), in which the HRP can catalyze the deposition of biotinylated tyramine on the nearby protein. Then the streptavidin (SA)-Au NPs was labeled on the deposited biotinylated tyramine as the intensive signal probe for ICP-MS measurement. Under the optimal experimental conditions, the limit of detection of the developed method for AFP was 1.85pg/mL and the linear range was 0.005-2ng/mL. The relative standard deviation for seven replicate detections of 0.01ng/mL AFP was 5.2%. The proposed method was successfully applied to the detection of AFP in human serum with good recoveries. This strategy is highly sensitive and easy to operate, and can be extended to the sensitive detection of other biomolecules in human serum. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Evidence for Non-Opponent Coding of Colour Information in Human Visual Cortex: Selective Loss of "Green" Sensitivity in a Subject with Damaged Ventral Occipito-Temporal Cortex.

    Science.gov (United States)

    Rauscher, Franziska G; Plant, Gordon T; James-Galton, Merle; Barbur, John L

    2011-01-01

    Damage to ventral occipito-temporal extrastriate visual cortex leads to the syndrome of prosopagnosia often with coexisting cerebral achromatopsia. A patient with this syndrome resulting in a left upper homonymous quadrantanopia, prosopagnosia, and incomplete achromatopsia is described. Chromatic sensitivity was assessed at a number of locations in the intact visual field using a dynamic luminance contrast masking technique that isolates the use of colour signals. In normal subjects chromatic detection thresholds form an elliptical contour when plotted in the Commission Internationale d'Eclairage, (x-y), chromaticity diagram. Because the extraction of colour signals in early visual processing involves opponent mechanisms, subjects with Daltonism (congenital red/green loss of sensitivity) show symmetric increase in thresholds towards the long wavelength ("red") and middle wavelength ("green") regions of the spectrum locus. This is also the case with acquired loss of chromatic sensitivity as a result of retinal or optic nerve disease. Our patient's results were an exception to this rule. Whilst his chromatic sensitivity in the central region of the visual field was reduced symmetrically for both "red/green" and "yellow/blue" directions in colour space, the subject's lower left quadrant showed a marked asymmetry in "red/green" thresholds with the greatest loss of sensitivity towards the "green" region of the spectrum locus. This spatially localized asymmetric loss of "green" but not "red" sensitivity has not been reported previously in human vision. Such loss is consistent with selective damage of neural substrates in the visual cortex that process colour information, but are spectrally non-opponent.

  20. The liver X receptor agonist T0901317 acts as androgen receptor antagonist in human prostate cancer cells

    International Nuclear Information System (INIS)

    Chuu, Chih-pin; Chen, Rou-Yu; Hiipakka, Richard A.; Kokontis, John M.; Warner, Karen V.; Xiang, Jialing; Liao, Shutsung

    2007-01-01

    T0901317 is a potent non-steroidal synthetic liver X receptor (LXR) agonist. T0901317 blocked androgenic stimulation of the proliferation of androgen-dependent LNCaP 104-S cells and androgenic suppression of the proliferation of androgen-independent LNCaP 104-R2 cells, inhibited the transcriptional activation of an androgen-dependent reporter gene by androgen, and suppressed gene and protein expression of prostate specific antigen (PSA), a target gene of androgen receptor (AR) without affecting gene and protein expression of AR. T0901317 also inhibited binding of a radiolabeled androgen to AR, but inhibition was much weaker compared to the effect of the antiandrogens, bicalutamide and hydroxyflutamide. The LXR agonist T0901317, therefore, acts as an antiandrogen in human prostate cancer cells

  1. S-glutathionylation of troponin I (fast) increases contractile apparatus Ca2+ sensitivity in fast-twitch muscle fibres of rats and humans.

    Science.gov (United States)

    Mollica, J P; Dutka, T L; Merry, T L; Lamboley, C R; McConell, G K; McKenna, M J; Murphy, R M; Lamb, G D

    2012-03-15

    Oxidation can decrease or increase the Ca2+ sensitivity of the contractile apparatus in rodent fast-twitch (type II) skeletal muscle fibres, but the reactions and molecular targets involved are unknown. This study examined whether increased Ca2+ sensitivity is due to S-glutathionylation of particular cysteine residues. Skinned muscle fibres were directly activated in heavily buffered Ca2+ solutions to assess contractile apparatus Ca2+ sensitivity. Rat type II fibres were subjected to S-glutathionylation by successive treatments with 2,2′-dithiodipyridine (DTDP) and glutathione (GSH), and displayed a maximal increase in pCa50 (−log10 [Ca2+] at half-maximal force) of ∼0.24 pCa units, with little or no effect on maximum force or Hill coefficient. Partial similar effect was produced by exposure to oxidized gluthathione (GSSG, 10 mM) for 10 min at pH 7.1, and near-maximal effect by GSSG treatment at pH 8.5. None of these treatments significantly altered Ca2+ sensitivity in rat type I fibres. Western blotting showed that both the DTDP–GSH and GSSG–pH 8.5 treatments caused marked S-glutathionylation of the fast troponin I isoform (TnI(f)) present in type II fibres, but not of troponin C (TnC) or myosin light chain 2. Both the increased Ca2+ sensitivity and glutathionylation of TnI(f) were blocked by N-ethylmaleimide (NEM). S-nitrosoglutathione (GSNO) also increased Ca2+ sensitivity, but only in conditions where it caused S-glutathionylation of TnI(f). In human type II fibres from vastus lateralis muscle, DTDP–GSH treatment also caused similar increased Ca2+ sensitivity and S-glutathionylation of TnI(f). When the slow isoform of TnI in type I fibres of rat was partially substituted (∼30%) with TnI(f), DTDP–GSH treatment caused a significant increase in Ca2+ sensitivity (∼0.08 pCa units). TnIf in type II fibres from toad and chicken muscle lack Cys133 present in mammalian TnIf, and such fibres showed no change in Ca2+ sensitivity with DTDP–GSH nor any S

  2. Sensitivity of mountain ecosystems to human-accelerated soil erosion. Contrasting geomorphic response between tropical and semi-arid ecosystems.

    Science.gov (United States)

    Vanacker, Veerle; Bellin, Nicolas; Schoonejans, Jerome; Molina, Armando; Kubik, Peter W.

    2014-05-01

    Human-induced land cover changes are causing important adverse effects on the ecological services rendered by mountain ecosystems, and the number of case-studies of the impact of humans on soil erosion and sediment yield has mounted rapidly. A modelling framework that is specifically adapted to mountain environments is currently lacking. Most studies make use of general river basin models that were originally parameterized and calibrated for temperate, low relief landscapes. Transposing these modelling concepts directly to steep environments with shallow and stony soils often leads to unrealistic model predictions, as model input parameters are rarely calibrated for the range of environmental conditions found in mountain regions. Here, we present a conceptual model that evaluates erosion regulation as a function of human disturbances in vegetation cover. The basic idea behind this model is that soil erosion mechanisms are independent of human impact, but that the frequency-magnitude distributions of erosion rates change as a response to human disturbances. Pre-disturbance (or natural) erosion rates are derived from in-situ produced 10Be concentrations in river sediment, while post-disturbance (or modern) erosion rates are derived from sedimentation rates in small catchments. In its simplicity, the model uses vegetation cover change as a proxy of human disturbance in a given vegetation system. The model is then calibrated with field measurements from two mountainous sites with strongly different vegetation dynamics, climatic and geological settings: the Tropical Andes, and the Spanish Betic Cordillera. Natural erosion processes are important in mountainous sites, and natural erosion benchmarks are primordial to assess human-induced changes in erosion rates. While the Spanish Betic Cordillera is commonly characterized as a degraded landscape, there is no significant change in erosion due to human disturbance for uncultivated sites. The opposite is true for the

  3. The human cyclin B1 protein modulates sensitivity of DNA mismatch repair deficient prostate cancer cell lines to alkylating agents.

    Science.gov (United States)

    Rasmussen, L J; Rasmussen, M; Lützen, A; Bisgaard, H C; Singh, K K

    2000-05-25

    DNA damage caused by alkylating agents results in a G2 checkpoint arrest. DNA mismatch repair (MMR) deficient cells are resistant to killing by alkylating agents and are unable to arrest the cell cycle in G2 phase after alkylation damage. We investigated the response of two MMR-deficient prostate cancer cell lines DU145 and LNCaP to the alkylating agent MNNG. Our studies reveal that DU145 cancer cells are more sensitive to killing by MNNG than LNCaP. Investigation of the underlying reasons for lower resistance revealed that the DU145 cells contain low endogenous levels of cyclin B1. We provide direct evidence that the endogenous level of cyclin B1 modulates the sensitivity of MMR-deficient prostate cancer cells to alkylating agents.

  4. An in vitro drug sensitivity test using a higher 3H-TdR incorporation and a modified human tumor stem cell assay

    International Nuclear Information System (INIS)

    Wang Enzhong

    1991-01-01

    An in vitro drug sensitivity test was developed to evaluate the lethal effects of drugs on human pulmonary carcinoma cells (HPCC). This method was a variant and combination of Human Tumor Stem (HTSCA) and a short-term test using 3 H-TdR incorporation. It consisted of a cell containing liquid top layer and a soft agar bottom layer in 24-well microplates. The medium was RPMI 1640 supplemented with 20% malignant pleural effusion, which could enhance 3 H-TdR incorporation into malignant cells. When 50%, 40%, 30% and 30% of cell survival rate defined as sensitivity-threshold for VCR, MMC, DDP and ADM respectively, in the vitro effectiveness were close to those of clinical single-drug treatment in HPCC by Wright et al. This method was also compared with HTSCA in ten human lung cancer cell lines and four pulmonary carcinoma tissues. The agreement rates were 83% and 100% respectively. Thus we presume this system is more useful for oncological clinics than the others

  5. Synthesis and evaluation of translocator 18 kDa protein (TSPO) positron emission tomography (PET) radioligands with low binding sensitivity to human single nucleotide polymorphism rs6971.

    Science.gov (United States)

    Zanotti-Fregonara, Paolo; Zhang, Yi; Jenko, Kimberly J; Gladding, Robert L; Zoghbi, Sami S; Fujita, Masahiro; Sbardella, Gianluca; Castellano, Sabrina; Taliani, Sabrina; Martini, Claudia; Innis, Robert B; Da Settimo, Federico; Pike, Victor W

    2014-10-15

    The imaging of translocator 18 kDa protein (TSPO) in living human brain with radioligands by positron emission tomography (PET) has become an important means for the study of neuroinflammatory conditions occurring in several neuropsychiatric disorders. The widely used prototypical PET radioligand [(11)C](R)-PK 11195 ([(11)C](R)-1; [N-methyl-(11)C](R)-N-sec-butyl-1-(2-chlorophenyl)-N-methylisoquinoline-3-carboxamide) gives a low PET signal and is difficult to quantify, whereas later generation radioligands have binding sensitivity to a human single nucleotide polymorphism (SNP) rs6971, which imposes limitations on their utility for comparative quantitative PET studies of normal and diseased subjects. Recently, azaisosteres of 1 have been developed with improved drug-like properties, including enhanced TSPO affinity accompanied by moderated lipophilicity. Here we selected three of these new ligands (7-9) for labeling with carbon-11 and for evaluation in monkey as candidate PET radioligands for imaging brain TSPO. Each radioligand was readily prepared by (11)C-methylation of an N-desmethyl precursor and was found to give a high proportion of TSPO-specific binding in monkey brain. One of these radioligands, [(11)C]7, the direct 4-azaisostere of 1, presents many radioligand properties that are superior to those reported for [(11)C]1, including higher affinity, lower lipophilicity, and stable quantifiable PET signal. Importantly, 7 was also found to show very low sensitivity to the human SNP rs6971 in vitro. Therefore, [(11)C]7 now warrants evaluation in human subjects with PET to assess its utility for imaging TSPO in human brain, irrespective of subject genotype.

  6. Auxetic Foam-Based Contact-Mode Triboelectric Nanogenerator with Highly Sensitive Self-Powered Strain Sensing Capabilities to Monitor Human Body Movement

    KAUST Repository

    Zhang, Steven L.; Lai, Ying-Chih; He, Xu; Liu, Ruiyuan; Zi, Yunlong; Wang, Zhong Lin

    2017-01-01

    The first contact-mode triboelectric self-powered strain sensor using an auxetic polyurethane foam, conductive fabric, and polytetrafluroethylene (PTFE) is fabricated. Utilizing the auxetic properties of the polyurethane foam, the auxetic polyurethane foam would expand into the PTFE when the foam is stretched, causing contact electrification. Due to a larger contact area between the PTFE and the foam as the foam is stretched, this device can serve effectively as a strain sensor. The sensitivity of this method is explored, and this sensor has the highest sensitivity in all triboelectric nanogenerator devices that are used previously as a strain sensor. Different applications of this strain sensor are shown, and this sensor can be used as a human body monitoring system, self-powered scale to measure weight, and a seat belt to measure body movements inside a car seat.

  7. Auxetic Foam-Based Contact-Mode Triboelectric Nanogenerator with Highly Sensitive Self-Powered Strain Sensing Capabilities to Monitor Human Body Movement

    KAUST Repository

    Zhang, Steven L.

    2017-05-15

    The first contact-mode triboelectric self-powered strain sensor using an auxetic polyurethane foam, conductive fabric, and polytetrafluroethylene (PTFE) is fabricated. Utilizing the auxetic properties of the polyurethane foam, the auxetic polyurethane foam would expand into the PTFE when the foam is stretched, causing contact electrification. Due to a larger contact area between the PTFE and the foam as the foam is stretched, this device can serve effectively as a strain sensor. The sensitivity of this method is explored, and this sensor has the highest sensitivity in all triboelectric nanogenerator devices that are used previously as a strain sensor. Different applications of this strain sensor are shown, and this sensor can be used as a human body monitoring system, self-powered scale to measure weight, and a seat belt to measure body movements inside a car seat.

  8. Study on sensitivity of southern blotting hybridization using a 32P-labeled probe of PCR products in detecting human cytomegalovirus

    International Nuclear Information System (INIS)

    Bu Hengfu; Chen Juan; Shen Rongsen; Ma Liren; Xu Yongqiang

    1996-01-01

    Southern blotting hybridization (SBH) using a 32 P-labeled probe is one of the most practical methods for genetic diagnosis of pathogen. On the basis of establishing PCR and nested PCR for detecting human cytomegalovirus (HCMV), a 32 P-labeled probe was prepared with the amplified products of 613 bp PCR outer primers and hybridized with 300 bp inner primer amplified product, resulting in increase in detecting sensitivity from 17 ng (in 1.2% agarose electrophoresis) before SBH to 500 pg (autoradiographed), in other words, increasing the sensitivity of detecting HCMV by 10 2 dilutions after using SBH. The method of PCR and SBH using a 32 P-labeled probe could detect less than 1 gene copy of HCMV, therefore, it is a rapid and reliable diagnosis method for detecting HCMV latent infection

  9. A single amino acid of human immunodeficiency virus type 2 capsid protein affects conformation of two external loops and viral sensitivity to TRIM5α.

    Directory of Open Access Journals (Sweden)

    Tadashi Miyamoto

    Full Text Available We previously reported that human immunodeficiency virus type 2 (HIV-2 carrying alanine or glutamine but not proline at position 120 of the capsid protein (CA could grow in the presence of anti-viral factor TRIM5α of cynomolgus monkey (CM. To elucidate details of the interaction between the CA and TRIM5α, we generated mutant HIV-2 viruses, each carrying one of the remaining 17 possible amino acid residues, and examined their sensitivity to CM TRIM5α-mediated restriction. Results showed that hydrophobic residues or those with ring structures were associated with sensitivity, while those with small side chains or amide groups conferred resistance. Molecular dynamics simulation study revealed a structural basis for the differential TRIM5α sensitivities. The mutations at position 120 in the loop between helices 6 and 7 (L6/7 affected conformation of the neighboring loop between helices 4 and 5 (L4/5, and sensitive viruses had a common L4/5 conformation. In addition, the common L4/5 structures of the sensitive viruses were associated with a decreased probability of hydrogen bond formation between the 97th aspartic acid in L4/5 and the 119th arginine in L6/7. When we introduced aspartic acid-to-alanine substitution at position 97 (D97A of the resistant virus carrying glutamine at position 120 to disrupt hydrogen bond formation, the resultant virus became moderately sensitive. Interestingly, the virus carrying glutamic acid at position 120 showed resistance, while its predicted L4/5 conformation was similar to those of sensitive viruses. The D97A substitution failed to alter the resistance of this particular virus, indicating that the 120th amino acid residue itself is also involved in sensitivity regardless of the L4/5 conformation. These results suggested that a hydrogen bond between the L4/5 and L6/7 modulates the overall structure of the exposed surface of the CA, but the amino acid residue at position 120 is also directly involved in CM TRIM5

  10. Development of an in vitro skin sensitization test using human cell lines; human Cell Line Activation Test (h-CLAT). II. An inter-laboratory study of the h-CLAT.

    Science.gov (United States)

    Sakaguchi, H; Ashikaga, T; Miyazawa, M; Yoshida, Y; Ito, Y; Yoneyama, K; Hirota, M; Itagaki, H; Toyoda, H; Suzuki, H

    2006-08-01

    Recent regulatory changes have placed a major emphasis on in vitro safety testing and alternative models. In regard to skin sensitization tests, dendritic cells (DCs) derived from human peripheral blood have been considered in the development of new in vitro alternatives. Human cell lines have been also reported recently. In our previous study, we suggested that measuring CD86 and/or CD54 expression on THP-1 cells (human monocytic leukemia cell line) could be used as an in vitro skin sensitization method. An inter-laboratory study among two laboratories was undertaken in Japan in order to further develop an in vitro skin sensitization model. In the present study, we used two human cell lines: THP-1 and U-937 (human histiocytic lymphoma cell line). First we optimized our test protocol (refer to the related paper entitled "optimization of the h-CLAT protocol" within this journal) and then we did an inter-laboratory validation with nine chemicals using the optimized protocol. We measured the expression of CD86 and CD54 on the above cells using flow cytometry after a 24h and 48h exposure to six known allergens (e.g., DNCB, pPD, NiSO(4)) and three non-allergens (e.g., SLS, tween 80). For the sample test concentration, four doses (0.1x, 0.5x, 1x, and 2x of the 50% inhibitory concentration (IC(50))) were evaluated. IC(50) was calculated using MTT assay. We found that allergens/non-allergens were better predicted using THP-1 cells compared to U-937 cells following a 24 h and a 48 h exposure. We also found that the 24h treatment time tended to have a better accuracy than the 48 h treatment time for THP-1 cells. Expression of CD86 and CD54 were good predictive markers for THP-1 cells, but for U-937 cells, expression of CD86 was a better predictor than CD54, at the 24h and the 48 h treatment time. The accuracy also improved when both markers (CD86 and CD54) were used as compared with a single marker for THP-1 cells. Both laboratories gave a good prediction of allergen

  11. In Situ CaptureRT-qPCR: A new simple and sensitive method to detect human norovirus in oysters

    Science.gov (United States)

    Human noroviruses (HuNoVs) are the major cause for the non-bacterial acute gastroenteritis worldwide. RT-qPCR is a widely used method to detect HuNoVs. However, the method is unable to discriminate between infectious and non-infectious viruses. Previously, we reported that the receptor mediated in s...

  12. Sensitization of multidrug-resistant human cancer cells to Hsp90 inhibitors by down-regulation of SIRT1

    Science.gov (United States)

    Kim, Hak-Bong; Lee, Su-Hoon; Um, Jee-Hyun; Oh, Won Keun; Kim, Dong-Wan; Kang, Chi-Dug; Kim, Sun-Hee

    2015-01-01

    The effectiveness of Hsp90 inhibitors as anticancer agents was limited in multidrug-resistant (MDR) human cancer cells due to induction of heat shock proteins (Hsps) such as Hsp70/Hsp27 and P-glycoprotein (P-gp)-mediated efflux. In the present study, we showed that resistance to Hsp90 inhibitors of MDR human cancer cells could be overcome with SIRT1 inhibition. SIRT1 knock-down or SIRT1 inhibitors (amurensin G and EX527) effectively suppressed the resistance to Hsp90 inhibitors (17-AAG and AUY922) in several MDR variants of human lymphoblastic leukemia and human breast cancer cell lines. SIRT1 inhibition down-regulated the expression of heat shock factor 1 (HSF1) and subsequently Hsps and facilitated Hsp90 multichaperone complex disruption via hyperacetylation of Hsp90/Hsp70. These findings were followed by acceleration of ubiquitin ligase CHIP-mediated mutant p53 (mut p53) degradation and subsequent down-regulation of P-gp in 17-AAG-treated MDR cancer cells expressing P-gp and mut p53 after inhibition of SIRT1. Therefore, combined treatment with Hsp90 inhibitor and SIRT1 inhibitor could be a more effective therapeutic approach for Hsp90 inhibitor-resistant MDR cells via down-regulation of HSF1/Hsps, mut p53 and P-gp. PMID:26416354

  13. Antioxidants keep the potentially probiotic but highly oxygen-sensitive human gut bacterium Faecalibacterium prausnitzii alive at ambient air

    NARCIS (Netherlands)

    Khan, M. Tanweer; van Dijl, Jan Maarten; Harmsen, Hermie J M

    2014-01-01

    The beneficial human gut microbe Faecalibacterium prausnitzii is a 'probiotic of the future' since it produces high amounts of butyrate and anti-inflammatory compounds. However, this bacterium is highly oxygen-senstive, making it notoriously difficult to cultivate and preserve. This has so far

  14. A Household-Based Distribution-Sensitive Human Development Index: An Empirical Application to Mexico, Nicaragua and Peru

    Science.gov (United States)

    Lopez-Calva, Luis F.; Ortiz-Juarez, Eduardo

    2012-01-01

    In measuring human development, one of the main concerns relates to the inclusion of a measure that penalizes inequalities in the distribution of achievements across the population. Using indicators from nationally representative household surveys and census data, this paper proposes a straightforward methodology to estimate a household-based…

  15. Sensitive genotyping of foodborne-associated human noroviruses and hepatitis A virus using an array-based platform

    Science.gov (United States)

    The viral pathogens, human norovirus (NoV) and hepatitis A virus (HAV), are significant contributors of foodborne associated outbreaks. To develop a typing tool for foodborne viruses, a focused, low-density DNA microarray was developed in conjunction with a rapid and high-throughput fluorescent meth...

  16. Using detergent to enhance detection sensitivity of African trypanosomes in human CSF and blood by loop-mediated isothermal amplification (LAMP.

    Directory of Open Access Journals (Sweden)

    Dennis J Grab

    2011-08-01

    Full Text Available The loop-mediated isothermal amplification (LAMP assay, with its advantages of simplicity, rapidity and cost effectiveness, has evolved as one of the most sensitive and specific methods for the detection of a broad range of pathogenic microorganisms including African trypanosomes. While many LAMP-based assays are sufficiently sensitive to detect DNA well below the amount present in a single parasite, the detection limit of the assay is restricted by the number of parasites present in the volume of sample assayed; i.e. 1 per µL or 10(3 per mL. We hypothesized that clinical sensitivities that mimic analytical limits based on parasite DNA could be approached or even obtained by simply adding detergent to the samples prior to LAMP assay.For proof of principle we used two different LAMP assays capable of detecting 0.1 fg genomic DNA (0.001 parasite. The assay was tested on dilution series of intact bloodstream form Trypanosoma brucei rhodesiense in human cerebrospinal fluid (CSF or blood with or without the addition of the detergent Triton X-100 and 60 min incubation at ambient temperature. With human CSF and in the absence of detergent, the LAMP detection limit for live intact parasites using 1 µL of CSF as the source of template was at best 10(3 parasites/mL. Remarkably, detergent enhanced LAMP assay reaches sensitivity about 100 to 1000-fold lower; i.e. 10 to 1 parasite/mL. Similar detergent-mediated increases in LAMP assay analytical sensitivity were also found using DNA extracted from filter paper cards containing blood pretreated with detergent before card spotting or blood samples spotted on detergent pretreated cards.This simple procedure for the enhanced detection of live African trypanosomes in biological fluids by LAMP paves the way for the adaptation of LAMP for the economical and sensitive diagnosis of other protozoan parasites and microorganisms that cause diseases that plague the developing world.

  17. Radiation and/or hyperthermia sensitivity of human melanoma cells grown for several days in media with reduced pH

    International Nuclear Information System (INIS)

    Zoelzer, F.; Streffer, C.

    1999-01-01

    MeWo cells were used throughout. Cells were incubated for 0, 3 or 6 days in media with pH 6.5, 6.7, 6.9, 7.1 or 7.3. The sensitivity against 250 kV X-rays and hyperthermia at 43 C were determined in the colony-forming assay. The intracellular pH was measured flow cytometrically using 5(and 6)-carboxyfluoroescein. Calibration curves were established with cells incubated in different buffers containing nigericin to equilibrate intra- and extracellular pH. Results: Cell growth was optimal with pH 7.3 and 7.1 in the medium, somewhat reduced at pH 6.9, and largely inhibited at pH 6.7 and 6.5. Radiation and/or hyperthermia sensitivities were noticeably increased after several days of incubation at reduced pH; the surviving fraction after 4 Gy and 1 h at 43 C either alone or in combination being a factor of 2 to 4 lower at pH 6.5 than at 7.3. These changes in sensitivity could not be correlated with changes in the intracellular pH. Cells seemed to be capable of regulating this parameter very well; the flow cytometric measurements revealed that the intracellular pH was 7.2±0.2 irrespective of the extracellular pH in the range considered here. Conclusions: In contrast to the almost generally accepted hypothesis that intracellular pH is decisive for the heat sensitivity, the human melanoma cells studied here became sensitive after a few days of incubation under acidic conditions without changes in the intracellular pH. Other factors seem to be influencing the cellular response to radiation and/or heat under chronically low pH. (orig./MG) [de

  18. Development and validation of a sensitive spectrofluorimetric method for the determination of cilazapril of human plasma, urine, in pure and pharmaceutical preparations

    Science.gov (United States)

    Karasakal, A.

    2015-08-01

    A selective and sensitive spectrofluorimetric method was developed and validated for the determination of cilazapril in human plasma urine, in pure and pharmaceutical preparations. The proposed method is based on derivatization using 1-dimethylaminonaphthalene-5-sulphonyl chloride (dansyl chloride) as fluorogenic agent and measuring the fluorescence of the products at emission wavelengths of 503 nm after excitation at 374 nm. The method was validated for linearity, limit of detection, limit of quantification, precision, accuracy, recovery. The calibration curves were linear over a concentration range of 100-500 and 50-250 ng/mL for plasma and urine, respectively. The limits of detection were calculated to be 0.26 and 31.59 ng/mL for plasma and urine, respectively. The proposed method was applied to study of cilazapril in pure, human plasma, urine, and pharmaceutical preparations.

  19. Quantification of rifampicin in human plasma and cerebrospinal fluid by a highly sensitive and rapid liquid chromatographic-tandem mass spectrometric method.

    Science.gov (United States)

    Srivastava, Abhishek; Waterhouse, David; Ardrey, Alison; Ward, Stephen A

    2012-11-01

    A highly sensitive and rapid liquid chromatography tandem mass spectrometry (LC-MS/MS) method has been developed to measure the levels of the antitubercular drug rifampicin (RIF) in human plasma and cerebrospinal fluid (CSF). The analyte and internal standard (IS) were isolated from plasma and CSF by a simple organic solvent based precipitation of proteins followed by centrifugation. Detection was carried out by electrospray positive ionization mass spectrometry in the multiple-reaction monitoring (MRM) mode. The assay was linear in the concentration range 25-6400 ng/mL with intra- and inter-day precision of <7% and <8%, respectively. The validated method was applied to the study of RIF pharmacokinetics in human CSF and plasma over 25 h period after a 10 mg/kg oral dose. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. Single-cell analysis of targeted transcriptome predicts drug sensitivity of single cells within human myeloma tumors.

    Science.gov (United States)

    Mitra, A K; Mukherjee, U K; Harding, T; Jang, J S; Stessman, H; Li, Y; Abyzov, A; Jen, J; Kumar, S; Rajkumar, V; Van Ness, B

    2016-05-01

    Multiple myeloma (MM) is characterized by significant genetic diversity at subclonal levels that have a defining role in the heterogeneity of tumor progression, clinical aggressiveness and drug sensitivity. Although genome profiling studies have demonstrated heterogeneity in subclonal architecture that may ultimately lead to relapse, a gene expression-based prediction program that can identify, distinguish and quantify drug response in sub-populations within a bulk population of myeloma cells is lacking. In this study, we performed targeted transcriptome analysis on 528 pre-treatment single cells from 11 myeloma cell lines and 418 single cells from 8 drug-naïve MM patients, followed by intensive bioinformatics and statistical analysis for prediction of proteasome inhibitor sensitivity in individual cells. Using our previously reported drug response gene expression profile signature at the single-cell level, we developed an R Statistical analysis package available at https://github.com/bvnlabSCATTome, SCATTome (single-cell analysis of targeted transcriptome), that restructures the data obtained from Fluidigm single-cell quantitative real-time-PCR analysis run, filters missing data, performs scaling of filtered data, builds classification models and predicts drug response of individual cells based on targeted transcriptome using an assortment of machine learning methods. Application of SCATT should contribute to clinically relevant analysis of intratumor heterogeneity, and better inform drug choices based on subclonal cellular responses.

  1. Estimated Aortic Stiffness is Independently Associated with Cardiac Baroreflex Sensitivity in Humans: Role of Aging and Habitual Endurance Exercise

    Science.gov (United States)

    Pierce, Gary L.; Harris, Stephen A.; Seals, Douglas R.; Casey, Darren P.; Barlow, Patrick B.; Stauss, Harald M.

    2016-01-01

    We hypothesized that differences in cardiac baroreflex sensitivity (BRS) would be independently associated with aortic stiffness and augmentation index (AI), clinical biomarkers of cardiovascular disease (CVD) risk, among young sedentary and middle-aged/older sedentary and endurance-trained adults. A total of 36 healthy middle-aged/older (age 55-76 years, n=22 sedentary; n=14 endurance-trained) and 5 young sedentary (age 18-31 years) adults were included in a cross-sectional study. A subset of the middle-aged/older sedentary adults (n=12) completed an 8-week aerobic exercise intervention. Invasive brachial artery blood pressure waveforms were used to compute spontaneous cardiac BRS (via sequence technique) and estimated aortic pulse wave velocity (PWV) and AI (AI, via brachial-aortic transfer function and wave separation analysis). In the cross-sectional study, cardiac BRS was 71% lower in older compared with young sedentary adults (Pendurance exercise (P=0.03). In a regression model that included age, sex, resting heart rate, mean arterial pressure (MAP), body mass index and maximal exercise oxygen uptake, estimated aortic PWV (β±SE = −5.76 ± 2.01, P=0.01) was the strongest predictor of BRS (Model R2=0.59, Pendurance exercise-related differences in cardiac BRS are independently associated with corresponding alterations in aortic PWV among healthy adults, consistent with a mechanistic link between variations in the sensitivity of the baroreflex and aortic stiffness with age and exercise. PMID:26911535

  2. Estimated aortic stiffness is independently associated with cardiac baroreflex sensitivity in humans: role of ageing and habitual endurance exercise.

    Science.gov (United States)

    Pierce, G L; Harris, S A; Seals, D R; Casey, D P; Barlow, P B; Stauss, H M

    2016-09-01

    We hypothesised that differences in cardiac baroreflex sensitivity (BRS) would be independently associated with aortic stiffness and augmentation index (AI), clinical biomarkers of cardiovascular disease risk, among young sedentary and middle-aged/older sedentary and endurance-trained adults. A total of 36 healthy middle-aged/older (age 55-76 years, n=22 sedentary and n=14 endurance-trained) and 5 young sedentary (age 18-31 years) adults were included in a cross-sectional study. A subset of the middle-aged/older sedentary adults (n=12) completed an 8-week-aerobic exercise intervention. Invasive brachial artery blood pressure waveforms were used to compute spontaneous cardiac BRS (via sequence technique), estimated aortic pulse wave velocity (PWV) and AI (AI, via brachial-aortic transfer function and wave separation analysis). In the cross-sectional study, cardiac BRS was 71% lower in older compared with young sedentary adults (Pendurance exercise (P=0.03). In a regression model that included age, sex, resting heart rate, mean arterial pressure (MAP), body mass index and maximal exercise oxygen uptake, estimated aortic PWV (β±s.e.=-5.76±2.01, P=0.01) was the strongest predictor of BRS (model R(2)=0.59, Pendurance-exercise-related differences in cardiac BRS are independently associated with corresponding alterations in aortic PWV among healthy adults, consistent with a mechanistic link between variations in the sensitivity of the baroreflex and aortic stiffness with age and exercise.

  3. Sensitive monitoring of monoterpene metabolites in human urine using two-step derivatisation and positive chemical ionisation-tandem mass spectrometry

    International Nuclear Information System (INIS)

    Schmidt, Lukas; Belov, Vladimir N.; Göen, Thomas

    2013-01-01

    Highlights: •Sensitive monitoring of 10 metabolites of (R)-limonene, α-pinene, and Δ 3 -carene in human urine samples. •Fast and simple sample preparation and derivatisation procedure using two-step silylation for unreactive tertiary hydroxyl groups. •Synthesis of reference substances and isotopically labelled internal standards of (R)-limonene, α-pinene, and Δ 3 -carene metabolites. •Study on (R)-limonene, α-pinene, and Δ 3 -carene metabolite background exposure of 36 occupationally unexposed volunteers. -- Abstract: A gas chromatographic–positive chemical ionisation-tandem mass spectrometric (GC–PCI-MS/MS) method for the simultaneous determination of 10 oxidative metabolites of the monoterpenoid hydrocarbons α-pinene, (R)-limonene, and Δ 3 -carene ((+)-3-carene) in human urine was developed and tested for the monoterpene biomonitoring of the general population (n = 36). The method involves enzymatic cleavage of the glucuronides followed by solid-supported liquid–liquid extraction and derivatisation using a two-step reaction with N,O-bis(trimethylsilyl)-trifluoroacetamide and N-(trimethylsilyl)imidazole. The method proved to be both sensitive and reliable with detection limits ranging from 0.1 to 0.3 μg L −1 . In contrast to the frequent and distinct quantities of (1S,2S,4R)-limonene-1,2-diol, the (1R,2R,4R)-stereoisomer could not be detected. The expected metabolite of (+)-3-carene, 3-caren-10-ol was not detected in any of the samples. All other metabolites were detected in almost all urine samples. The procedure enables for the first time the analysis of trace levels of a broad spectrum of mono- and bicyclic monoterpenoid metabolites (alcohols, diols, and carboxylic acids) in human urine. This analytical procedure is a powerful tool for population studies as well as for the discovery of human metabolism and toxicokinetics of monoterpenes

  4. TAS-116, a novel Hsp90 inhibitor, selectively enhances radio-sensitivity of human cancer cells to X-rays and carbon ion radiation

    Science.gov (United States)

    Lee, Younghyun; Sunada, Shigeaki; Hirakawa, Hirokazu; Fujimori, Akira; Nickoloff, Jac A.; Okayasu, Ryuichi

    2016-01-01

    Hsp90 inhibitors have been investigated as cancer therapeutics in mono-therapy and to augment radiotherapy, however serious adverse effects of early generation Hsp90 inhibitors limited their development. TAS-116 is a novel Hsp90 inhibitor with lower adverse effects than other Hsp90 inhibitors, and here we investigated the radio-sensitizing effects of TAS-116 in low LET X-ray, and high LET carbon ion irradiated human cancer cells and mouse tumor xenografts. TAS-116 decreased cell survival of both X-ray and carbon ion-irradiated human cancer cell lines (HeLa and H1299 cells), and similar to other Hsp90 inhibitors, it did not affect radiosensitivity of non-cancerous human fibroblasts. TAS-116 increased the number of radiation-induced γ-H2AX foci, and delayed the repair of DNA double-strand breaks (DSBs). TAS-116 reduced the expression of proteins that mediate repair of DSBs by homologous recombination (RAD51) and non-homologous end joining (Ku, DNA-PKcs), and suppressed formation of RAD51 foci and phosphorylation/activation of DNA-PKcs. TAS-116 also decreased expression of the cdc25 cell cycle progression marker, markedly increasing G2/M arrest. Combined treatment of mouse tumor xenografts with carbon ions and TAS-116 showed promising delay in tumor growth compared to either individual treatment. These results demonstrate that TAS-116 radio-sensitizes human cancer cells to both X rays and carbon ions by inhibiting the two major DSB repair pathways, and these effects were accompanied by marked cell cycle arrest. The promising results of combination TAS-116 + carbon ion radiation therapy of tumor xenografts justify further exploration of TAS-116 as an adjunct to radiotherapy using low or high LET radiation. PMID:28062703

  5. New sensitive and specific assay for human immunodeficiency virus antibodies using labeled recombinant fusion protein and time-resolved fluoroimmunoassay.

    OpenAIRE

    Siitari, H; Turunen, P; Schrimsher, J; Nunn, M

    1990-01-01

    A new, rapid method for the detection of human immunodeficiency virus type 1 (HIV-1) antibody by time-resolved fluoroimmunoassay (TR-FIA) was developed. In this assay format, microtitration strips were coated with a recombinant fusion protein, and the same protein was labeled with europium and added into the wells simultaneously with the test specimens. The recombinant fusion protein contained the HIV-1 p24 gag protein sequence that carried an insertion, near the carboxyl terminus, of a 23-am...

  6. Amygdala Lesions Reduce Anxiety-like Behavior in a Human Benzodiazepine-Sensitive Approach-Avoidance Conflict Test.

    Science.gov (United States)

    Korn, Christoph W; Vunder, Johanna; Miró, Júlia; Fuentemilla, Lluís; Hurlemann, Rene; Bach, Dominik R

    2017-10-01

    Rodent approach-avoidance conflict tests are common preclinical models of human anxiety disorder. Their translational validity mainly rests on the observation that anxiolytic drugs reduce rodent anxiety-like behavior. Here, we capitalized on a recently developed approach-avoidance conflict computer game to investigate the impact of benzodiazepines and of amygdala lesions on putative human anxiety-like behavior. In successive epochs of this game, participants collect monetary tokens on a spatial grid while under threat of virtual predation. In a preregistered, randomized, double-blind, placebo-controlled trial, we tested the effect of a single dose (1 mg) of lorazepam (n = 59). We then compared 2 patients with bilateral amygdala lesions due to Urbach-Wiethe syndrome with age- and gender-matched control participants (n = 17). Based on a previous report, the primary outcome measure was the effect of intra-epoch time (i.e., an adaptation to increasing potential loss) on presence in the safe quadrant of the spatial grid. We hypothesized reduced loss adaptation in this measure under lorazepam and in patients with amygdala lesions. Lorazepam and amygdala lesions reduced loss adaptation in the primary outcome measure. We found similar results in several secondary outcome measures. The relative reduction of anxiety-like behavior in patients with amygdala lesions was qualitatively and quantitatively indistinguishable from an impact of anterior hippocampus lesions found in a previous report. Our results establish the translational validity of human approach-avoidance conflict tests in terms of anxiolytic drug action. We identified the amygdala, in addition to the hippocampus, as a critical structure in human anxiety-like behavior. Copyright © 2017 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

  7. Sensitivity to the visual field origin of natural image patches in human low-level visual cortex

    Directory of Open Access Journals (Sweden)

    Damien J. Mannion

    2015-06-01

    Full Text Available Asymmetries in the response to visual patterns in the upper and lower visual fields (above and below the centre of gaze have been associated with ecological factors relating to the structure of typical visual environments. Here, we investigated whether the content of the upper and lower visual field representations in low-level regions of human visual cortex are specialised for visual patterns that arise from the upper and lower visual fields in natural images. We presented image patches, drawn from above or below the centre of gaze of an observer navigating a natural environment, to either the upper or lower visual fields of human participants (n = 7 while we used functional magnetic resonance imaging (fMRI to measure the magnitude of evoked activity in the visual areas V1, V2, and V3. We found a significant interaction between the presentation location (upper or lower visual field and the image patch source location (above or below fixation; the responses to lower visual field presentation were significantly greater for image patches sourced from below than above fixation, while the responses in the upper visual field were not significantly different for image patches sourced from above and below fixation. This finding demonstrates an association between the representation of the lower visual field in human visual cortex and the structure of the visual input that is likely to be encountered below the centre of gaze.

  8. In vitro activation of the neuro-transduction mechanism in sensitive organotypic human skin model.

    Science.gov (United States)

    Martorina, Francesca; Casale, Costantino; Urciuolo, Francesco; Netti, Paolo A; Imparato, Giorgia

    2017-01-01

    Recent advances in tissue engineering have encouraged researchers to endeavor the production of fully functional three-dimensional (3D) thick human tissues in vitro. Here, we report the fabrication of a fully innervated human skin tissue in vitro that recapitulates and replicates skin sensory function. Previous attempts to innervate in vitro 3D skin models did not demonstrate an effective functionality of the nerve network. In our approach, we initially engineer functional human skin tissue based on fibroblast-generated dermis and differentiated epidermis; then, we promote rat dorsal root ganglion (DRG) neurons axon ingrowth in the de-novo developed tissue. Neurofilaments network infiltrates the entire native dermis extracellular matrix (ECM), as demonstrated by immunofluorescence and second harmonic generation (SHG) imaging. To prove sensing functionality of the tissue, we use topical applications of capsaicin, an agonist of transient receptor protein-vanilloid 1 (TRPV1) channel, and quantify calcium currents resulting from variations of Ca ++ concentration in DRG neurons innervating our model. Calcium currents generation demonstrates functional cross-talking between dermis and epidermis compartments. Moreover, through a computational fluid dynamic (CFD) analysis, we set fluid dynamic conditions for a non-planar skin equivalent growth, as proof of potential application in creating skin grafts tailored on-demand for in vivo wound shape. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. MSH3 mismatch repair protein regulates sensitivity to cytotoxic drugs and a histone deacetylase inhibitor in human colon carcinoma cells.

    Directory of Open Access Journals (Sweden)

    Jae Myung Park

    Full Text Available MSH3 is a DNA mismatch repair (MMR gene that undergoes frequent somatic mutation in colorectal cancers (CRCs with MMR deficiency. MSH3, together with MSH2, forms the MutSβ heteroduplex that interacts with interstrand cross-links induced by drugs such as cisplatin. To date, the impact of MSH3 on chemosensitivity is unknown.We utilized isogenic HCT116 (MLH1-/MSH3- cells where MLH1 is restored by transfer of chromosome 3 (HCT116+ch3 and also MSH3 by chromosome 5 (HCT116+3+5. We generated HCT116+3+5, SW480 (MLH1+/MSH3+ and SW48 (MLH1-/MSH3+ cells with shRNA knockdown of MSH3. Cells were treated with 5-fluorouracil (5-FU, SN-38, oxaliplatin, or the histone deacetylase (HDAC inhibitor PCI-24781 and cell viability, clonogenic survival, DNA damage and apoptosis were analyzed.MSH3-deficient vs proficient CRC cells showed increased sensitivity to the irinotecan metabolite SN-38 and to oxaliplatin, but not 5-FU, as shown in assays for apoptosis and clonogenic survival. In contrast, suppression of MLH1 attenuated the cytotoxic effect of 5-FU, but did not alter sensitivity to SN-38 or oxaliplatin. The impact of MSH3 knockdown on chemosensitivity to SN-38 and oxaliplatin was maintained independent of MLH1 status. In MSH3-deficient vs proficient cells, SN-38 and oxaliplatin induced higher levels of phosphorylated histone H2AX and Chk2, and similar results were found in MLH1-proficient SW480 cells. MSH3-deficient vs proficient cells showed increased 53BP1 nuclear foci after irradiation, suggesting that MSH3 can regulate DNA double strand break (DSB repair. We then utilized PCI-24781 that interferes with homologous recombination (HR indicated by a reduction in Rad51 expression. The addition of PCI-24781 to oxaliplatin enhanced cytotoxicity to a greater extent compared to either drug alone.MSH3 status can regulate the DNA damage response and extent of apoptosis induced by chemotherapy. The ability of MSH3 to regulate chemosensitivity was independent of MLH1

  10. Evaluation of a sensitive and specific radioimmunoassay for pancreatic glucagon in human plasma and its clinical application

    International Nuclear Information System (INIS)

    Borghi, V.C.; Wajchenberg, B.L.; Albuquerque, R.H.

    1984-01-01

    A glucagon radioimmunoassay employing antiserum specific for pancreatic glucagon is described. Glucagon was radioiodinated by the chloramine T technique and purified on QAE-Sephadex A 25 to a specific activity of 225 μCi/μg. The standard curve allowed measurements from 12 to 500 pg/ml with sensitivity of 17.5 pg/ml, precision of 6.3-14.9% (CV, within-assay) and 5.6-10.7% (CV, between-assay). Recovery was between 82 and 112%. Fasting plasma glucagon levels in diabetics, obese subjects, acromegalics and patients with Cushing's syndrome were greater than in normals (22.0+-91 pg/ml: mean+-SD). Very low glucagon levels after oral glucose suppression (15.2+-3.1 pg/ml) in normals and greatly increased values after arginine in insulin-dependent diabetics (271.0+-132.3 pg/ml) could be determined. (Auth.)

  11. Evaluation of a sensitive and specific radioimmunoassay for pancreatic glucagon in human plasma and its clinical application

    Energy Technology Data Exchange (ETDEWEB)

    Borghi, V.C.; Wajchenberg, B.L.; Albuquerque, R.H. (Instituto de Pesquisas Energeticas e Nucleares, Sao Paulo (Brazil))

    1984-01-16

    A glucagon radioimmunoassay employing antiserum specific for pancreatic glucagon is described. Glucagon was radioiodinated by the chloramine T technique and purified on QAE-Sephadex A 25 to a specific activity of 225 ..mu..Ci/..mu..g. The standard curve allowed measurements from 12 to 500 pg/ml with sensitivity of 17.5 pg/ml, precision of 6.3-14.9% (CV, within-assay) and 5.6-10.7% (CV, between-assay). Recovery was between 82 and 112%. Fasting plasma glucagon levels in diabetics, obese subjects, acromegalics and patients with Cushing's syndrome were greater than in normals (22.0+-91 pg/ml: mean+-SD). Very low glucagon levels after oral glucose suppression (15.2+-3.1 pg/ml) in normals and greatly increased values after arginine in insulin-dependent diabetics (271.0+-132.3 pg/ml) could be determined.

  12. Macrophages and Adipocytes in Human Obesity Adipose Tissue Gene Expression and Insulin Sensitivity During Calorie Restriction and Weight Stabilization

    DEFF Research Database (Denmark)

    Capel, F.; Klimcakova, E.; Viguerie, N.

    2009-01-01

    OBJECTIVE-We investigated the regulation of adipose tissue gene expression during different phases of a dietary weight loss program and its relation with insulin sensitivity. RESEARCH DESIGN AND METHODS-Twenty-two obese women followed a dietary intervention program composed of an energy restriction...... expression profiling was performed using a DNA microarray in a subgroup of eight women. RT-quantitative PCR was used for determination of mRNA levels of 31 adipose tissue macrophage markers (n = 22). RESULTS-Body weight, fat mass, and C-reactive protein level decreased and glucose disposal rate increased...... during the dietary intervention program. Transcriptome profiling revealed two main patterns of variations. The first involved 464 mostly adipocyte genes involved in metabolism that were downregulated during energy restriction, upregulated during weight stabilization, and unchanged during the dietary...

  13. Genetic variation in PNPLA3 (adiponutrin) confers sensitivity to weight loss-induced decrease in liver fat in humans.

    Science.gov (United States)

    Sevastianova, Ksenia; Kotronen, Anna; Gastaldelli, Amalia; Perttilä, Julia; Hakkarainen, Antti; Lundbom, Jesper; Suojanen, Laura; Orho-Melander, Marju; Lundbom, Nina; Ferrannini, Eleuterio; Rissanen, Aila; Olkkonen, Vesa M; Yki-Järvinen, Hannele

    2011-07-01

    The rs738409 C→G single nucleotide polymorphism in the patatin-like phospholipase domain-containing 3 (PNPLA3; adiponutrin) leads to a missense mutation (I148M), which is associated with increased liver fat but not insulin resistance. The I148M mutation impedes triglyceride hydrolysis in vitro, and its carriers have an increased risk of developing severe liver disease. We explored whether the rs738409 PNPLA3 G allele influences the ability of weight loss to decrease liver fat or change insulin sensitivity. We recruited 8 subjects who were homozygous for the rs738409 PNPLA3 G allele (PNPLA3-148MM) and 10 who were homozygous for the rs738409 PNPLA3 C allele (PNPLA3-148II). To allow comparison of changes in liver fat, the groups were matched with respect to baseline age, sex, body mass index, and liver fat. The subjects were placed on a hypocaloric low-carbohydrate diet for 6 d. Liver fat content (proton magnetic resonance spectroscopy), whole-body insulin sensitivity of glucose metabolism (euglycemic clamp technique), and lipolysis ([(2)H(5)]glycerol infusion) were measured before and after the diet. At baseline, fasting serum insulin and C-peptide concentrations were significantly lower in the PNPLA3-148MM group than in the PNPLA3-148II group, as predicted by study design. Weight loss was not significantly different between groups (PNPLA3-148MM: -3.1 ± 0.5 kg; PNPLA3-148II: -3.1 ± 0.4 kg). Liver fat decreased by 45% in the PNPLA3-148MM group (P loss is effective in decreasing liver fat in subjects who are homozygous for the rs738409 PNPLA3 G or C allele. This trial was registered at www.hus.fi as 233775.

  14. Sensiprobe—a miniature thermal device incorporating Peltier technology as a diagnostic tool for studying human oesophageal sensitivity

    International Nuclear Information System (INIS)

    Reeves, J W; Birch, M J; Al-Zinaty, M; Woodland, P; Sifrim, D; Aziz, Q

    2014-01-01

    Heightened perception of gastrointestinal sensation is termed visceral hypersensitivity (VH) and is commonly observed in patients with gastrointestinal disorders. VH is thought to be a major contributory factor in oesophageal disease, particularly gastro-oesophageal reflux disease that does not respond to standard (proton pump inhibitor) treatment, and in functional heartburn. Clinical tools that can help phenotype according to the mechanism of chronic pain and thus allow targeted drug treatment (e.g. with pain modulator therapy) would be very desirable. A technique that produces repeatable and controllable thermal stimuli within the oesophagus could meet this need. The aims of this study were to develop a method for linear control of the heat stimulation in the oesophagus, to assess the reproducibility of this method, and obtain normal thermal sensitivity values in the distal and proximal oesophagus. The 7 mm diameter Peltier-based thermal device was investigated on 27 healthy subjects using a heating ramp of 0.2 °C s −1 . The pain detection threshold (PDT) temperature was recorded. To assess the reproducibility of the device, each subject underwent the procedure twice, with a minimum of two weeks between each procedure. The mean PDT temperature measured in the distal oesophagus, was 53.8 ± 2.9 °C and 53.6 ± 2.6 °C, for visits 1 and 2 respectively. The mean PDT temperature measured in the proximal oesophagus was 54.1 ± 2.4 °C and 54.0 ± 2.8 °C, for visits 1 and 2 respectively. The reproducibility of the PDT temperature in the distal and proximal oesophagus, was good (intra-class correlation >0.6). Future studies should be aimed to determine whether oesophageal thermal sensitivity can act as a biomarker of transient receptor potential vallanoid 1 upregulation. (paper)

  15. The dual PI3K/mTOR inhibitor PKI-587 enhances sensitivity to cetuximab in EGFR-resistant human head and neck cancer models.

    Science.gov (United States)

    D'Amato, V; Rosa, R; D'Amato, C; Formisano, L; Marciano, R; Nappi, L; Raimondo, L; Di Mauro, C; Servetto, A; Fusciello, C; Veneziani, B M; De Placido, S; Bianco, R

    2014-06-10

    Cetuximab is the only targeted agent approved for the treatment of head and neck squamous cell carcinomas (HNSCC), but low response rates and disease progression are frequently reported. As the phosphoinositide 3-kinase (PI3K) and the mammalian target of rapamycin (mTOR) pathways have an important role in the pathogenesis of HNSCC, we investigated their involvement in cetuximab resistance. Different human squamous cancer cell lines sensitive or resistant to cetuximab were tested for the dual PI3K/mTOR inhibitor PF-05212384 (PKI-587), alone and in combination, both in vitro and in vivo. Treatment with PKI-587 enhances sensitivity to cetuximab in vitro, even in the condition of epidermal growth factor receptor (EGFR) resistance. The combination of the two drugs inhibits cells survival, impairs the activation of signalling pathways and induces apoptosis. Interestingly, although significant inhibition of proliferation is observed in all cell lines treated with PKI-587 in combination with cetuximab, activation of apoptosis is evident in sensitive but not in resistant cell lines, in which autophagy is pre-eminent. In nude mice xenografted with resistant Kyse30 cells, the combined treatment significantly reduces tumour growth and prolongs mice survival. Phosphoinositide 3-kinase/mammalian target of rapamycin inhibition has an important role in the rescue of cetuximab resistance. Different mechanisms of cell death are induced by combined treatment depending on basal anti-EGFR responsiveness.

  16. Apoptosis-related molecular differences for response to tyrosin kinase inhibitors in drug-sensitive and drug-resistant human bladder cancer cells

    Directory of Open Access Journals (Sweden)

    Jixia Li

    2013-01-01

    Full Text Available Context: The epidermal growth factor receptor (EGFR family is reportedly overexpressed in bladder cancer, and tyrosine kinaseinhibitors (TKIs have been suggested as treatment. Gefitinib is a selective inhibitor of the EGFR and lapatinib is a dual inhibitor of both the EGFR and HER2 (human EGFR type 2 receptor. Both compounds compete with the binding of adenosine triphosphate (ATP to the tyrosine kinase domain of the respective receptors to inhibit receptor autophosphorylation causing suppression of signal transduction. Unfortunately, resistance to these inhibitors is a major clinical problem. Aims: To compare the apoptosis signaling pathway(s induced by gefitinib and lapatinib, in UM-UC-5 (drug-sensitive and UM-UC-14 (drug-resistant bladder cancer cells and to identify molecular differences that might be useful predictors of their efficacy. Materials and Methods: Cell proliferation, cell cycle and apoptosis assay were used to detect the effect of TKIs on UM-UC-5 and UM-UC-14 cells. Molecular differences for response to TKIs were examined by protein array. Results: TKIs strongly inhibited cell proliferation and induced cell cycle G1 arrest and apoptosis in UM-UC-5 cells. Most notable apoptosis molecular differences included decreased claspin, trail, and survivin by TKIs in the sensitive cells. In contrast, TKIs had no effect on resistant cells. Conclusions: Claspin, trail, and survivin might be used to determine the sensitivity of bladder cancers to TKIs.

  17. Panel-reactive antibody levels and renal transplantation rates in sensitized patients after desensitization and human leucocyte antigen amino acid residue matching.

    Science.gov (United States)

    Shang, Wenjun; Dong, Laidong; Feng, Guiwen; Wang, Yue; Pang, Xinlu; Li, Jinfeng; Liu, Lei; Zhang, Weihong

    2013-08-01

    To determine whether a new desensitization protocol (mycophenolate mofetil [MMF], plasmapheresis and antithymocyte globulin [ATG], complemented with human leucocyte antigen [HLA] amino acid residue matching) could reduce panel-reactive antibody (PRA) levels in sensitized patients, to facilitate successful renal transplantation. Patients awaiting transplantation with PRA levels >10% received treatment with MMF; those with PRA levels >30% were also treated with plasmapheresis. Patients whose PRA level was desensitization were eligible for transplantation. When a donor became available, traditional HLA matching and HLA amino acid residue matching were performed. All patients received ATG induction therapy postoperatively. Thirty-two sensitized patients were enrolled. Desensitization produced a significant decrease in PRA levels; 27 patients (84.4%) became eligible for transplantation and 26 (81.2%) subsequently underwent successful transplantation. Residue matching improved the proportion with a mismatch number of 0-1 from 7.7% to 65.4%, compared with traditional HLA matching. Postoperatively, all patients showed immediate graft function. Acute rejection occurred in three patients (11.5%) and infections in seven patients (25.9%); all were treated successfully. The combination of a desensitization protocol (MMF, plasmapheresis and ATG) and residue matching appears to be an effective strategy for sensitized patients awaiting renal transplantation.

  18. Regulating Chondrogenesis of Human Mesenchymal Stromal Cells with a Retinoic Acid Receptor-Beta Inhibitor: Differential Sensitivity of Chondral Versus Osteochondral Development

    Directory of Open Access Journals (Sweden)

    Solvig Diederichs

    2014-05-01

    Full Text Available Aim: Main objective was to investigate whether the synthetic retinoic acid receptor (RAR-β antagonist LE135 is able to drive in vitro chondrogenesis of human mesenchymal stromal cells (MSCs or improve differentiation by suppressing hypertrophic chondrocyte development. Methods: Chondrogenesis of human bone marrow and adipose tissue-derived MSCs was induced in micromass pellet culture for six weeks. Effects of LE135 alone and in combinatorial treatment with TGF-β on deposition of cartilaginous matrix including collagen type II and glycosaminoglycans, on deposition of non-hyaline cartilage collagens type I and X, and on hypertrophy markers including alkaline phosphatase (ALP, indian hedghehog (IHH and matrix metalloproteinase (MMP-13 were assessed. Results: LE135 was no inducer of chondrogenesis and failed to stimulate deposition of collagen type II and glycosaminoglycans. Moreover, addition of LE135 to TGF-β-treated pellets inhibited cartilaginous matrix deposition and gene expression of COL2A1. In contrast, non-hyaline cartilage collagens were less sensitive to LE135 and hypertrophy markers remained unaffected. Conclusion: This demonstrates a differential sensitivity of chondral versus endochondral differentiation pathways to RARβ signaling; however, opposite to the desired direction. The relevance of trans-activating versus trans-repressing RAR signaling, including effects on activator protein (AP-1 is discussed and implications for overcoming current limits of hMSC chondrogenesis are considered.

  19. Regulating chondrogenesis of human mesenchymal stromal cells with a retinoic Acid receptor-Beta inhibitor: differential sensitivity of chondral versus osteochondral development.

    Science.gov (United States)

    Diederichs, Solvig; Zachert, Kerstin; Raiss, Patric; Richter, Wiltrud

    2014-01-01

    Main objective was to investigate whether the synthetic retinoic acid receptor (RAR)-β antagonist LE135 is able to drive in vitro chondrogenesis of human mesenchymal stromal cells (MSCs) or improve differentiation by suppressing hypertrophic chondrocyte development. Chondrogenesis of human bone marrow and adipose tissue-derived MSCs was induced in micromass pellet culture for six weeks. Effects of LE135 alone and in combinatorial treatment with TGF-β on deposition of cartilaginous matrix including collagen type II and glycosaminoglycans, on deposition of non-hyaline cartilage collagens type I and X, and on hypertrophy markers including alkaline phosphatase (ALP), indian hedghehog (IHH) and matrix metalloproteinase (MMP)-13 were assessed. LE135 was no inducer of chondrogenesis and failed to stimulate deposition of collagen type II and glycosaminoglycans. Moreover, addition of LE135 to TGF-β-treated pellets inhibited cartilaginous matrix deposition and gene expression of COL2A1. In contrast, non-hyaline cartilage collagens were less sensitive to LE135 and hypertrophy markers remained unaffected. This demonstrates a differential sensitivity of chondral versus endochondral differentiation pathways to RARβ signaling; however, opposite to the desired direction. The relevance of trans-activating versus trans-repressing RAR signaling, including effects on activator protein (AP)-1 is discussed and implications for overcoming current limits of hMSC chondrogenesis are considered. © 2014 S. Karger AG, Basel.

  20. Autophagic cell death induced by reactive oxygen species is involved in hyperthermic sensitization to ionizing radiation in human hepatocellular carcinoma cells.

    Science.gov (United States)

    Yuan, Guang-Jin; Deng, Jun-Jian; Cao, De-Dong; Shi, Lei; Chen, Xin; Lei, Jin-Ju; Xu, Xi-Ming

    2017-08-14

    To investigate whether autophagic cell death is involved in hyperthermic sensitization to ionizing radiation in human hepatocellular carcinoma cells, and to explore the underlying mechanism. Human hepatocellular carcinoma cells were treated with hyperthermia and ionizing radiation. MTT and clonogenic assays were performed to determine cell survival. Cell autophagy was detected using acridine orange staining and flow cytometric analysis, and the expression of autophagy-associated proteins, LC3 and p62, was determined by Western blot analysis. Intracellular reactive oxygen species (ROS) were quantified using the fluorescent probe DCFH-DA. Treatment with hyperthermia and ionizing radiation significantly decreased cell viability and surviving fraction as compared with hyperthermia or ionizing radiation alone. Cell autophagy was significantly increased after ionizing radiation combined with hyperthermia treatment, as evidenced by increased formation of acidic vesicular organelles, increased expression of LC3II and decreased expression of p62. Intracellular ROS were also increased after combined treatment with hyperthermia and ionizing radiation. Pretreatment with N-acetylcysteine, an ROS scavenger, markedly inhibited the cytotoxicity and cell autophagy induced by hyperthermia and ionizing radiation. Autophagic cell death is involved in hyperthermic sensitization of cancer cells to ionizing radiation, and its induction may be due to the increased intracellular ROS.

  1. Transcriptional differences between smokers and non-smokers and variance by obesity as a risk factor for human sensitivity to environmental exposures.

    Science.gov (United States)

    Nikodemova, Maria; Yee, Jeremiah; Carney, Patrick R; Bradfield, Christopher A; Malecki, Kristen Mc

    2018-04-01

    Obesity has been shown to alter response to air pollution and smoking but underlying biological mechanisms are largely unknown and few studies have explored mechanisms by which obesity increases human sensitivity to environmental exposures. Overall study goals were to investigate whole blood gene expression in smokers and non-smokers to examine associations between cigarette smoke and changes in gene expression by obesity status and test for effect modification. Relative fold-change in mRNA expression levels of 84 genes were analyzed using a Toxicity and Stress PCR array among 50 21-54 year old adults. Data on smoking status was confirmed using urinary cotinine levels. Adjusted models included age, gender, white blood cell count and body-mass index. Models comparing gene expression of smokers vs. non-smokers identified six differentially expressed genes associated with smoking after adjustments for covariates. Obesity was associated with 29 genes differentially expressed compared to non-obese. We also identified 9 genes with significant smoking/obesity interactions influencing mRNA levels in adjusted models comparing expression between smokers vs non-smokers for four DNA damage related genes (GADD45A, DDB2, RAD51 and P53), two oxidative stress genes (FTH1, TXN), two hypoxia response genes (BN1P3lL, ARNT), and one gene associated with unfolded protein response (ATF6B). Findings suggest that obesity alters human sensitivity to smoke exposures through several biological pathways by modifying gene expression. Additional studies are needed to fully understand the clinical impact of these effects, but risk assessments should consider underlying phenotypes, such as obesity, that may modulate sensitivity of vulnerable populations to environmental exposures. Copyright © 2018 Elsevier Ltd. All rights reserved.

  2. Prediction of preservative sensitization potential using surface marker CD86 and/or CD54 expression on human cell line, THP-1.

    Science.gov (United States)

    Sakaguchi, Hitoshi; Miyazawa, Masaaki; Yoshida, Yukiko; Ito, Yuichi; Suzuki, Hiroyuki

    2007-02-01

    Preservatives are important components in many products, but have a history of purported allergy. Several assays [e.g., guinea pig maximization test (GPMT), local lymph node assay (LLNA)] are used to evaluate allergy potential of preservatives. We recently developed the human Cell Line Activation Test (h-CLAT), an in vitro skin sensitization test using human THP-1 cells. This test evaluates the augmentation of CD86 and CD54 expression, which are key events in the sensitization process, as an indicator of allergy following treatment with test chemical. Earlier, we found that a sub-toxic concentration was needed for the up-regulation of surface marker expression. In this study, we further evaluate the capability of h-CLAT to predict allergy potential using eight preservatives. Cytotoxicity was determined using propidium iodide with flow cytometry analysis and five doses that produce a 95, 85, 75, 65, and 50% cell viability were selected. If a material did not have any cytotoxicity at the highest technical dose (HTD), five doses are set using serial 1.3 dilutions of the HTD. The test materials used were six known allergic preservatives (e.g., methylchloroisothiazolinone/methylisothiazolinone, formaldehyde), and two non-allergic preservatives (methylparaben and 4-hydroxybenzoic acid). All allergic preservatives augmented CD86 and/or CD54 expression, indicating h-CLAT correctly identified the allergens. No augmentation was observed with the non-allergic preservatives; also correctly identified by h-CLAT. In addition, we report two threshold concentrations that may be used to categorize skin sensitization potency like the LLNA estimated concentration that yield a three-fold stimulation (EC3) value. These corresponding values are the estimated concentration which gives a relative fluorescence intensity (RFI) = 150 for CD86 and an RFI = 200 for CD54. These data suggest that h-CLAT, using THP-1 cells, may be able to predict the allergy potential of preservatives and

  3. A virus-sensitive suppressor cell is involved in the regulation of human allospecific T cell-mediated cytotoxicity

    International Nuclear Information System (INIS)

    Muluk, S.C.; Bernstein, D.C.; Shearer, G.M.

    1989-01-01

    The in vitro generation of allospecific CTL by human PBMC was enhanced 4- to 16-fold by sequential plastic and nylon wool adherence, which depleted the PBMC of macrophages and B cells. The enhanced CTL response was suppressed by adding back irradiated, unfractionated PBMC or adherent cells to the depleted cells. This finding suggests that the enhanced CTL response was not simply a consequence of enrichment of T cells, but was instead due to active suppression by radioresistant cells contained in the adherent fraction. Of note is the finding that, unlike the CTL response, the proliferative response to allostimulation was not affected by the removal of adherent cells. The suppressor function could be abrogated by preincubation of irradiated PBMC with influenza A virus before the coculture with depleted cells. Furthermore, costimulation of unfractionated PBMC with influenza A virus and allogeneic stimulators augmented allospecific CTL activity. Thus, in the adherent fraction of human PBMC, there appears to be a native suppressor population that can be functionally inactivated by virus. This result may account for the clinical observation of increased allograft rejection after certain viral infections

  4. Solar cycle predicts folate-sensitive neonatal genotypes at discrete phases of the first trimester of pregnancy: a novel folate-related human embryo loss hypothesis.

    Science.gov (United States)

    Lucock, Mark; Glanville, Tracey; Yates, Zoë; Walker, James; Furst, John; Simpson, Nigel

    2012-08-01

    Folate, a key periconceptional nutrient, is ultraviolet light (UV-R) sensitive. We therefore hypothesise that a relationship exists between sunspot activity, a proxy for total solar irradiance (particularly UV-R) reaching Earth, and the occurrence of folate-sensitive, epigenomic-related neonatal genotypes during the first trimester of pregnancy. Limited data is provided to support the hypothesis that the solar cycle predicts folate-related human embryo loss: 379 neonates born at latitude 54°N between 1998 and 2000 were examined for three folate-sensitive, epigenome-related polymorphisms, with solar activity for trimester one accessed via the Royal Greenwich Observatory-US Air force/National Oceanic and Atmospheric Administration Sunspot Database (34,110 total observation days). Logistic regression showed solar activity predicts C677T-methylenetetrahydrofolate reductase (C677T-MTHFR) and A66G-methionine synthase reductase (A66G-MSR) genotype at discrete phases of trimester one. Total and maximal sunspot activity predicts C677T-MTHFR genotype for days 31-60 of trimester one (p=0.0181 and 0.0366, respectively) and A66G-MSR genotype for days 61-90 of trimester one (p=0.0072 and 0.0105, respectively). Loss of UV-R sensitive folate associated with the sunspot cycle might therefore interact with variant folate genes to perturb DNA methylation and/or elaboration of the primary base sequence (thymidylate synthesis), as well as increase embryo-toxic homocysteine. We hypothesise that this may influence embryo viability leading to 677CC-MTHFR and 66GG-MSR embryo loss at times of increased solar activity. This provides an interesting and plausible link between well recognised 'folate gene originated developmental disorders' and 'solar activity/seasonality modulated developmental disorders'. Copyright © 2012 Elsevier Ltd. All rights reserved.

  5. The use of a sensitive equilibrium dialysis method for the measurement of free testosterone levels in healthy, cycling women and in human immunodeficiency virus-infected women.

    Science.gov (United States)

    Sinha-Hikim, I; Arver, S; Beall, G; Shen, R; Guerrero, M; Sattler, F; Shikuma, C; Nelson, J C; Landgren, B M; Mazer, N A; Bhasin, S

    1998-04-01

    Measurements of total and free testosterone levels in women have lacked precision and accuracy because of limited assay sensitivity. The paucity of normative data on total and free testosterone levels in healthy women has confounded interpretation of androgen levels in women with human immunodeficiency virus (HIV) infection and other disease states. Therefore, the objectives of this study were to develop sensitive assays for the measurement of the low total and free testosterone levels in women to define the range for these hormones during the normal menstrual cycle and assess the total and free testosterone levels in HIV-infected women. By using a larger volume of serum, increasing the incubation time, and reducing the antibody concentration, the sensitivity of the total testosterone assay was increased to 0.008 nmol/L, and that of the free testosterone assay was increased to 2 pmol/L. The mean percent free testosterone was 1.0 +/- 0.1% of the total testosterone. Serum total and free testosterone levels in the follicular and luteal phases were not significantly different, but both demonstrated a modest preovulatory increase, 3 days before the LH peak. Serum total [0.50 +/- 0.32 (14.60 +/- 9.22) vs. 1.2 +/- 0.7 nmol/L (34.3 +/- 21.0 ng/dL); P copy number. Serum FSH, but not LH, levels were significantly higher in HIV-infected women than in controls. Using assays with sufficient sensitivity, we defined the range for total and free testosterone levels during the normal menstrual cycle. Serum total and free testosterone levels are lower in HIV-infected women and correlate inversely with plasma HIV ribonucleic acid levels. The hypothesis that androgen deficiency contributes to wasting in HIV-infected women remains to be tested.

  6. Distinct Mechanism of Cysteine Oxidation-Dependent Activation and Cold Sensitization of Human Transient Receptor Potential Ankyrin 1 Channel by High and Low Oxaliplatin

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    Takahito Miyake

    2017-11-01

    Full Text Available Oxaliplatin, a third-generation platinum-based chemotherapeutic agent, displays unique acute peripheral neuropathy triggered or enhanced by cold, and accumulating evidence suggests that transient receptor potential ankyrin 1 (TRPA1 is responsible. TRPA1 is activated by oxaliplatin via a glutathione-sensitive mechanism. However, oxaliplatin interrupts hydroxylation of a proline residue located in the N-terminal region of TRPA1 via inhibition of prolyl hydroxylase (PHD, which causes sensitization of TRPA1 to reactive oxygen species (ROS. Furthermore, PHD inhibition endows cold-insensitive human TRPA1 (hTRPA1 with ROS-dependent cold sensitivity. Since cysteine oxidation and proline hydroxylation regulate its activity, their association with oxaliplatin-induced TRPA1 activation and acquirement of cold sensitivity were investigated in the present study. A high concentration of oxaliplatin (1 mM induced outward-rectifier whole-cell currents and increased the intracellular Ca2+ concentration in hTRPA1-expressing HEK293 cells, but did not increase the probability of hTRPA1 channel opening in the inside-out configuration. Oxaliplatin also induced the rapid generation of hydrogen peroxide, and the resultant Ca2+ influx was prevented in the presence of glutathione and in cysteine-mutated hTRPA1 (Cys641Ser-expressing cells, whereas proline-mutated hTRPA1 (Pro394Ala-expressing cells showed similar whole-cell currents and Ca2+ influx. By contrast, a lower concentration of oxaliplatin (100 μM did not increase the intracellular Ca2+ concentration but did confer cold sensitivity on hTRPA1-expressing cells, and this was inhibited by PHD2 co-overexpression. Cold sensitivity was abolished by the mitochondria-targeting ROS scavenger mitoTEMPO and was minimal in cysteine-mutated hTRPA1 (Cys641Ser or Cys665Ser-expressing cells. Thus, high oxaliplatin evokes ROS-mediated cysteine oxidation-dependent hTRPA1 activation independent of PHD activity, while a lower

  7. PAR-2 activation enhances weak acid-induced ATP release through TRPV1 and ASIC sensitization in human esophageal epithelial cells.

    Science.gov (United States)

    Wu, Liping; Oshima, Tadayuki; Shan, Jing; Sei, Hiroo; Tomita, Toshihiko; Ohda, Yoshio; Fukui, Hirokazu; Watari, Jiro; Miwa, Hiroto

    2015-10-15

    Esophageal visceral hypersensitivity has been proposed to be the pathogenesis of heartburn sensation in nonerosive reflux disease. Protease-activated receptor-2 (PAR-2) is expressed in human esophageal epithelial cells and is believed to play a role in inflammation and sensation. PAR-2 activation may modulate these responses through adenosine triphosphate (ATP) release, which is involved in transduction of sensation and pain. The transient receptor potential vanilloid receptor 1 (TRPV1) and acid-sensing ion channels (ASICs) are both acid-sensitive nociceptors. However, the interaction among these molecules and the mechanisms of heartburn sensation are still not clear. We therefore examined whether ATP release in human esophageal epithelial cells in response to acid is modulated by TRPV1 and ASICs and whether PAR-2 activation influences the sensitivity of TRPV1 and ASICs. Weak acid (pH 5) stimulated the release of ATP from primary human esophageal epithelial cells (HEECs). This effect was significantly reduced after pretreatment with 5-iodoresiniferatoxin (IRTX), a TRPV1-specific antagonist, or with amiloride, a nonselective ASIC blocker. TRPV1 and ASIC3 small interfering RNA (siRNA) transfection also decreased weak acid-induced ATP release. Pretreatment of HEECs with trypsin, tryptase, or a PAR-2 agonist enhanced weak acid-induced ATP release. Trypsin treatment led to the phosphorylation of TRPV1. Acid-induced ATP release enhancement by trypsin was partially blocked by IRTX, amiloride, or a PAR-2 antagonist. Conversely, acid-induced ATP release was augmented by PAR-2 activation through TRPV1 and ASICs. These findings suggested that the pathophysiology of heartburn sensation or esophageal hypersensitivity may be associated with the activation of PAR-2, TRPV1, and ASICs. Copyright © 2015 the American Physiological Society.

  8. Network analysis of genes involved in the enhancement of hyperthermia sensitivity by the knockdown of BAG3 in human oral squamous cell carcinoma cells.

    Science.gov (United States)

    Yunoki, Tatsuya; Tabuchi, Yoshiaki; Hayashi, Atsushi; Kondo, Takashi

    2016-07-01

    BCL2-associated athanogene 3 (BAG3), a co-chaperone of the heat shock 70 kDa protein (HSPA) family of proteins, is a cytoprotective protein that acts against various stresses, including heat stress. The aim of the present study was to identify gene networks involved in the enhancement of hyperthermia (HT) sensitivity by the knockdown (KD) of BAG3 in human oral squamous cell carcinoma (OSCC) cells. Although a marked elevation in the protein expression of BAG3 was detected in human the OSCC HSC-3 cells exposed to HT at 44˚C for 90 min, its expression was almost completely suppressed in the cells transfected with small interfering RNA against BAG3 (siBAG) under normal and HT conditions. The silencing of BAG3 also enhanced the cell death that was increased in the HSC-3 cells by exposure to HT. Global gene expression analysis revealed many genes that were differentially expressed by >2-fold in the cells exposed to HT and transfected with siBAG. Moreover, Ingenuity® pathways analysis demonstrated two unique gene networks, designated as Pro-cell death and Anti-cell death, which were obtained from upregulated genes and were mainly associated with the biological functions of induction and the prevention of cell death, respectively. Of note, the expression levels of genes in the Pro-cell death and Anti-cell death gene networks were significantly elevated and reduced in the HT + BAG3-KD group compared to those in the HT control group, respectively. These results provide further insight into the molecular mechanisms involved in the enhancement of HT sensitivity by the silencing of BAG3 in human OSCC cells.

  9. Sub-apoptotic dosages of pro-oxidant vitamin cocktails sensitize human melanoma cells to NK cell lysis.

    Science.gov (United States)

    Tremante, Elisa; Santarelli, Lory; Lo Monaco, Elisa; Sampaoli, Camilla; Ingegnere, Tiziano; Guerrieri, Roberto; Tomasetti, Marco; Giacomini, Patrizio

    2015-10-13

    Alpha-tocopheryl succinate (αTOS), vitamin K3 (VK3) and vitamin C (ascorbic acid, AA) were previously shown to synergistically promote different death pathways in carcinoma cells, depending on their concentrations and combinations. Similar effects were observed herein in melanoma cells, although αTOS behaved as an antagonist. Interestingly, suboptimal cell death-inducing concentrations (1.5 μM αTOS/20 μM AA/0.2 μM VK3) effectively up-regulated activating Natural Killer (NK) cell ligands, including MICA (the stress-signaling ligand of the NKG2D receptor), and/or the ligands of at least one of the natural cytotoxicity receptors (NKp30, NKp44 and NKp46) in 5/6 melanoma cell lines. Only an isolated MICA down-regulation was seen. HLA class I, HLA class II, ULBP1, ULBP2, ULBP3, Nectin-2, and PVR displayed little, if any, change in expression. Ligand up-regulation resulted in improved lysis by polyclonal NK cells armed with the corresponding activating receptors. These results provide the first evidence for concerted induction of cell death by cell-autonomous and extrinsic (immune) mechanisms. Alarming the immune system much below the cell damage threshold may have evolved as a sensitive readout of neoplastic transformation and oxidative stress. Cocktails of vitamin analogues at slightly supra-physiological dosages may find application as mild complements of melanoma treatment, and in chemoprevention.

  10. Proton MRS detects Metabolic Changes in Hormone Sensitive and Resistant Human Prostate Cancer Model CWR22 and CWR22r

    Science.gov (United States)

    Le, H. Carl; Lupu, Mihaela; Kotedia, Khushali; Rosen, Neal; Solit, David; Koutcher, Jason A.

    2010-01-01

    17-Allylamino, 17-Demethoxygeldanamycin (17-AAG), an effective inhibitor of the heat shock protein hsp90, preferentially inhibiting tumor hsp90 compared to hsp90 from normal cells (1), has shown promising results against several cancers, including hormone resistant prostate cancer. Levels of several oncogenic proteins critical to tumor growth and progression, such as AR (androgen receptor) and HER2/neu, were reduced 4 hours post 17-AAG treatment. Post treatment metabolic changes have also been observed in several tumor cell lines. In this study total choline (t-cho) distributions in hormone sensitive CWR22 and hormone resistant CWR22r prostate cancer xenograft tumors in mice were measured before, 4 hours and 48 hours after a single bolus 17-AAG treatment at 100 mg/kg using proton MRS. Our results show that tumor t-cho levels declined 4 hours after the treatment for CWR22 (P = 0.001) and 48 hours post treatment for CWR22r (P=0.003). Metabolic changes, in particular of t-cho intensity detected by 1H MRSI, are consistent with the observed immunohistochemistry changes, tumor growth inhibition for CWR22r (P=0.01 at 14 days post treatment) and a constant PSA level versus increasing PSA for control CWR22 (P=0.01). Metabolic changes in t-cho by proton MRSI can be used as an early biomarker of response for advanced stage prostate cancer in targeted therapy such as 17-AAG. PMID:19780165

  11. Sensitivity of populations of bats (Mammalia: Chiroptera) in relation to human development in northern Paraná, southern Brazil.

    Science.gov (United States)

    Reis, N R; Gallo, P H; Peracchi, A L; Lima, L P; Fregonezi, M N

    2012-08-01

    Most natural forests have been converted for human use, restricting biological life to small forest fragments. Many animals, including some species of bats are disappearing and the list of these species grows every day. It seems that the destruction of the habitat is one of its major causes. This study aimed to analyze how this community of bats was made up in environments with different sizes and quality of habitat. Data from studies conducted in the region of Londrina, Parana, Brazil, from 1982 to 2000 were used. Originally, this area was covered by a semi deciduous forest, especially Aspidosperma polyneuron (Apocynaceae), Ficus insipida (Moraceae), Euterpe edulis (Arecaceae), Croton floribundus (Euforbiaceae), and currently, only small remnants of the original vegetation still exist. The results showed a decline in the number of species caught in smaller areas compared to the largest remnant. In about 18 years of sampling, 42 species of bats were found in the region, representing 67% of the species that occur in Paraná and 24.4% in Brazil. There were two species of Noctilionidae; 21 of Phyllostoma; 11 Vespertilionidae and eight Molossidae. Eight of these were captured only in the largest fragment, Mata dos Godoy State Park (680 ha). Ten species had a low capture rate in the smaller areas with less than three individuals. Of the total sampled, 14 species were found in human buildings, and were able to tolerate modified environments, foraging and even using them as shelter. As the size of the forest area increases, there is a greater variety of ecological opportunities and their physical conditions become more stable, i.e., conditions favorable for growth and survival of a greater number of species. Forest fragmentation limits and creates subpopulations, preserving only long-lived K-strategist animals for some time, where the supporting capacity of the environment is a limiting factor. The reduction of habitats, species and genetic diversity resulting from human

  12. Sensitivity of populations of bats (Mammalia: Chiroptera in relation to human development in northern Paraná, southern Brazil

    Directory of Open Access Journals (Sweden)

    NR. Reis

    Full Text Available Most natural forests have been converted for human use, restricting biological life to small forest fragments. Many animals, including some species of bats are disappearing and the list of these species grows every day. It seems that the destruction of the habitat is one of its major causes. This study aimed to analyze how this community of bats was made up in environments with different sizes and quality of habitat. Data from studies conducted in the region of Londrina, Parana, Brazil, from 1982 to 2000 were used. Originally, this area was covered by a semi deciduous forest, especially Aspidosperma polyneuron (Apocynaceae, Ficus insipida (Moraceae, Euterpe edulis (Arecaceae, Croton floribundus (Euforbiaceae, and currently, only small remnants of the original vegetation still exist. The results showed a decline in the number of species caught in smaller areas compared to the largest remnant. In about 18 years of sampling, 42 species of bats were found in the region, representing 67% of the species that occur in Paraná and 24.4% in Brazil. There were two species of Noctilionidae; 21 of Phyllostoma; 11 Vespertilionidae and eight Molossidae. Eight of these were captured only in the largest fragment, Mata dos Godoy State Park (680 ha. Ten species had a low capture rate in the smaller areas with less than three individuals. Of the total sampled, 14 species were found in human buildings, and were able to tolerate modified environments, foraging and even using them as shelter. As the size of the forest area increases, there is a greater variety of ecological opportunities and their physical conditions become more stable, i.e., conditions favorable for growth and survival of a greater number of species. Forest fragmentation limits and creates subpopulations, preserving only long-lived K-strategist animals for some time, where the supporting capacity of the environment is a limiting factor. The reduction of habitats, species and genetic diversity

  13. Glycogen synthase kinase-3 inhibition sensitizes human induced pluripotent stem cells to thiol-containing antioxidants induced apoptosis.

    Science.gov (United States)

    Tu, Chengyi; Xu, Robert; Koleti, Meghana; Zoldan, Janet

    2017-08-01

    Inhibition of glycogen synthase kinase 3 (GSK3) is an extensively used strategy to activate Wnt pathway for pluripotent stem cell (PSC) differentiation. However, the effects of such inhibition on PSCs, besides upregulating the Wnt pathway, have rarely been investigated despite that GSK3 is broadly involved in other cellular activities such as insulin signaling and cell growth/survival regulation. Here we describe a previously unknown synergistic effect between GSK3 inhibition (e.g., Chir99021 and LY2090314) and various normally non-toxic thiol-containing antioxidants (e.g., N-acetylcysteine, NAC) on the induction of apoptosis in human induced pluripotent stem cells (iPSCs). Neither Chir99021 nor the antioxidants individually induced significant apoptosis, whereas their combined treatment resulted in rapid and extensive apoptosis, with substantial caspase 3 activity observed within 3h and over 90% decrease in cell viability after 24h. We confirmed the generality of this phenomenon with multiple independent iPSCs lines, various thiol-based antioxidants and distinct GSK3 inhibitors. Mechanistically, we demonstrated that rapamycin treatment could substantially reduce cell death, suggesting the critical role of mammalian target of rapamycin (mTOR). Akt dysregulation was also found to partially contribute to cell apoptosis but was not the primary cause. Further, this coordinated proapoptotic effect was not detected in mouse ESCs but was present in another human cells line: a breast cancer cell line (MDA-MB-231). Given the wide use of GSK3 inhibition in biomedical research: from iPSC differentiation to cancer intervention and the treatment of neuronal diseases, researchers can potentially take advantage of or avoid this synergistic effect for improved experimental or clinical outcome. Copyright © 2017. Published by Elsevier B.V.

  14. Impact of anemia prevention by recombinant human erythropoietin on the sensitivity of xenografted glioblastomas to fractionated irradiation

    International Nuclear Information System (INIS)

    Stueben, G.; Poettgen, C.; Knuehmann, K.; Sack, H.; Stuschke, M.; Thews, O.; Vaupel, P.

    2003-01-01

    Background: Pronounced oxygen deficiency in tumors which might be caused by a diminished oxygen transport capacity of the blood (e.g., in anemia) reduces the efficacy of ionizing radiation. The aim of this study was to analyze whether anemia prevention by recombinant human erythropoietin (rHuEPO) affects the radiosensitivity of human glioblastoma xenografts during fractionated irradiation. Material and Methods: Anemia was induced by total body irradiation (TBI, 2 x 4 Gy) of mice prior to tumor implantation into the subcutis of the hind leg. In one experimental group, the development of anemia was prevented by rHuEPO (750 U/kg s.c.) given three times weekly starting 10 days prior to TBI. 13 days after tumor implantation (tumor volume approx. 40 mm 3 ), fractionated irradiation (4 x 7 Gy, one daily fraction) of the glioblastomas was performed resulting in a growth delay with subsequent regrowth of the tumors. Results: Compared to nonanemic control animals (hemoglobin concentration cHb = 14.7 g/dl), the growth delay in anemic mice (cHb = 9.9 g/dl) was significantly shorter (49 ± 5 days vs. 79 ± 4 days to reach four times the initial tumor volume) upon fractionated radiation. The prevention of anemia by rHuEPO treatment (cHb = 13.3 g/dl) resulted in a significantly prolonged growth delay (61 ± 5 days) compared to the anemia group, even though the growth inhibition found in control animals was not completely achieved. Conclusions: These data indicate that moderate anemia significantly reduces the efficacy of radiotherapy. Prevention of anemia with rHuEPO partially restores the radiosensitivity of xenografted glioblastomas to fractionated irradiation. (orig.)

  15. Vulnerability or Sensitivity to the Environment? Methodological Issues, Trends, and Recommendations in Gene–Environment Interactions Research in Human Behavior

    Directory of Open Access Journals (Sweden)

    Caroline Leighton

    2017-06-01

    Full Text Available Research on the potential role of gene–environment interactions (GxE in explaining vulnerability to psychopathology in humans has witnessed a shift from a diathesis-stress perspective to differential susceptibility approaches. This paper critically reviews methodological issues and trends in this body of research. Databases were screened for studies of GxE in the prediction of personality traits, behavior, and mental health disorders in humans published between January 2002 and January 2015. In total, 315 papers were included. Results showed that 34 candidate genes have been included in GxE studies. Independent of the type of environment studied (early or recent life events, positive or negative environments, about 67–83% of studies have reported significant GxE interactions, which is consistent with a social susceptibility model. The percentage of positive results does not seem to differ depending on the gene studied, although publication bias might be involved. However, the number of positive findings differs depending on the population studied (i.e., young adults vs. older adults. Methodological considerations limit the ability to draw strong conclusions, particularly as almost 90% (n = 283/315 of published papers are based on samples from North America and Europe, and about 70% of published studies (219/315 are based on samples that were also used in other reports. At the same time, there are clear indications of methodological improvements over time, as is shown by a significant increase in longitudinal and experimental studies as well as in improved minimum genotyping. Recommendations for future research, such as minimum quality assessment of genes and environmental factors, specifying theoretical models guiding the study, and taking into account of cultural, ethnic, and lifetime perspectives, are formulated.

  16. Relationship of dopamine type 2 receptor binding potential with fasting neuroendocrine hormones and insulin sensitivity in human obesity.

    Science.gov (United States)

    Dunn, Julia P; Kessler, Robert M; Feurer, Irene D; Volkow, Nora D; Patterson, Bruce W; Ansari, Mohammad S; Li, Rui; Marks-Shulman, Pamela; Abumrad, Naji N

    2012-05-01

    Midbrain dopamine (DA) neurons, which are involved with reward and motivation, are modulated by hormones that regulate food intake (insulin, leptin, and acyl ghrelin [AG]). We hypothesized that these hormones are associated with deficits in DA signaling in obesity. We assessed the relationships between fasting levels of insulin and leptin, and AG, BMI, and insulin sensitivity index (S(I)) with the availability of central DA type 2 receptor (D2R). We measured D2R availability using positron emission tomography and [(18)F]fallypride (radioligand that competes with endogenous DA) in lean (n = 8) and obese (n = 14) females. Fasting hormones were collected prior to scanning and S(I) was determined by modified oral glucose tolerance test. Parametric image analyses revealed associations between each metabolic measure and D2R. The most extensive findings were negative associations of AG with clusters involving the striatum and inferior temporal cortices. Regional regression analyses also found extensive negative relationships between AG and D2R in the caudate, putamen, ventral striatum (VS), amygdala, and temporal lobes. S(I) was negatively associated with D2R in the VS, while insulin was not. In the caudate, BMI and leptin were positively associated with D2R availability. The direction of associations of leptin and AG with D2R availability are consistent with their opposite effects on DA levels (decreasing and increasing, respectively). After adjusting for BMI, AG maintained a significant relationship in the VS. We hypothesize that the increased D2R availability in obese subjects reflects relatively reduced DA levels competing with the radioligand. Our findings provide evidence for an association between the neuroendocrine hormones and DA brain signaling in obese females.

  17. Proton MRS detects metabolic changes in hormone sensitive and resistant human prostate cancer models CWR22 and CWR22r.

    Science.gov (United States)

    Le, H Carl; Lupu, Mihaela; Kotedia, Khushali; Rosen, Neal; Solit, David; Koutcher, Jason A

    2009-11-01

    17-Allylamino, 17-demethoxygeldanamycin (17-AAG), an effective inhibitor of the heat shock protein hsp90, preferentially inhibiting tumor hsp90 compared to hsp90 from normal cells, has shown promising results against several cancers, including hormone-resistant prostate cancer. Levels of several oncogenic proteins critical to tumor growth and progression, such as androgen receptor and HER2/neu, were reduced 4 h post 17-allylamino, 17-demethoxygeldanamycin treatment. Posttreatment metabolic changes have also been observed in several tumor cell lines. In this study, total choline distributions in hormone sensitive CWR22 and hormone resistant CWR22r prostate cancer xenograft tumors in mice were measured before and at 4 h and 48 h after a single-bolus 17-allylamino, 17-demethoxygeldanamycin treatment at 100 mg/kg, using proton MR spectroscopy. Our results show that tumor total choline levels declined 4 h after the treatment for CWR22 (P = 0.001) and 48 h post treatment for CWR22r (P = 0.003). Metabolic changes, in particular of total choline intensity detected by proton magnetic resonance spectroscopic imaging (MRSI), are consistent with the observed immunohistochemistry changes, tumor growth inhibition for CWR22r (P = 0.01 at 14 days post treatment), and a constant prostate specific antigen level versus increasing prostate specific antigen for control CWR22 (P = 0.01). Metabolic changes in total choline by proton MRSI can be used as an early biomarker of response for advanced-stage prostate cancer in targeted therapy such as 17-allylamino, 17-demethoxygeldanamycin. (c) 2009 Wiley-Liss, Inc.

  18. Highly Sensitive Micellar Enhanced Spectrofluorimetric Method for Determination of Mirtazapine in Tablets and Human Urine: Application to In Vitro Drug Release and Content Uniformity Test

    Directory of Open Access Journals (Sweden)

    Hany W. Darwish

    2016-01-01

    Full Text Available A highly sensitive and simple micelle enhanced spectrofluorimetric method was developed for assaying mirtazapine (MRZ in REMERON® tablets and spiked human urine directly without the need of derivatizing agent. The basis of the current procedure is the examination of the relative fluorescence intensity (RFI of MRZ in sodium lauryl sulphate (SLS micellar medium. The RFI of MRZ in water was enhanced markedly on addition of SLS. The RFI was measured at 403 nm after excitation at 320 nm. The fluorescence-concentration relationship was linear over the range 1–500 ng/mL, with lower detection limit of 0.399 ng/mL. The proposed method was successfully applied to the determination of MRZ in dosage form and spiked human urine. Recovery percentages of MRZ utilizing the current method were 99.05±1.83, 98.37±1.96, and 100.41±2.61% for pure powder, pharmaceutical dosage form, and spiked human urine, respectively. The application of the proposed method was extended to test content uniformity and the in vitro drug release of REMERON tablets, according to USP guidelines.

  19. High-sensitivity simultaneous liquid chromatography–tandem mass spectrometry assay of ethinyl estradiol and levonorgestrel in human plasma

    Directory of Open Access Journals (Sweden)

    Abhishek Gandhi

    2015-10-01

    Full Text Available A sensitive and simultaneous liquid chromatography–tandem mass spectrometry method was developed and validated for quantification of ethinyl estradiol and levonorgestrel. The analytes were extracted with methyl-tert-butyl ether: n-hexane (50:50, v/v solvent mixture, followed by dansyl derivatization. The chromatographic separation was performed on a Kinetex C18 (50 mm×4.6 mm, 2.6 µm column with a mobile phase of 0.1% (v/v formic acid in water and acetonitrile in gradient composition. The mass transitions were monitored in electrospray positive ionization mode. The assay exhibited a linear range of 0.100–20.0 ng/mL for levonorgestrel and 4.00–500 pg/mL for ethinyl estradiol in human plasma. A run time of 9.0 min for each sample made it possible to analyze a throughput of more than 100 samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic and bioequivalence studies. Keywords: Ethinyl estradiol, Levonorgestrel, LC–MS/MS, Human plasma, Derivatization

  20. Alterations of DNA content in human endometrial stromal cells transfected with a temperature-sensitive SV40: tetraploidization and physiological consequences.

    Science.gov (United States)

    Rinehart, C A; Mayben, J P; Butler, T D; Haskill, J S; Kaufman, D G

    1992-01-01

    The normal genomic stability of human cells is reversed during neoplastic transformation. The SV40 large T antigen alters the DNA content in human endometrial stromal cells in a manner that relates to neoplastic progression. Human endometrial stromal cells were transfected with a plasmid containing the A209 temperature-sensitive mutant of SV40 (tsSV40), which is also defective in the viral origin of replication. Ninety-seven clonal transfectants from seven different primary cell strains were isolated. Initial analysis revealed that 20% of the clonal populations (19/97) had an apparent diploid DNA content, 35% (34/97) had an apparent tetraploid DNA content, and the remainder were mixed populations of diploid and tetraploid cells. No aneuploid populations were observed. Diploid tsSV40 transformed cells always give rise to a population of cells with a tetraploid DNA content when continuously cultured at the permissive temperature. The doubling of DNA content can be vastly accelerated by the sudden reintroduction of large T antigen activity following a shift from non-permissive to permissive temperature. Tetraploid tsSV40 transfected cells have a lower capacity for anchorage-independent growth and earlier entry into 'crisis' than diploid cells. These results indicate that during the pre-crisis, extended lifespan phase of growth, the SV40 large T antigen causes a doubling of DNA content. This apparent doubling of DNA content does not confer growth advantage during the extended lifespan that precedes 'crisis'.