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Sample records for receptor-mediated clonogenic radioresistance

  1. The phosphatidylinositol-3 kinase pathway is not essential for insulin-like growth factor I receptor-mediated clonogenic radioresistance

    International Nuclear Information System (INIS)

    Yu, Dong; Watanabe, Hiroshi; Shibuya, Hitoshi; Miura, Masahiko

    2002-01-01

    The insulin-like growth factor I receptor (IGF-IR) is known to induce clonogenic radioresistance in cells following ionizing irradiation. To explore the downstream signaling pathways, we focused on the phosphatidylinositol-3 kinase (PI3-K) pathway, which is thought to be the primary cell survival signal originating from the receptor. For this purpose, R- cells deficient in the endogenous IGF-IR were used as a recipient of the human IGF-IR with or without mutations at potential PI3-K activation sites: NPXY 950 and Y 1316 XXM. Mutats with double mutation at Y950/Y1316 exhibited not abrogated, but reduced activation of insulin receptor substance-1 (IRS-1), PI3-K, and Akt upon IGF-I stimulation. However, the mutants had the same clonogenic radioresistance as cells with wild type (WT) receptors. Neither wortmannin nor LY294002, specific inhibitors of PI3-K, affected the radioresistance of cells with WT receptors at concentrations specific for PI3-K. Collectively, these results indicate that the PI3-K pathway is not essential for IGF-IR-mediated clonogenic radioresistance. (author)

  2. Clonogenic assay: adherent cells.

    Science.gov (United States)

    Rafehi, Haloom; Orlowski, Christian; Georgiadis, George T; Ververis, Katherine; El-Osta, Assam; Karagiannis, Tom C

    2011-03-13

    The clonogenic (or colony forming) assay has been established for more than 50 years; the original paper describing the technique was published in 1956. Apart from documenting the method, the initial landmark study generated the first radiation-dose response curve for X-ray irradiated mammalian (HeLa) cells in culture. Basically, the clonogenic assay enables an assessment of the differences in reproductive viability (capacity of cells to produce progeny; i.e. a single cell to form a colony of 50 or more cells) between control untreated cells and cells that have undergone various treatments such as exposure to ionising radiation, various chemical compounds (e.g. cytotoxic agents) or in other cases genetic manipulation. The assay has become the most widely accepted technique in radiation biology and has been widely used for evaluating the radiation sensitivity of different cell lines. Further, the clonogenic assay is commonly used for monitoring the efficacy of radiation modifying compounds and for determining the effects of cytotoxic agents and other anti-cancer therapeutics on colony forming ability, in different cell lines. A typical clonogenic survival experiment using adherent cells lines involves three distinct components, 1) treatment of the cell monolayer in tissue culture flasks, 2) preparation of single cell suspensions and plating an appropriate number of cells in petri dishes and 3) fixing and staining colonies following a relevant incubation period, which could range from 1-3 weeks, depending on the cell line. Here we demonstrate the general procedure for performing the clonogenic assay with adherent cell lines with the use of an immortalized human keratinocyte cell line (FEP-1811). Also, our aims are to describe common features of clonogenic assays including calculation of the plating efficiency and survival fractions after exposure of cells to radiation, and to exemplify modification of radiation-response with the use of a natural antioxidant

  3. Characterization of radioresistant variant from U251 human glioblastoma cell line and the role of antioxdant enzymes in its radioresistancy

    International Nuclear Information System (INIS)

    Lee, Hyung Chahn; Park, In Chul; Park, Myung Jin; Woo, Sang Hyeok; Rhee, Chang Hum; Hong, Seok-II

    2004-01-01

    To investigate the radioresistant mechanism in glioblastoma multiforme(GBM), we isolated the radioresistant clone (RRC) from U251 human glioblastoma cell line by exposing to repeated fractions of 3 Gy γ-radiation for six months. RRC had higher radioresistance than the parent cell line as measured by clonogenic survival assay. FACS analysis showed that RRC had a delayed G2 arrest after radiation. Antioxidant enzymes, such as SOD, catalase, glutathione peroxidase (GPX), glutathione reductase (GR), were activated up to 5 folds in RRC after radiation. Erk 1/2 activation was higher in RRC than in the parent cell. Therefore, radioresistancy in RRC might be due to the delayed cell cycle, the coordinated high activation of antioxidant enzyme rather than a single enzyme alone,and higher activation of Erk 1/2

  4. Radioresistance and hypoxic cells

    International Nuclear Information System (INIS)

    Ando, Koichi

    1989-01-01

    Current progress to explore further understanding of tumor hypoxia was reviewed. At subcellular level, hypoxia induces specific proteins, inhibits DNA synthesis as well as initiation of DNA replicon. Radioresistant characteristics of hypoxic cells is questioned in condition where irradiated cells were kept hypoxia during colony formation. Chronically hypoxic cells recovered from the inner layer of V79 multicellular spheroids are more sensitive to radiation than those from the oxic, outer layer. A novel sandwich culture method, which enables to reoxygenate chronic hypoxia, implies that chronically hypoxic cells are less sensitive to radiation after reoxygenation than oxic cells. For in vivo tumor, two types of tumor hypoxia are reported: diffusion-limited, chronic hypoxia and perfusion-limited, acute hypoxia. Evidence supporting the existence of perfusion-limited hypoxia is provided by an elegant method using vital staining and cell sorter. Data of our own laboratory also implies 2 types of tumor hypoxia; fractional hypoxia and incomplete hypoxia. Fractional hypoxia corresponds to a radioresistant tail on a biphasic tumor cell survival curves while tumors with incomplete hypoxia demonstrate only single component with radioresistant characteristics, instead. (author)

  5. Radioresistant canine hematopoietic cells

    International Nuclear Information System (INIS)

    Kawakami, T.G.; Shimizu, J.; Rosenblatt, L.S.; Goldman, M.

    1987-01-01

    Survival of dogs that are continuously exposed to a moderate dose-rate of gamma radiation (10 cGy/day) is dependent on the age of the dog at the time of exposure. Most dogs exposed postpartum to gamma radiation suffered from suppressed hematopoiesis and died of aplasia. On the other hand, none of the in utero-exposed dogs suffered from suppressed hematopoiesis and most became long-term survivors, tolerating 10-fold greater total dose, but dying of myeloproliferative disease (MPD). Using acute gamma irradiation of hematopoietic cells and colony forming unit cell assay (CFU), they observed that a canine hematopoietic cell line established from a myeloid leukemic dog that was a long-term survivor of continuous irradiation was approximately 4-fold more radioresistant than a hematopoietic cell line established from a dog with nonradiation-induced myeloid leukemia or hematopoietic cells from normal canine bone marrow. In utero dogs that are long-term survivors of continuous irradiation have radioresistant hematopoietic cells, and radioresistance that is a constitutive property of the cells

  6. Radioresistant microorganisms and food irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Ito, H [Japan Atomic Energy Research Inst., Takasaki, Gunma. Takasaki Radiation Chemistry Research Establishment

    1976-01-01

    This paper deals with Micrococcus radiodurans, Arthrobacter radiotolerance, etc., which were isolated and discovered as radioresistant microorganisms. As for the explanation of the mechanism of radioresistance of these microorganisms, the consideration that these organisms have marked repair power of the damaged DNA and have many opportunity to repair the damaged DNA because of their long fission term were cited. The relationship between the radioresistance of microorganisms and food irradiation was also mentioned.

  7. Selection of radioresistant tumor cells and presence of ALDH1 activity in vitro

    International Nuclear Information System (INIS)

    Mihatsch, Julia; Toulany, Mahmoud; Bareiss, Petra M.; Grimm, Sabrina; Lengerke, Claudia; Kehlbach, Rainer; Rodemann, H. Peter

    2011-01-01

    Background: Tumor resistance to radiotherapy has been hypothesized to be mediated by a tumor subpopulation, called cancer stem cells (CSCs). Based on the proposed function of CSCs in radioresistance, we explored the cancer stem cell properties of cells selected for radioresistance phenotype. Materials and methods: A549 and SK-BR-3 cells were radioselected with four single doses of 4 or 3 Gy in intervals of 10-12 days and used for colony formation assay and γ-H2AX foci formation assay. Expression of putative stem cell markers, i.e. Sox2, Oct4, ALDH1, and CD133 were analyzed using Western blotting. A549 and SK-BR-3 cells sorted based on their ALDH1 activity were analyzed in clonogenic survival assays. Results: Radioselected A549 and SK-BR-3 cells (A549-R, SK-BR-3-R) showed increased radioresistance and A549-R cells presented enhanced repair of DNA-double strand breaks. PI3K inhibition significantly reduced radioresistance of A549-R cells. Cell line specific differences in the expression of the putative CSC markers Sox2 and Oct4 were observed when parental and radioselected cells were compared but could not be directly correlated to the radioresistant phenotype. However, enzyme activity of the putative stem cell marker ALDH1 showed a correlation to radioresistance. Conclusions: Subpopulations of pooled radioresistant colonies, selected by various radiation exposures were analyzed for the presence of putative stem cell markers. Although the pattern of Sox2, Oct4, and CD133 expression was not generally associated with radioresistance, presence of ALDH1 seems to be indicative for subpopulations with increased radioresistance.

  8. On the analysis of clonogenic survival data: Statistical alternatives to the linear-quadratic model

    International Nuclear Information System (INIS)

    Unkel, Steffen; Belka, Claus; Lauber, Kirsten

    2016-01-01

    The most frequently used method to quantitatively describe the response to ionizing irradiation in terms of clonogenic survival is the linear-quadratic (LQ) model. In the LQ model, the logarithm of the surviving fraction is regressed linearly on the radiation dose by means of a second-degree polynomial. The ratio of the estimated parameters for the linear and quadratic term, respectively, represents the dose at which both terms have the same weight in the abrogation of clonogenic survival. This ratio is known as the α/β ratio. However, there are plausible scenarios in which the α/β ratio fails to sufficiently reflect differences between dose-response curves, for example when curves with similar α/β ratio but different overall steepness are being compared. In such situations, the interpretation of the LQ model is severely limited. Colony formation assays were performed in order to measure the clonogenic survival of nine human pancreatic cancer cell lines and immortalized human pancreatic ductal epithelial cells upon irradiation at 0-10 Gy. The resulting dataset was subjected to LQ regression and non-linear log-logistic regression. Dimensionality reduction of the data was performed by cluster analysis and principal component analysis. Both the LQ model and the non-linear log-logistic regression model resulted in accurate approximations of the observed dose-response relationships in the dataset of clonogenic survival. However, in contrast to the LQ model the non-linear regression model allowed the discrimination of curves with different overall steepness but similar α/β ratio and revealed an improved goodness-of-fit. Additionally, the estimated parameters in the non-linear model exhibit a more direct interpretation than the α/β ratio. Dimensionality reduction of clonogenic survival data by means of cluster analysis was shown to be a useful tool for classifying radioresistant and sensitive cell lines. More quantitatively, principal component analysis allowed

  9. Radiogenic neoplasia in thyroid and mammary clonogens

    International Nuclear Information System (INIS)

    Clifton, K.H.

    1993-01-01

    The induction of cancer by ionizing radiation is a matter of great practical importance to the nuclear industry, to national defense, to radiological medicine and to the general public. It is increasingly apparent that carcinogenesis is one of the leading dose-limiting effects of radiation exposure (Co90). Quantitative information at the cellular level is essential to an understanding of the mechanisms of radiogenic neoplastic initiation and the stages of promotion and progression to overt neoplasia. We have developed two experimental models, the rat thyroid and rat mammary clonogen transplant systems, for the quantitative study of radiation carcinogenesis at the cellular level in vivo (C185). The most important steps taken or completed during the current grant year include: (a) demonstration of the high age-dependent radiosensitivity of prepubertal rat mammary clonogens to radiogenic damage which may influence their susceptibility to neoplastic initiation, and (b) demonstration of the feasibility of using a molecular test for clonogenicity in which Simple Sequence Repeats in the DNA serve as identifying signals of the genotypic origin of the cells. We have also (c) set up a large carcinogenesis experiment to test the effect of close intercellular contact in thyroid glands in situ on promotion-progression of radiogenically initiated clonogens, (d) achieved considerable further concentration of thyroid clonogens, and (e) begun to explore whether thyroid cells can be induced to give rise to three dimensional multicellular structures in culture in reconstituted basement membrane. These are discussed in this report

  10. Radiogenic neoplasia in thyroid and mammary clonogens

    International Nuclear Information System (INIS)

    Clifton, K.H.

    1992-01-01

    We have developed rat thyroid and mammary clonogen transplantation systems for the study of radiogenic cancer induction at the target cell level in vivo. The epithelial cell populations of both glands contain small subpopulations of cells which are capable of giving rise to monoclonal glandular structures when transplanted and stimulated with appropriate hormones. Previous results indicated that these clonogens are the precursor cells of radiogenic cancer, and that initiation, is common event at the clonegenic cell level. Detailed information on the physiologic control of clonogen proliferation, differentiation, and total numbers is thus essential to an understanding of the carcinogenic process. We report here studies on investigations on the relationships between grafted thyroid cell number and the rapidity and degree of reestablishment of the thyroid-hypothalamus-pituitary feedback axis in thyroidectomized rats maintained on a normal diet or an iodine deficient diet; studies of the persistence of, and the differentiation potential and functional characteristics of, the TSH-(thyrotropin-) responsive sub- population of clonogens during goitrogenesis, the plateau-phase of goiter growth, and goiter involution; studies of changes in the size of the clonogen sub-population during goitrogenesis, goiter involution and the response to goitrogen rechallenge; and a large carcinogenesis experiment on the nature of the grafted thyroid cell number-dependent suppression of promotion/progression to neoplasia in grafts of radiation-initiated thyroid cells. Data from these studies will be used in the design of future carcinogenesis experiments on neoplastic initiation by high and low LET radiations and on cell interactions during the neoplastic process

  11. Radiogenic neoplasia in thyroid and mammary clonogens

    International Nuclear Information System (INIS)

    Clifton, K.H.

    1991-01-01

    We have developed rat thyroid and mammary clonogen transplantation systems for the study of radiogenic cancer induction at the target cell level in vivo. The epithelial cell populations of both glands contain small subpopulations of cells which are capable of giving rise to monoclonal glandular structures when transplanted and stimulated with appropriate hormones. During the end of the last grant year and the first half of the current grant year, we have completed analyses and summarized for publication: investigations on the relationship between grafted thyroid cell number and the rapidity and degree of reestablishment of the thyroid-hypothalamicpituitary axis in thyroidectomized rats maintained on a normal diet or an iodine deficient diet; studies of the persistence of, and the differentiation potential and functional characteristics of, the TSH- (thyrotropin-) responsive sub-population of clonogens during goitrogenesis, the plateau-phase of goiter growth, and goiter involution; studies of changes in the size of the clonogen sub-population during goitrogenesis, goiter involution and the response to goitrogen rechallenge; and the results of the large carcinogenesis experiment on the nature of the grafted thyroid cell number-dependent suppression of promotion/progression to neoplasia in grafts of radiation-initiated thyroid cells. We are testing new techniques for the culture, cytofluorescent analysis and characterization mammary epithelial cells and of clonogens in a parallel project, and plan to apply similar technology to the thyroid epithelial cells and clonogen population. Data from these studies will be used in the design of future carcinogenesis experiments on neoplastic initiation by high and low LET radiations and on cells interactions during the neoplastic process

  12. Canonical autophagy does not contribute to cellular radioresistance

    International Nuclear Information System (INIS)

    Schaaf, Marco B.E.; Jutten, Barry; Keulers, Tom G.; Savelkouls, Kim G.M.; Peeters, Hanneke J.M.; Beucken, Twan van den; Schooten, Frederik-Jan van; Godschalk, Roger W.; Vooijs, Marc; Rouschop, Kasper M.A.

    2015-01-01

    Background: (Pre)clinical studies indicate that autophagy inhibition increases response to anti-cancer therapies. Although promising, due to contradicting reports, it remains unclear if radiation therapy changes autophagy activity and if autophagy inhibition changes the cellular intrinsic radiosensitivity. Discrepancies may result from different assays and models through off-target effects and influencing other signaling routes. In this study, we directly compared the effects of genetic and pharmacological inhibition of autophagy after irradiation in human cancer cell lines. Materials and methods: Changes in autophagy activity after ionizing radiation (IR) were assessed by flux analysis in eight cell lines. Clonogenic survival, DNA damage (COMET-assay) and H2AX phosphorylation were assessed after chloroquine or 3-methyladenine pretreatment and after ATG7 or LC3b knockdown. Results: IR failed to induce autophagy and chloroquine failed to change intrinsic radiosensitivity of cells. Interestingly, 3-methyladenine and ATG7- or LC3b-deficiency sensitized cancer cells to irradiation. Surprisingly, the radiosensitizing effect of 3-methyladenine was also observed in ATG7 and LC3b deficient cells and was associated with attenuated γ-H2AX formation and DNA damage repair. Conclusion: Our data demonstrate that the anti-tumor effects of chloroquine are independent of changes in intrinsic radioresistance. Furthermore, ATG7 and LC3b support radioresistance independent of canonical autophagy that involves lysosomal degradation

  13. Transient elevation of glycolysis confers radio-resistance by facilitating DNA repair in cells

    International Nuclear Information System (INIS)

    Bhatt, Anant Narayan; Chauhan, Ankit; Khanna, Suchit; Rai, Yogesh; Singh, Saurabh; Soni, Ravi; Kalra, Namita; Dwarakanath, Bilikere S

    2015-01-01

    Cancer cells exhibit increased glycolysis for ATP production (the Warburg effect) and macromolecular biosynthesis; it is also linked with therapeutic resistance that is generally associated with compromised respiratory metabolism. Molecular mechanisms underlying radio-resistance linked to elevated glycolysis remain incompletely understood. We stimulated glycolysis using mitochondrial respiratory modifiers (MRMs viz. di-nitro phenol, DNP; Photosan-3, PS3; Methylene blue, MB) in established human cell lines (HEK293, BMG-1 and OCT-1). Glucose utilization and lactate production, levels of glucose transporters and glycolytic enzymes were investigated as indices of glycolysis. Clonogenic survival, DNA repair and cytogenetic damage were studied as parameters of radiation response. MRMs induced the glycolysis by enhancing the levels of two important regulators of glucose metabolism GLUT-1 and HK-II and resulted in 2 fold increase in glucose consumption and lactate production. This increase in glycolysis resulted in resistance against radiation-induced cell death (clonogenic survival) in different cell lines at an absorbed dose of 5 Gy. Inhibition of glucose uptake and glycolysis (using fasentin, 2-deoxy-D-glucose and 3-bromopyruvate) in DNP treated cells failed to increase the clonogenic survival of irradiated cells, suggesting that radio-resistance linked to inhibition of mitochondrial respiration is glycolysis dependent. Elevated glycolysis also facilitated rejoining of radiation-induced DNA strand breaks by activating both non-homologous end joining (NHEJ) and homologous recombination (HR) pathways of DNA double strand break repair leading to a reduction in radiation-induced cytogenetic damage (micronuclei formation) in these cells. These findings suggest that enhanced glycolysis generally observed in cancer cells may be responsible for the radio-resistance, partly by enhancing the repair of DNA damage

  14. Characterization of the radioresistance in the radioresistant bacterium deinococcus radiodurans

    International Nuclear Information System (INIS)

    Kong Xiangrong; Du Zeji

    1999-01-01

    The radioresistance of wild type Deinococcus radiodurans KD8301 and the factors affecting the radioresistance were investigated. KH3111 which was a DNA repair mutant of KD8301 (Zeji Du, 1998) was used to be compared with KD8301. Deinococcus radiodurans was discovered by Anderson et al (1956) in X-ray sterilized canned meat that was found to have undergone spoilage. this bacterium and other species of this genus share extreme resistance to ionizing radiation and other agents that damage DNA. Wild type KD8301 and its sensitive mutant KH3111 were irradiated with 60 Co γ-ray at the dose range 0.5 ∼ 10 kGy. Dose-survival fraction curves were made and the radio resistances were determined by LD 99 . The relative contents of DNA in cells were measured by Fluorescence Spectrophotometry (Freedman and Bruce, 1971). The results indicated that wild type KD8301 possesses more radioresistant than its mutant KH3111, LD99 were 9.5 kGy and 2.4 kGy respectively. KD8301 grown at exponential phase showed a decreased resistance to radiation, and the LD99 was 5.1 kGy. No differences of DNA/protein in cells were found between the exponential phase and the stationary phase. The results could be concluded that wild type KD8301 possesses remarkable radioresistance, but this ability was decreased or disappeared after mutation (in KH3111). None DNA relative content other than the growth stages were determinant factors of radioresistance in Deinococcus radiodurans. This results were different from other report (Dickie N et al, 1990). The cellular mechanisms might be the deference's of the bacterium cell morphology between the exponential phase and the stationary phase. Recently, the mutation site of KH3111 which was mutated chemically from wild type KD8301 was identified (Zeji Du, 1998). One base pair changed in the novel gene pprA which was isolated from KD8301 genomic DNA. This point mutation was confirmed to be responsible for the sensitivity of KH3111 to γ-ray and other DNA

  15. A radiobiological comparison of human tumor soft-agar clonogenic assays.

    Science.gov (United States)

    West, C M; Sutherland, R M

    1986-06-15

    Radiation survival curves have been generated for 3 human tumor cell lines as a means of comparing and evaluating the validity of human tumor soft-agar clonogenic assays. The assays investigated were the Hamburger-Salmon, Courtenay-Mills, Courtenay-Mills plus additions, soft agar (no additions), and soft agar plus additions. The additions were formulated to supplement the media used in soft agar assays of primary ovarian and cervical carcinoma specimens. Supplementing the media with additions led to a 2- to 3-fold increase in PE of CaSki cells but had no effect on the PEs of ME180 and OWI cells. Radiation survival curves were similar in all assays for CaSki and OWI but differed for ME180 cells. For ME180 cells, the Courtenay-Mills and soft agar assays plus additions produced the most radioresistant curves (Do = 2.2 Gy); the cells were more responsive when assayed by the Hamburger-Salmon method (Do = 1.5 Gy), and the soft agar and Courtenay-Mills assays gave the most radiosensitive curves (Do = 1.2 Gy). These results demonstrate that the PE of human tumor cell lines may be increased with no effect on radiation survival; radiation survival may be altered without changes in PE and neither may be altered by applying modifications and supplements to existing clonogenic assays.

  16. Heterogeneity of clonogenic cells in acute myeloblastic leukemia.

    OpenAIRE

    Sabbath, K D; Ball, E D; Larcom, P; Davis, R B; Griffin, J D

    1985-01-01

    The expression of differentiation-associated surface antigens by the clonogenic leukemic cells from 20 patients with acute myeloblastic leukemia (AML) was studied with a panel of seven cytotoxic monoclonal antibodies (anti-Ia, -MY9, -PM-81, -AML-2-23, -Mol, -Mo2, and -MY3). The surface antigen phenotypes of the clonogenic cells were compared with the phenotypes of the whole leukemic cell population, and with the phenotypes of normal hematopoietic progenitor cells. In each case the clonogenic ...

  17. Targeting the AKT/GSK3β/Cyclin D1/Cdk4 Survival Signaling Pathway for Eradication of Tumor Radioresistance Acquired by Fractionated Radiotherapy

    International Nuclear Information System (INIS)

    Shimura, Tsutomu; Kakuda, Satoshi; Ochiai, Yasushi; Kuwahara, Yoshikazu; Takai, Yoshihiro; Fukumoto, Manabu

    2011-01-01

    Purpose: Radioresistance is a major cause of treatment failure of radiotherapy (RT) in human cancer. We have recently revealed that acquired radioresistance of tumor cells induced by fractionated radiation is attributable to cyclin D1 overexpression as a consequence of the downregulation of GSK3β-dependent cyclin D1 proteolysis mediated by a constitutively activated serine-threonine kinase, AKT. This prompted us to hypothesize that targeting the AKT/GSK3β/cyclin D1 pathway may improve fractionated RT by suppressing acquired radioresistance of tumor cells. Methods and Materials: Two human tumor cell lines with acquired radioresistance were exposed to X-rays after incubation with either an AKT inhibitor, AKT/PKB signaling inhibitor-2 (API-2), or a Cdk4 inhibitor (Cdk4-I). Cells were then subjected to immunoblotting, clonogenic survival assay, cell growth analysis, and cell death analysis with TUNEL and annexin V staining. In vivo radiosensitivity was assessed by growth of human tumors xenografted into nude mice. Results: Treatment with API-2 resulted in downregulation of cyclin D1 expression in cells with acquired radioresistance. Cellular radioresistance disappeared completely both in vitro and in vivo with accompanying apoptosis when treated with API-2. Furthermore, inhibition of cyclin D1/Cdk4 by Cdk4-I was sufficient for abolishing radioresistance. Treatment with either API-2 or Cdk4-I was also effective in suppressing resistance to cis-platinum (II)-diamine-dichloride in the cells with acquired radioresistance. Interestingly, the radiosensitizing effect of API-2 was canceled by overexpression of cyclin D1 whereas Cdk4-I was still able to sensitize cells with cyclin D1 overexpression. Conclusion: Cyclin D1/Cdk4 is a critical target of the AKT survival signaling pathway responsible for tumor radioresistance. Targeting the AKT/GSK3β/cyclin D1/Cdk4 pathway would provide a novel approach to improve fractionated RT and would have an impact on tumor eradication in

  18. In vitro stemness characterization of radio-resistant clones isolated from a medulloblastoma cell line ONS-76

    International Nuclear Information System (INIS)

    Sun, Lue; Suzuki, Kenshi; Gerelchuluun, Ariungerel; Hong, Zhengshan; Moritake, Takashi; Zenkoh, Junko; Tsuboi, Koji; Zheng, Yun-Wen; Taniguchi, Hideki

    2013-01-01

    One-third of patients with medulloblastoma die due to recurrence after various treatments including radiotherapy. Although it has been postulated that cancer stem-like cells are radio-resistant and play an important role in tumor recurrence, the 'stemness' of medulloblastoma cells surviving irradiation has not yet been elucidated. Using a medulloblastoma cell line ONS-76, cells that survived gamma irradiation were investigated on their 'stemness' in vitro. From 10 500 cells, 20 radio-resistant clones were selected after gamma ray irradiation (5 Gy x two fractions) using the replica micro-well technique. These 20 resistant clones were screened for CD133 positivity by flow cytometry followed by side population assay, tumor sphere formation assay and clonogenic survival assay. Results revealed CD133 fractions were significantly elevated in three clones, which also exhibited significantly increased levels of tumor sphere formation ability and side population fraction. Clonogenic survival assay demonstrated that their radio-resistance was significantly higher than the parental ONS-76. This may support the hypothesis that a small number of cancer stem-like cells (CSCs) are the main culprits in local recurrence after radiotherapy, and disruption of the resistance mechanism of these CSCs is a critical future issue in improving the outcome of patients with medulloblastoma. (author)

  19. Gene expression profile associated with radioresistance and malignancy in melanoma

    International Nuclear Information System (INIS)

    Ibañez, I.L.; Molinari, B.; Notcovich, C.; García, F.M.; Bracalente, C.; Zuccato, C.F.; Durán, H.

    2015-01-01

    The incidence of melanoma has substantially increased over the last decades. Melanomas respond poorly to treatments and no effective therapy exists to inhibit its metastatic spread. The aim of this study was to evaluate the association between radioresistance of melanoma cells and malignancy. A melanoma model developed in our laboratory from A375 human amelanotic melanoma cells was used. It consists in two catalase-overexpressing cell lines with the same genetic background, but with different phenotypes: A375-A7, melanotic and non-invasive and A375-G10, amelanotic and metastatic; and A375-PCDNA3 (transfected with empty plasmid) as control. Radiosensitivity was determined by clonogenic assay after irradiating these cells with a “1”3”7 Cs gamma source. Survival curves were fitted to the linear-quadratic model and surviving fraction at 2 Gy (SF2) was calculated. Results showed that A375-G10 cells were significantly more radioresistant than both A375-A7 and control cells, demonstrated by SF2 and α parameter of survival curves: SF2=0.32±0.03, 0.43±0.16 and 0.89±0.05 and α=0.45±0.05, 0.20±0.05 and 0 for A375-PCDNA3, A375-A7 and A375-G10 respectively. Bioinformatic analysis of whole genome expression microarrays data (Affymetrix) from these cells was performed. A priori defined gene sets associated with cell cycle, apoptosis and MAPK signaling pathway were collected from KEGG (Kyoto Encyclopedia of Genes and Genomes) to evaluate significant differences in gene set expression between cells by GSEA (Gene Set Enrichment Analysis). A375-G10 showed significant decrease in the expression of genes related to DNA damage response (ATM, TP53BP1 and MRE11A) compared to A375-A7 and controls. Moreover, A375-G10 exhibited down-regulated gene sets that are involved in DNA repair, checkpoint and negative regulation of cell cycle and apoptosis. In conclusion, A375-G10 gene expression profile could be involved in radioresistance mechanisms of these cells. Thus, this expression

  20. Stress and radioresistance (genetic aspects)

    International Nuclear Information System (INIS)

    Lyubaschevsky, N.M.; Grigorkina, E.B.

    1995-01-01

    Inbred mice and randomly bred stock were exposed to long-term adrenaline and hypothermia stress effects for unification of their functional status, followed by acute total gamma irradiation by equal-effect doses (LD 50/30 ). The significant reduction of lethality distribution in inbred mice after combined stress and radiation effects was observed. The spread of lethality in random bred stock was increased as compared to the control group. The post-irradiation mortality in experimental animals differed only slightly from the values in the control groups. Conclusions are drawn about the principal meaning of genotype in radiosensitivity, about the role of an organism's functional status in determination of death date and about the possible epigenetic structuralisation of linear mice by radioresistance. (Author)

  1. Radiations, human cancerization and radioresistance

    International Nuclear Information System (INIS)

    Vuillaume, M.

    1991-01-01

    Active oxygen species have been implicated in inflammatory diseases, cellular transformation and carcinogenesis. Experiments studying their direct (for instance, by ionizing radiation) or indirect influence (for instance, by the decrease(s) in activity of cellular defence enzymes), in vivo, and/or, in cell cultures (fibroblasts, keratinocytes, hepatocytes,...) have been positive. To the contrary, reactions or products, which decrease level(s) of free radicals/H 2 O 2 (i) directly, (ii) indirectly by an increase of SOD, catalase, and peroxydase activites suppress the above-described phenomena. This is the case for the domain number 2 (that contains copper, MW 53KDa) of the Scorpion's blood pigment (hemocyanin), which possesses SOD-, catalase-, and peroxydase-like properties, resistant to, at least, 4000 Gy, and thus may explain the especially high radioresistance of scorpions. Enzymatic decrease(s) and the domain number 2 of hemocyanin are under current investigation in several laboratories [fr

  2. E-Cadherin loss associated with EMT promotes radioresistance in human tumor cells

    International Nuclear Information System (INIS)

    Theys, Jan; Jutten, Barry; Habets, Roger; Paesmans, Kim; Groot, Arjan J.; Lambin, Philippe; Wouters, Brad G.; Lammering, Guido; Vooijs, Marc

    2011-01-01

    Background and purpose: Hypoxia is a hallmark of solid cancers and associated with metastases and treatment failure. During tumor progression epithelial cells often acquire mesenchymal features, a phenomenon known as epithelial-to-mesenchymal transition (EMT). Intratumoral hypoxia has been linked to EMT induction. We hypothesized that signals from the tumor microenvironment such as growth factors and tumor oxygenation collaborate to promote EMT and thereby contribute to radioresistance. Materials and methods: Gene expression changes under hypoxia were analyzed using microarray and validated by qRT-PCR. Conversion of epithelial phenotype upon hypoxic exposure, TGFβ addition or oncogene activation was investigated by Western blot and immunofluorescence. Cell survival following ionizing radiation was assayed using clonogenic survival. Results: Upon hypoxia, TGFβ addition or EGFRvIII expression, MCF7, A549 and NMuMG epithelial cells acquired a spindle shape and lost cell-cell contacts. Expression of epithelial markers such as E-cadherin decreased, whereas mesenchymal markers such as vimentin and N-cadherin increased. Combining hypoxia with TGFβ or EGFRvIII expression, lead to more rapid and pronounced EMT-like phenotype. Interestingly, E-cadherin expression and the mesenchymal appearance were reversible upon reoxygenation. Mesenchymal conversion and E-cadherin loss were associated with radioresistance. Conclusions: Our findings describe a mechanism by which the tumor microenvironment may contribute to tumor radioresistance via E-cadherin loss and EMT.

  3. Energetic heavy ions overcome tumor radioresistance caused by overexpression of Bcl-2

    International Nuclear Information System (INIS)

    Hamada, Nobuyuki; Hara, Takamitsu; Omura-Minamisawa, Motoko; Funayama, Tomoo; Sakashita, Tetsuya; Sora, Sakura; Yokota, Yuichiro; Nakano, Takashi

    2008-01-01

    Background and purpose: Overexpression of Bcl-2 is frequent in human cancers and has been associated with radioresistance. Here we investigated the potential impact of heavy ions on Bcl-2 overexpressing tumors. Materials and methods: Bcl-2 cells (Bcl-2 overexpressing HeLa cells) and Neo cells (neomycin resistant gene-expressing HeLa cells) exposed to γ-rays or heavy ions were assessed for the clonogenic survival, apoptosis and cell cycle distribution. Results: Whereas Bcl-2 cells were more resistant to γ-rays (0.2 keV/μm) and helium ions (16.2 keV/μm) than Neo cells, heavy ions (76.3-1610 keV/μm) yielded similar survival regardless of Bcl-2 overexpression. Carbon ions (108 keV/μm) decreased the difference in the apoptotic incidence between Bcl-2 and Neo cells, and prolonged G 2 /M arrest that occurred more extensively in Bcl-2 cells than in Neo cells. Conclusions: High-LET heavy ions overcome tumor radioresistance caused by Bcl-2 overexpression, which may be explained at least in part by the enhanced apoptotic response and prolonged G 2 /M arrest. Thus, heavy-ion therapy may be a promising modality for Bcl-2 overexpressing radioresistant tumors

  4. Cytotoxicity and radiosensitization effect of TRA-8 on radioresistant human larynx squamous carcinoma cells.

    Science.gov (United States)

    Wu, F; Hu, Y; Long, J; Zhou, Y J; Zhong, Y H; Liao, Z K; Liu, S Q; Zhou, F X; Zhou, Y F; Xie, C H

    2009-02-01

    TRAIL induces apoptosis in a variety of tumorigenic and transformed cell lines, but not in many normal cells. Recent studies have demonstrated that death receptor 5 (DR5), one of the two death receptors bound by TRAIL, showed expression in most malignantly transformed cells. This study evaluated effects of a monoclonal antibody (TRA-8) to human death receptor 5, combined with ionizing radiation, on radioresistant human larynx squamous carcinoma cell line (Hep-2R). Cells were treated with TRA-8 alone or in combination with radiation, cell viability inhibition was measured by MTT assay, and the induction of apoptosis was determined by Annexin V staining. Radionsensitivity of Hep-2R cells treated with TRA-8 were investigated with long-term clonogenic assays. Regulation of DR5 expression in cells after radiation was analyzed by indirect immunofluorescence using murine TRA-8 in combination with flow cytometry. The results suggested that TRA-8 enhanced radionsensitivity of Hep-2R cells, and that TRA-8 regulated Hep-2R cell cycle arrest at G2/M phase. Irradiation up-regulated the expression of DR5, and when combined with TRA-8 yielded optimal survival benefit. Therefore, TRA-8 can be used in combination with irradiation in radioresistant human larynx squamous carcinoma cells. Monoclonal antibodies such as TRA-8 may play an important role in the development of an effective treatment strategy for patients with radioresistant cancers.

  5. Dexamethasone-induced radioresistance occurring independent of human papilloma virus gene expression in cervical carcinoma cells

    International Nuclear Information System (INIS)

    Rutz, H.P.; Mariotta, M.; Mirimanoff, R.O.; Knebel Doeberitz, M. von

    1998-01-01

    The aim of this study was to investigate the role of HPV 18 E6 and E7 gene products with respect to radiosensitivity of two cervical carcinoma cell lines. The two cervical carcinoma lines C4-1 and SW 756 were used in which treatment with dexamethasone allows to modulate expression levels of HPV 18 E6 and E7 genes: Upregulation in C4-1, down-regulation in SW 756. Effects of treatment with dexamethasone on plating efficiency and radiosensitivity were assessed using a clonogenic assay. Treatment with dexamethasone increased plating efficiency of the C4-1 cells, but did not affect plating efficiency of SW 756 cells. Treatment with dexamethasone induced enhanced radioresistance in both cell lines. Thus, in C4-1 cells the observed changes in radioresistance correlate to the enhancement in expression of HPV 18 genes E6/E7, whereas in SW 756, a reduced expression correlates negatively with the enhanced radioresistance. (orig./MG) [de

  6. Radioresistant cell strain of human fibrosarcoma cells obtained after long-term exposure to X-rays

    International Nuclear Information System (INIS)

    Wei, K.; Kodym, R.

    1998-01-01

    A radioresistant cell strain from human fibrosarcoma HT1080 has been obtained after prolonged exposure to x-rays for 7 months (2 Gy per day, 5 days per week). This new strain, HT1080R, differs from HT1080 in a significantly increased ability of clonogenical survival, with coefficient α decreasing from 0.161 to 0.123 Gy -1 and coefficient β decreasing from 0.0950 to 0.0565 Gy -2 . Furthermore, the radioresistance of HT1080R proved to be stable in long-term passaged cultures as well as in frozen samples. Differences between the two cell lines are also observed in the G-banded karyotype; the new cell line shows monosomy of chromosome 17 and loss of 5p + and 11q + present in the parental cells. These data suggest that the radioresistance may have been caused by radiation-induced cell mutation and that the resistant cells may have been selected by repeated irradiations. In order to characterize this new strain, the ability of the cells to rejoin DNA double-strand breaks, the cell cycle distribution and the amount of apoptosis after irradiation have been estimated; however, no differences are observed between these two cell strains. Although the mechanism of the elevated radioresistance remains unknown, this pair of cell strains can provide a new model system for further investigations with regard to the mechanisms of cellular radioresistance. The results also show that any type of irradiation similar to the schedules used in radiotherapy can lead to the formation and selection of more radioresistant cell clones in vitro, a phenomenon with possible implications for radiotherapy. (orig.)

  7. Establishment and cell cycle distribution pattern of a radioresistant subline from human lung cancer D6 cell line

    International Nuclear Information System (INIS)

    Wei Qichun; Zheng Shu

    2003-01-01

    Objective: To establish a radioresistant cell subline from a human D6 lung cancer cell line and investigate the mechanism of radioresistance. Methods: D6 human NSCLC cells were exposed to X-rays generated by a linear accelerator(650 cGy per fraction). After a total exposure dose of 5200 cGy, a monoclone was obtained. The radiosensitivity and cell cycle distribution of this clone, together with its parent D6 cells, were measured by clonogenic assay and flow cytometry. Results: The new clone, namely D 6 -R subline, had a higher D 0 (D 0 =2.08 Gy) and a broader initial shoulder(Dq=1.64 Gy, N=2.20) than those of the parent D6 cell line (D 0 =1.84 Gy, Dq=0.34 Gy, N=1.20), being 1.65-fold increase in radioresistance as regards to the SF 2 . The D6-R subline also showed higher percentage of cells in S phase(53.4% vs 37.8%), but lower percentages in G 1 (44.1% vs 57.2%) and G 2 /M(2.5% vs 5%) phases. Conclusion: The new subline D6-R is more radioresistant as compare to its parent D6 cell line, and has a different cell cycle distribution

  8. Induction of aryl hydrocarbon receptor-mediated and estrogen receptor-mediated activities, and modulation of cell proliferation by dinaphthofurans

    Czech Academy of Sciences Publication Activity Database

    Vondráček, Jan; Chramostová, Kateřina; Plíšková, M.; Bláha, L.; Brack, W.; Kozubík, Alois; Machala, M.

    2004-01-01

    Roč. 23, č. 9 (2004), s. 2214-2220 ISSN 0730-7268 R&D Projects: GA ČR GA525/03/1527 Institutional research plan: CEZ:AV0Z5004920 Keywords : aryl hydrocarbon receptor-mediated activity * estrogenicity * intercellular communication inhibition Subject RIV: BO - Biophysics Impact factor: 2.121, year: 2004

  9. Bmi-1 confers adaptive radioresistance to KYSE-150R esophageal carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Guanyu [Department of General Surgery, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Liu, Luying [Department of Radiotherapy, Zhejiang Cancer Hospital, Hangzhou (China); Sharma, Sherven [David Geffen School of Medicine at UCLA, and the Department of Veterans Affairs, Los Angeles, CA (United States); Liu, Hai; Yang, Weifang; Sun, Xiaonan [Department of Radiotherapy, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou (China); Dong, Qinghua, E-mail: dongqinghua@zju.edu.cn [Biomedical Research Center, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou (China)

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer Adaptive radioresistant KYSE-150R cells expressed high level of Bmi-1. Black-Right-Pointing-Pointer Bmi-1 depletion sensitized KYSE-150R cells to RT. Black-Right-Pointing-Pointer Bmi-1 depletion increased the generation of ROS in KYSE-150R cells exposed to radiation. Black-Right-Pointing-Pointer Bmi-1 depletion impaired DNA repair capacities in KYSE-150R cells exposed to radiation. -- Abstract: Radiotherapy (RT) is a major modality of cancer treatment. However, tumors often acquire radioresistance, which causes RT to fail. The exact mechanisms by which tumor cells subjected to fractionated irradiation (FIR) develop an adaptive radioresistance are largely unknown. Using the radioresistant KYSE-150R esophageal squamous cell carcinoma (ESCC) model, which was derived from KYSE-150 parental cells using FIR, the role of Bmi-1 in mediating the radioadaptive response of ESCC cells to RT was investigated. The results showed that the level of Bmi-1 expression was significantly higher in KYSE-150R cells than in the KYSE-150 parental cells. Bmi-1 depletion sensitized the KYSE-150R cells to RT mainly through the induction of apoptosis, partly through the induction of senescence. A clonogenic cell survival assay showed that Bmi-1 depletion significantly decreased the radiation survival fraction in KYSE-150R cells. Furthermore, Bmi-1 depletion increased the generation of reactive oxygen species (ROS) and the expression of oxidase genes (Lpo, Noxo1 and Alox15) in KYSE-150R cells exposed to irradiation. DNA repair capacities assessed by {gamma}-H2AX foci formation were also impaired in the Bmi-1 down-regulated KYSE-150R cells. These results suggest that Bmi-1 plays an important role in tumor radioadaptive resistance under FIR and may be a potent molecular target for enhancing the efficacy of fractionated RT.

  10. Heterogeneity of clonogenic cells in acute myeloblastic leukemia.

    Science.gov (United States)

    Sabbath, K D; Ball, E D; Larcom, P; Davis, R B; Griffin, J D

    1985-02-01

    The expression of differentiation-associated surface antigens by the clonogenic leukemic cells from 20 patients with acute myeloblastic leukemia (AML) was studied with a panel of seven cytotoxic monoclonal antibodies (anti-Ia, -MY9, -PM-81, -AML-2-23, -Mol, -Mo2, and -MY3). The surface antigen phenotypes of the clonogenic cells were compared with the phenotypes of the whole leukemic cell population, and with the phenotypes of normal hematopoietic progenitor cells. In each case the clonogenic leukemic cells were found within a distinct subpopulation that was less "differentiated" than the total cell population. Clonogenic leukemic cells from different patients could be divided into three phenotype groups. In the first group (7 of 20 cases), the clonogenic cells expressed surface antigens characteristic of the normal multipotent colony-forming cell (Ia, MY9). These cases tended to have "undifferentiated" (FAB M1) morphology, and the total cell population generally lacked expression of "late" monocyte antigens such as MY3 and Mo2. A second group (seven cases) of clonogenic cells expressed surface antigens characteristic of an "early" (day 14) colony-forming unit granulocyte-monocyte (CFU-GM), and a third group (six cases) was characteristic of a "late" (day 7) CFU-GM. The cases in these latter two groups tended to have myelomonocytic (FAB M4) morphology and to express monocyte surface antigens. These results suggest that the clonogenic cells are a distinct subpopulation in all cases of AML, and may be derived from normal hematopoietic progenitor cells at multiple points in the differentiation pathway. The results further support the possibility that selected monoclonal antibodies have the potential to purge leukemic clonogenic cells from bone marrow in some AML patients without eliminating critical normal progenitor cells.

  11. An improved method for staining cell colonies in clonogenic assays

    OpenAIRE

    Guda, Kishore; Natale, Leanna; Markowitz, Sanford D.

    2007-01-01

    Clonogenic assay is a widely used experimental approach to test for the effects of drugs/genes on the growth and proliferative characteristics of cells in vitro. Accurate quantitation of treatment effects in clonogeneic assays depends on the ability to visualize and count cell colonies precisely. We report a novel method (referred as ETeB) for staining cell colonies grown on plastic and specially coated substrates like collagen. Using colon cancer cell lines grown on plastic and collagen, we ...

  12. Radioresistance in Central Asian tortoise

    Energy Technology Data Exchange (ETDEWEB)

    Turdyev, A.A.; Bogdanova-Berezovskaya, I.G.; Dvornikova, L.I.

    Season-related studies were conducted on the phenomenal radioresistance of the Central Asian tortoise (Testudo horsfieldii), in order to obtain greater understanding of the factors that determine this physiological uniqueness. Test animals were exposed to gamma radiation from a Co-60 source with dosages ranging from 12.9 to 25.8 Cl/kg in the spring, summer, fall, and winter. Using Pearson's ..gamma..-distribution coefficient calculations of the mean lethal dose for the season yielded 16.33 Cl/kg for winter, 10.94 Cl/kg for spring, 7.1 Cl/kg for summer, and 19.71 Cl/kg for fall. The animals were thus seen to be most susceptible to radiation damage in the spring and summer. Correlation with histological and hematological studies showed that animals irradiated in spring and summer succumbed as a result of damage to the GI and the hematopoietic systems, whereas animals that died as a result of irradiation during the fall and winter season did so because of depletion of hepatic glycogen stores. 5 references, 1 figure.

  13. Receptor-mediated gene delivery using chemically modified chitosan

    International Nuclear Information System (INIS)

    Kim, T H; Jiang, H L; Nah, J W; Cho, M H; Akaike, T; Cho, C S

    2007-01-01

    Chitosan has been investigated as a non-viral vector because it has several advantages such as biocompatibility, biodegradability and low toxicity with high cationic potential. However, the low specificity and low transfection efficiency of chitosan need to be solved prior to clinical application. In this paper, we focused on the galactose or mannose ligand modification of chitosan for enhancement of cell specificity and transfection efficiency via receptor-mediated endocytosis in vitro and in vivo

  14. Association of anti-apoptotic Mcl-1L isoform expression with radioresistance of oral squamous carcinoma cells

    International Nuclear Information System (INIS)

    Palve, Vinayak C; Teni, Tanuja R

    2012-01-01

    Oral cancer is a common cancer and a major health problem in the Indian subcontinent. At our laboratory Mcl-1, an anti-apoptotic member of the Bcl-2 family has been demonstrated to be overexpressed in oral cancers and to predict outcome in oral cancer patients treated with definitive radiotherapy. To study the role of Mcl-1 isoforms in radiation response of oral squamous carcinoma cells (OSCC), we investigated in the present study, the association of Mcl-1 isoform expression with radiosensitivity of OSCC, using siRNA strategy. The time course expression of Mcl-1 splice variants (Mcl-1L, Mcl-1S & Mcl-1ES) was studied by RT-PCR, western blotting & immunofluorescence, post-irradiation in oral cell lines [immortalized FBM (radiosensitive) and tongue cancer AW8507 & AW13516 (radioresistant)]of relatively differing radiosensitivities. The effect of Mcl-1L knockdown alone or in combination with ionizing radiation (IR) on cell proliferation, apoptosis & clonogenic survival, was investigated in AW8507 & AW13516 cells. Further the expression of Mcl-1L protein was assessed in radioresistant sublines generated by fractionated ionizing radiation (FIR). Three to six fold higher expression of anti-apoptotic Mcl-1L versus pro-apoptotic Mcl-1S was observed at mRNA & protein levels in all cell lines, post-irradiation. Sustained high levels of Mcl-1L, downregulation of pro-apoptotic Bax & Bak and a significant (P < 0.05) reduction in apoptosis was observed in the more radioresistant AW8507, AW13516 versus FBM cells, post-IR. The ratios of anti to pro-apoptotic proteins were high in AW8507 as compared to FBM. Treatment with Mcl-1L siRNA alone or in combination with IR significantly (P < 0.01) increased apoptosis viz. 17.3% (IR), 25.3% (siRNA) and 46.3% (IR plus siRNA) and upregulated pro-apoptotic Bax levels in AW8507 cells. Combination of siRNA & IR treatment significantly (P < 0.05) reduced cell proliferation and clonogenic survival of radioresistant AW8507 & AW13516 cells

  15. Age and radiation sensitivity of rat mammary clonogenic cells

    International Nuclear Information System (INIS)

    Shimada, Yoshiya; Yasukawa-Barnes, J.; Kim, R.Y.; Gould, M.N.; Clifton, K.H.

    1994-01-01

    The relative risk of breast cancer is very high among women who were exposed to ionizing radiation during or before puberty. In the current studies, the surviving fractions of clonogenic mammary cells of groups of virgin rats were estimated after single exposures to 137 Cs γ rays at intervals from 1 to 12 weeks after birth. The radiosensitivity of clonogens from prepubertal rats was high and changed with the onset of puberty at between 4 and 6 weeks of age. By this time, the increase in the size of the clonogenic cell subpopulation was slowing and differentiation of terminal mammary end buds and alveolar structures was occurring. Analysis of the relationship of clonogen survival and radiation dose according to the α/β model showed that the exponential αD term predominated at the second and fourth weeks of age. By the eighth week of age, the βD 2 term had come to predominate and the survival curve had a pronounced initial convex shoulder. Further experiments are required to determine whether there is an association between the high sensitivity of the prepubertal and pubertal mammary clonogens to radiation killing and a high susceptibility to radiogenic initiation of cancer. 24 refs., 4 figs., 1 tab

  16. Thermoresistance in radioresistant strains of 'Drosophila nebulosa'

    International Nuclear Information System (INIS)

    Kratz, F.L.

    1977-01-01

    The detection of thermoresistance in radioresistant strains of 'D. nebulosa' is described, as well as some conclusions on the genetic nature of these differences are presented. The strains used in this experiment were MF 204, from 'Morro de Ferro', in Pocos de Caldas (MG) (one of the biggest radioactive anomalies in the world) whose radioresistance is due to its additive genetic components (Kratz, 1973 and 1975); 85(87) R, an induced radioresistant strain; and MF K a control 'pooled' strain obtained near 'Morro do Ferro'. Survival tests, 72 hours after temperature shocks, performed in the interval of 36 0 C to 39 0 C showed a decreasing gradient of thermoresistance with the following regression coefficients: MF 204 b= - 5,4; 85(87)R b= - 7,2 and MF K b= - 7,9. Bifactorial analysis (strains and sexes) performed at 38 0 C and 39 0 C confirmed differences among strains (P [pt

  17. Role of lipids in bacterial radioresistance

    International Nuclear Information System (INIS)

    Abushady, M.R.; Fawkia, M.E.; Tawfik, Z.S.

    1992-01-01

    The radioresistance of three bacterial isolates was determined. S. aureus was the most sensitive one (D 1 0 value 0.14 KGy), B. coagulans was moderate resistant (D 1 0 value 3.3 KGy) and the most resistant one was B.megaterium (D 1 0 value 3.7 KGy). Total lipids and lipid patterns of these bacteria were determined and the role of lipids in radioresistance was investigated. Least amount of total lipids was detected in the most sensitive organism (S. aureus). The increase in the bacterial content of total lipids was concomitant with high degrees of radioresistance. The most resistant organism (B. megaterium was characterized by high content of methyl esters of fatty acids, phosphatidylcholine and phosphatidylethanolamine, followed by appreciable amounts in the moderate resistant (B. coagulans) and the least amounts were detected in the most sensitive organism (S.aureus).6 fig., 3 tab

  18. The problem: Tumor radioresistance in clinical radiotherapy

    International Nuclear Information System (INIS)

    Peters, L.J.; Withers, H.R.; Thames, H.D.; Fletcher, G.H.; M.D. Anderson Hospital and Tumor Inst., Houston, TX)

    1983-01-01

    Tumor radioresistance in clinical radiotherapy implies failure to achieve local-regional disease control with radiation doses producing an acceptable degree of morbidity. Such radioresistance may be due to many different causes (biological and technical) which are reviewed in terms of possible remedial actions. Dose-response relationships for human cancers suggest that, in many sites, tumors are heterogeneous with respect to their cure-limiting characteristics. The case is developed that unless the predominant cure-limiting factor can be predicted, little benefit may be seen in trials of new treatment strategies using heterogeneous tumor populations. The fundamental problem of clinical radioresistance is therefore perceived as the inability to identify predictively its cause in the individual patient

  19. [Unique properties of highly radioresistant bacteria].

    Science.gov (United States)

    Romanovskaia, V A; Rokitko, P V; Malashenko, Iu R

    2000-01-01

    In connection with the Chernobyl Nuclear Power Plant (ChNPP) accident and the negative ecological after-effects for biota in this zone the interest has arisen to radioresistant bacteria, as to the most dynamic model of the given ecosystem, and to mechanisms which provide resistance of bacteria to ionizing radiation. The analysis of published data has shown that the radioresistant bacteria are not interrelated taxonomically and phylogenetically. The extreme radioresistant bacteria are represented by the Deinococcus species, which form a group phylogenetically close to the line Thermus-Meiothermus. Other radioresistant bacteria are the representatives of the genera Rubrobacter, Methylobacterium, Kocuria, Bacillus and some archebacteria. Data on natural habitats, of radioresistant bacteria are not numerous. In a number of cases it is difficult to distinguish their natural habitats, as they were isolated from the samples which were previously exposed to X-ray or gamma-irradiation, or from the ecosystems with the naturally raised radioactivity. To understand the strategy of survival of radioresistant bacteria, we briefly reviewed the mechanism of action of various species of radiation on cells and macromolecules; physiological signs of the cell damage caused by radiation; mechanisms eliminating (repairing) these damages. More details on mechanisms of the DNA repair in D. radiodurans are described. The extreme resistance of D. radiodurans to the DNA damaging factors is defined by 1) repair mechanisms which fundamentally differ from those in other procaryotes; 2) ability to increase the efficiency of a standard set of the DNA repairing proteins. Literary and own data on the effect of radiation on survival of various groups of bacteria in natural ecosystems are summarized. The ecological consequences of the ChNPP accident for soil bacteria in this region were estimated. The reduction of the number of soil bacteria and recession of microbial diversity under the effect of

  20. Radioresistance of immunized animals in internal irradiation

    International Nuclear Information System (INIS)

    Kal'nitskij, S.A.; Ponomareva, T.V.; Shubik, V.M.

    1981-01-01

    The influence of an immunization with bacterial vaccines and antimeasles-gamma-globulin on the radioresistance of raceless white mice was studied. In the vaccinated animals a higher survival rate and duration of life, a better general condition and a better curve of weight and stronger physical stamina were stated compared to the merely irradiated mice. The higher radioresistance is ascribed to the stimulation of cellular and humoral factors of the unspecific protection against infection, to the repair of the lymphoid tissue of the immunized animals and to the decrease in autosensibilization. (author)

  1. Radioresistance and radiosensitivity in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Reguly, M.L.

    1983-01-01

    Studying the mechanisms controlling radioresistant in Drosophila the sensibility of four strains of Drosophila melanogaster to sex-linked recessive lethal mutations induced by 5kR Cobalt-60 gamma radiation and 0,006 M EMS or 0,25% of caffeine was determined. (M.A.C.) [pt

  2. Radioresistance and radiosensitivity in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Reguly, M.L.; Marques, E.K.

    1987-01-01

    The mechanisms of radioresistance in Drosophila are studied. The mutagenic effects of 5KR of 60 Cobalt gamma radiation and of 0,006M dose of ethyl methanesulfonate (EMS) on four D. Melanogaster strains (RC 1 , CO 3 , BUE and LEN) are investigated. (M.A.C.) [pt

  3. The liver taxis of receptor mediated lactosaminated human growth hormone

    International Nuclear Information System (INIS)

    Chen Zelian; Shi Lin; Li Tongling; Pang Qijie; He Juying; Guan Changtian

    2002-01-01

    Radiography imaging is used to assess liver taxis mechanism of anti-dwarfism drug lactosaminated human growth hormone (L-rhGH). Both L-rhGH and rhGH labelled with 131 I are used to study their biodistribution in animals (including rabbits, cocks and rats). The results show that L-rhGH is of specific hepatic targeting property, and the maximum hepatic concentration rate is 76.8%, which is two times of rhGH. Its hepatic binding is receptor mediated

  4. Nicotine enhances proliferation, migration, and radioresistance of human malignant glioma cells through EGFR activation

    International Nuclear Information System (INIS)

    Khalil, A.A.; Jameson, M.J.; Broaddus, W.C.; Lin, P.S.; Chung, T.D.

    2013-01-01

    It has been suggested that continued tobacco use during radiation therapy contributes to maintenance of neoplastic growth despite treatment with radiation. Nicotine is a cigarette component that is an established risk factor for many diseases, neoplastic and otherwise. The hypothesis of this work is that nicotine promotes the proliferation, migration, and radioresistance of human malignant glioma cells. The effect of nicotine on cellular proliferation, migration, signaling, and radiation sensitivity were evaluated for malignant glioma U87 and GBM12 cells by use of the AlamarBlue, scratch healing, and clonogenic survival assays. Signal transduction was assessed by immunoblotting for activated EGFR, extracellular regulated kinase (ERK), and AKT. At concentrations comparable with those found in chronic smokers, nicotine induced malignant glioma cell migration, growth, colony formation, and radioresistance. Nicotine increased phosphorylation of EGFR tyr992 , AKT ser473 , and ERK. These molecular effects were reduced by pharmacological inhibitors of EGFR, PI3K, and MEK. It was therefore concluded that nicotine stimulates the malignant behavior of glioma cells in vitro by activation of the EGFR and downstream AKT and ERK pathways. (author)

  5. Stronger Dopamine D1 Receptor-Mediated Neurotransmission in Dyskinesia.

    Science.gov (United States)

    Farré, Daniel; Muñoz, Ana; Moreno, Estefanía; Reyes-Resina, Irene; Canet-Pons, Júlia; Dopeso-Reyes, Iria G; Rico, Alberto J; Lluís, Carme; Mallol, Josefa; Navarro, Gemma; Canela, Enric I; Cortés, Antonio; Labandeira-García, José L; Casadó, Vicent; Lanciego, José L; Franco, Rafael

    2015-12-01

    Radioligand binding assays to rat striatal dopamine D1 receptors showed that brain lateralization of the dopaminergic system were not due to changes in expression but in agonist affinity. D1 receptor-mediated striatal imbalance resulted from a significantly higher agonist affinity in the left striatum. D1 receptors heteromerize with dopamine D3 receptors, which are considered therapeutic targets for dyskinesia in parkinsonian patients. Expression of both D3 and D1-D3 receptor heteromers were increased in samples from 6-hydroxy-dopamine-hemilesioned rats rendered dyskinetic by treatment with 3, 4-dihydroxyphenyl-L-alanine (L-DOPA). Similar findings were obtained using striatal samples from primates. Radioligand binding studies in the presence of a D3 agonist led in dyskinetic, but not in lesioned or L-DOPA-treated rats, to a higher dopamine sensitivity. Upon D3-receptor activation, the affinity of agonists for binding to the right striatal D1 receptor increased. Excess dopamine coming from L-DOPA medication likely activates D3 receptors thus making right and left striatal D1 receptors equally responsive to dopamine. These results show that dyskinesia occurs concurrently with a right/left striatal balance in D1 receptor-mediated neurotransmission.

  6. Radioresistance increase in polymers at high pressures

    International Nuclear Information System (INIS)

    Milinchuk, V.; Kirjukhin, V.; Klinshpont, E.

    1977-01-01

    The effect was studied of very high pressures ranging within 100 and 2,700 MPa on the radioresistance of polytetrafluoroethylene, polypropylene and polyethylene in gamma irradiation. For experiments industrial polymers in the shape of blocks, films and fibres were used. It is shown that in easily breakable polymers, such as polytetrafluoroethylene and polypropylene 1.3 to 2 times less free radicals are formed as a result of gamma irradiation and a pressure of 150 MPa than at normal pressure. The considerably reduced radiation-chemical formation of radicals and the destruction suppression by cross-linking in polymers is the evidence of the polymer radioresistance in irradiation at high pressures. (J.B.)

  7. Radioresistance and immunization effectiveness under internal irradiation

    International Nuclear Information System (INIS)

    Kal'nitskij, S.A.

    1978-01-01

    The effect of preliminary immunization on the radioresistance of mice to internal irradiation from incorporated 137 Cs or 90 Sr was studied, and it was found that a preliminary single immunization with bacterial vaccines had a favorable effect on the outcome of radiation injury. The present results suggested that vaccination had a very pronounced radioprotective effect and so may be used as a means of biologic protection from internal irradiation

  8. Ivastimul used to increase radioresistance of mice

    International Nuclear Information System (INIS)

    Rotkovska, D.; Vacek, A.; Bartonickova, A.

    1989-01-01

    A study was made of the effect of ivastimul (IS), an aqueous extract from unicellular Chlorella algae, on the radioresistance and some haemopoiesis parameters of mice exposed to 60 Co-γ-radiation. With median and absolutely lethal radiation doses, IS was shown to produce a pronounced protective effect displayed by the increased survival rate. With sublethal doses, IS elevated the postirradiation formation of endogenous colonies and restoration of bone marrow and spleen cellularity and spleen mass

  9. Receptor-mediated endocytosis of trichosanthin in choriocarcinoma cells

    International Nuclear Information System (INIS)

    Chan, W.Y.; Huang Hai; Tam, S.-C.

    2003-01-01

    Trichosanthin (TCS) is a ribosome inactivating protein (RIP). It is generally believed that its many biological activities act through inhibition of ribosomes resulting in a decrease in protein synthesis. It has been hypothesized that the rate of entry of TCS into cells to reach ribosomes is an important factor in determining its biological activity. To prove this hypothesis, we have mapped out and compared the intracellular routing of TCS in two cell lines, namely the choriocarcinoma JAR cell line, which is known to be highly sensitive to the toxic effects of TCS, and the hepatoma H35 cell line, to which TCS shows minimal toxicity. Results from laser scanning confocal microscopy indicated that fluorescein isothiocyanate labeled TCS quickly accumulated inside JAR cells within 4 h of incubation while only a low level of fluorescent signals was detected in H35 cells during the same period of time. When TCS was conjugated with gold particles (Au) and its intracellular locations were traced with a transmission electron microscope, it was found that most of TCS were bound to coated pits on the JAR cell surface and were rapidly internalized within an hour. By 4 h, TCS reached almost every cytoplasmic region including ribosomes, and the JAR cell began to degenerate. In H35 cells, however, the binding of TCS to coated pits was not observed, but instead, a small amount of TCS was found to penetrate the cell non-specifically by direct diffusion across the cell membrane. Our observations suggest that most of TCS enter JAR cells via a specific receptor mediated pathway, which allows a swift transport of TCS across the membrane and a rapid accumulation of intracellular TCS, while in H35 cells, TCS takes a slow and non-specific route. The receptor-mediated uptake together with the specific intracellular routing of TCS may partly account for the differential vulnerability of the choriocarcinoma cell line towards the toxicity of TCS

  10. DMPD: Modulation of Toll-interleukin 1 receptor mediated signaling. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15662540 Modulation of Toll-interleukin 1 receptor mediated signaling. Li X, Qin J.... J Mol Med. 2005 Apr;83(4):258-66. Epub 2005 Jan 21. (.png) (.svg) (.html) (.csml) Show Modulation of Toll-i...nterleukin 1 receptor mediated signaling. PubmedID 15662540 Title Modulation of Toll-interleukin 1 receptor

  11. An improved method for staining cell colonies in clonogenic assays.

    Science.gov (United States)

    Guda, Kishore; Natale, Leanna; Markowitz, Sanford D

    2007-06-01

    Clonogenic assay is a widely used experimental approach to test for the effects of drugs/genes on the growth and proliferative characteristics of cells in vitro. Accurate quantitation of treatment effects in clonogeneic assays depends on the ability to visualize and count cell colonies precisely. We report a novel method (referred as ETeB) for staining cell colonies grown on plastic and specially coated substrates like collagen. Using colon cancer cell lines grown on plastic and collagen, we compared the colony staining efficiencies of the widely used methylene blue, and Ethidium bromide (ETeB) stains. Results show that the ETeB protocol works well on plastic and is extremely effective for staining colonies on collagen when compared to methylene blue. The key features and advantages of ETeB technique are; (a) reduction in background for colonies grown on collagen and possibly other substrates, (b) the whole procedure takes less than a minute, (c) no post-stain washing step is required which eliminates colony losses for cell lines that are loosely adherent, (d) colony visualization and counting can be done immediately following the staining procedure using a standard UV illuminator and software, and (e) the method works across a wide variety of cell lines. The simplicity and robustness of this procedure should warrant its usage in both small and large-scale clonogenic experiments.

  12. Hidden stressors in the clonogenic assay used in radiobiology experiments

    International Nuclear Information System (INIS)

    Potter, M.D.E.; Suchowerska, N.; Rizvi, S.; McKenzie, D.R.

    2011-01-01

    Full text: While clonogenic assays are extensively used in radiobiology, there is no widely accepted procedure for choosing the composition of the cell culture media. Cell line suppliers recommend a specific culture medium for each cell line, however a researcher will frequently customize this aspect of the protocol by supplementing the recommended support medium with additives. For example, many researchers add antibiotics, in order to avoid contamination of cells and the consequent loss of data, with little discussion of the influence of the antibiotics on the clonogenic survival of the cells. It is assumed that the effect of any variables in the growth medium on cell survival is taken into consideration by comparing the survival fraction relative to that of controls grown under the same conditions. In the search for better cancer treatment, the effect of various stressors on clonogenic cell survival is under investigation. This study seeks to identify and test potential stressors commonly introduced into the cell culture medium, which may confound the response to radiation. (author)

  13. The mechanisms of Micrococcus radiodurans radioresistance

    International Nuclear Information System (INIS)

    Samojlenko, I.I.

    1983-01-01

    Modern representations on molecular mechanisms of Micrococcus radiodurans viability under the effect of ionizing radiation have been considered. Factors conditioning a high level of micrococcus cell radiostability have been analyzed: peculiarities of structure of a cell wall, DNA, membranes; excess of genetic information; multiplicity of DNA implantation points to a membrane; high level of antioxidation and antiradical systems. It has been shown that on efficiency of accurate, properly balanced system of DNA repair combined with above Micrococcus radiodurant properties provide a high microorganism radioresistance

  14. Postradiation recovery and radioresistance of seeds

    International Nuclear Information System (INIS)

    Atayan, R.R.; Semerdzhyan, S.P.

    1990-01-01

    Problems on radiobiology of seeds were studied. Irradiation was shown to induce primary injuries in chromosomes sensitive to various modifying factors. Primary injuries can eliminate and form stable physico-chemical injuries as a result of physico-chemical reactions and under concomitant factors. These injuries can be reversible and nonreversible character. Reversible injuries turned out to be potential for they can be repaired at biochemical level. The cells of strong ability to repair postirradiation injuries under various equal conditions reveal a relatively high level of radioresistance. The mechanisms were shown to be basical in the observed differences of radiosensitivity of large and small seeds. 7 refs.; 3 figs

  15. Receptor-Mediated Endocytosis and Brain Delivery of Therapeutic Biologics

    Directory of Open Access Journals (Sweden)

    Guangqing Xiao

    2013-01-01

    Full Text Available Transport of macromolecules across the blood-brain-barrier (BBB requires both specific and nonspecific interactions between macromolecules and proteins/receptors expressed on the luminal and/or the abluminal surfaces of the brain capillary endothelial cells. Endocytosis and transcytosis play important roles in the distribution of macromolecules. Due to the tight junction of BBB, brain delivery of traditional therapeutic proteins with large molecular weight is generally not possible. There are multiple pathways through which macromolecules can be taken up into cells through both specific and nonspecific interactions with proteins/receptors on the cell surface. This review is focused on the current knowledge of receptor-mediated endocytosis/transcytosis and brain delivery using the Angiopep-2-conjugated system and the molecular Trojan horses. In addition, the role of neonatal Fc receptor (FcRn in regulating the efflux of Immunoglobulin G (IgG from brain to blood, and approaches to improve the pharmacokinetics of therapeutic biologics by generating Fc fusion proteins, and increasing the pH dependent binding affinity between Fc and FcRn, are discussed.

  16. Modulation of clonogenicity, growth, and radiosensitivity of three human epidermoid tumor cell lines by a fibroblastic environment

    International Nuclear Information System (INIS)

    Gery, Bernard; Little, John B.; Coppey, Jacques

    1996-01-01

    Purpose: To develop a model vitro system to examine the influence of fibroblasts on the growth and survival of human tumor cells after exposure to ionizing radiation. Methods and Materials: The cell system consists of three epidermoid carcinoma cell lines derived from head and neck tumors having differing growth potentials and intrinsic radiosensitivities, as well as a low passage skin fibroblast strain from a normal human donor. The tumor cells were seeded for five days prior to exposure to radiation: (a) in the presence of different numbers of fibroblasts, (b) in conditioned medium from stationary fibroblast cultures, and (c) on an extracted fibroblastic matrix. Results: When grown with fibroblasts, all three tumor cell lines showed increased clonogenicity and increased radioresistance. The radioprotective effect was maximal at a density of approximately 10 5 fibroblasts/100 mm Petri dish, and was greatest in the intrinsically radiosensitive tumor cell line. On the other hand, the effects of incubation with conditioned medium or on a fibroblastic matrix varied among the tumor cell lines. Thus, the protective effect afforded by coculture with fibroblasts must involve several cellular factors related to the fibroblast itself. Conclusions: These observations emphasize the importance of cultural conditions on the apparent radiosensitivity of human tumor cell lines, and suggest that the fibroblastic connective tissue enveloping the malignant cells should be considered when the aim is to establish a radiopredictive assay from surgical tumors fragments

  17. Criteria for estimation of radioresistance of testudo horsfieldi

    Energy Technology Data Exchange (ETDEWEB)

    Turdyev, A.A.; Bogdanova-Berezovskaya, I.G.; Dvornikova, L.I. (AN Uzbekskoj SSR, Tashkent. Inst. Zoologii i Parazitologii)

    A study was made of a phenomenal radioresistance of Testudo horsfieldi. The authors described the features of the postirradiation death at different seasons. A threshold number of the affected systems in the turtles was one order lower than that of mammals. A positive correlaton was found between specific affection of the systems and the degree of their radioresistance.

  18. Somatostatin receptor-mediated imaging and therapy: basic science, current knowledge, limitations and future perspectives

    Energy Technology Data Exchange (ETDEWEB)

    Breeman, W.A.P.; Jong, M. de; Kwekkeboom, D.J.; Valkema, R.; Bakker, W.H.; Kooij, P.P.M. [Dept. of Nuclear Medicine, Erasmus Medical Centre Rotterdam (Netherlands); Visser, T.J. [Dept. of Internal Medicine, Erasmus Medical Centre Rotterdam (Netherlands); Krenning, E.P. [Dept. of Nuclear Medicine, Erasmus Medical Centre Rotterdam (Netherlands); Dept. of Internal Medicine, Erasmus Medical Centre Rotterdam (Netherlands)

    2001-09-01

    In vivo somatostatin receptor-mediated scintigraphy has proven to be a valuable method for the visualisation of neuroendocrine tumours and their metastases. A new application is the use of radiolabelled analogues for somatostatin receptor-mediated therapy. This paper presents a review on the basic science, historical background and current knowledge of somatostatin receptor subtypes and their expression in neuroendocrine tumours. New somatostatin analogues, new chelators, ''new'' radionuclides and combinations thereof are also discussed. Due attention is given to limitations and future perspectives of somatostatin receptor-mediated imaging and therapy. (orig.)

  19. Somatostatin receptor-mediated imaging and therapy: basic science, current knowledge, limitations and future perspectives

    International Nuclear Information System (INIS)

    Breeman, W.A.P.; Jong, M. de; Kwekkeboom, D.J.; Valkema, R.; Bakker, W.H.; Kooij, P.P.M.; Visser, T.J.; Krenning, E.P.

    2001-01-01

    In vivo somatostatin receptor-mediated scintigraphy has proven to be a valuable method for the visualisation of neuroendocrine tumours and their metastases. A new application is the use of radiolabelled analogues for somatostatin receptor-mediated therapy. This paper presents a review on the basic science, historical background and current knowledge of somatostatin receptor subtypes and their expression in neuroendocrine tumours. New somatostatin analogues, new chelators, ''new'' radionuclides and combinations thereof are also discussed. Due attention is given to limitations and future perspectives of somatostatin receptor-mediated imaging and therapy. (orig.)

  20. Fcγ receptor-mediated inflammation inhibits axon regeneration.

    Directory of Open Access Journals (Sweden)

    Gang Zhang

    Full Text Available Anti-glycan/ganglioside antibodies are the most common immune effectors found in patients with Guillain-Barré Syndrome, which is a peripheral autoimmune neuropathy. We previously reported that disease-relevant anti-glycan autoantibodies inhibited axon regeneration, which echo the clinical association of these antibodies and poor recovery in Guillain-Barré Syndrome. However, the specific molecular and cellular elements involved in this antibody-mediated inhibition of axon regeneration are not previously defined. This study examined the role of Fcγ receptors and macrophages in the antibody-mediated inhibition of axon regeneration. A well characterized antibody passive transfer sciatic nerve crush and transplant models were used to study the anti-ganglioside antibody-mediated inhibition of axon regeneration in wild type and various mutant and transgenic mice with altered expression of specific Fcγ receptors and macrophage/microglia populations. Outcome measures included behavior, electrophysiology, morphometry, immunocytochemistry, quantitative real-time PCR, and western blotting. We demonstrate that the presence of autoantibodies, directed against neuronal/axonal cell surface gangliosides, in the injured mammalian peripheral nerves switch the proregenerative inflammatory environment to growth inhibitory milieu by engaging specific activating Fcγ receptors on recruited monocyte-derived macrophages to cause severe inhibition of axon regeneration. Our data demonstrate that the antibody orchestrated Fcγ receptor-mediated switch in inflammation is one mechanism underlying inhibition of axon regeneration. These findings have clinical implications for nerve repair and recovery in antibody-mediated immune neuropathies. Our results add to the complexity of axon regeneration in injured peripheral and central nervous systems as adverse effects of B cells and autoantibodies on neural injury and repair are increasingly recognized.

  1. Targeting DNA repair with PNKP inhibition sensitizes radioresistant prostate cancer cells to high LET radiation.

    Directory of Open Access Journals (Sweden)

    Pallavi Srivastava

    Full Text Available High linear energy transfer (LET radiation or heavy ion such as carbon ion radiation is used as a method for advanced radiotherapy in the treatment of cancer. It has many advantages over the conventional photon based radiotherapy using Co-60 gamma or high energy X-rays from a Linear Accelerator. However, charged particle therapy is very costly. One way to reduce the cost as well as irradiation effects on normal cells is to reduce the dose of radiation by enhancing the radiation sensitivity through the use of a radiomodulator. PNKP (polynucleotide kinase/phosphatase is an enzyme which plays important role in the non-homologous end joining (NHEJ DNA repair pathway. It is expected that inhibition of PNKP activity may enhance the efficacy of the charged particle irradiation in the radioresistant prostate cancer cell line PC-3. To test this hypothesis, we investigated cellular radiosensitivity by clonogenic cell survival assay in PC-3 cells.12Carbon ion beam of62 MeVenergy (equivalent 5.16 MeV/nucleon and with an entrance LET of 287 kev/μm was used for the present study. Apoptotic parameters such as nuclear fragmentation and caspase-3 activity were measured by DAPI staining, nuclear ladder assay and colorimetric caspase-3method. Cell cycle arrest was determined by FACS analysis. Cell death was enhanced when carbon ion irradiation is combined with PNKPi (PNKP inhibitor to treat cells as compared to that seen for PNKPi untreated cells. A low concentration (10μM of PNKPi effectively radiosensitized the PC-3 cells in terms of reduction of dose in achieving the same survival fraction. PC-3 cells underwent significant apoptosis and cell cycle arrest too was enhanced at G2/M phase when carbon ion irradiation was combined with PNKPi treatment. Our findings suggest that combined treatment of carbon ion irradiation and PNKP inhibition could enhance cellular radiosensitivity in a radioresistant prostate cancer cell line PC-3. The synergistic effect of PNKPi

  2. Dexamethasone-induced radioresistance occurring independent of human papilloma virus gene expression in cervical carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Rutz, H.P.; Mariotta, M.; Mirimanoff, R.O. [Lab. de Radiobiologie, Service de Radio-Oncologie, CHUV, Lausanne (Switzerland); Knebel Doeberitz, M. von [Deutsches Krebsforschungszentrum, Heidelberg (Germany). Inst. fuer Virusforschung

    1998-02-01

    The aim of this study was to investigate the role of HPV 18 E6 and E7 gene products with respect to radiosensitivity of two cervical carcinoma cell lines. The two cervical carcinoma lines C4-1 and SW 756 were used in which treatment with dexamethasone allows to modulate expression levels of HPV 18 E6 and E7 genes: Upregulation in C4-1, down-regulation in SW 756. Effects of treatment with dexamethasone on plating efficiency and radiosensitivity were assessed using a clonogenic assay. Treatment with dexamethasone increased plating efficiency of the C4-1 cells, but did not affect plating efficiency of SW 756 cells. Treatment with dexamethasone induced enhanced radioresistance in both cell lines. Thus, in C4-1 cells the observed changes in radioresistance correlate to the enhancement in expression of HPV 18 genes E6/E7, whereas in SW 756, a reduced expression correlates negatively with the enhanced radioresistance. (orig./MG) [Deutsch] Das Ziel dieser Studie lag darin, die Rolle der HPV-18-Gene E6 und E7 in bezug auf die Strahlenempfindlichkeit von menschlichen Zervixkarzinomzellen zu untersuchen. Wir verwendeten zwei menschliche Zervixkarzinomzellinien, C4-1 und SW 756, in welchen die Expression der viralen Gene HPV 18 E6 und E7 mit Dexamethason moduliert werden kann: In C4-1 bewirkt die Behandlung mit Dexamethason eine Erhoehung der Expression dieser Gene, in SW 756 eine Verminderung. Die Wirkung auf die Wachstumsfaehigkeit der Zellen und auf die Wachstumshemmung durch die Bestrahlung wurde unter Verwendung eines klonogenen Assays bestimmt. Dexamethason bewirkte eine erhoehte Wachstumsfaehigkeit der C4-1 Zellen, ohne die Wachstumsfaehigkeit der SW-756-Zellen zu beeinflussen, wie schon frueher beschrieben. Die Resistenz beider Zellinien gegenueber Bestrahlung wurde erhoeht. Somit besteht in den C4-1-Zellen eine Korrelation der Expression der viralen Gene mit der Zunahme der Strahlenresistenz, wogegen in den SW-756-Zellen die Abnahme der Expression im Gegensatz zu

  3. Radioresponsiveness at low doses. Hyper-radiosensitivity and increased radioresistance in mammalian cells

    International Nuclear Information System (INIS)

    Skov, K.A.

    1999-01-01

    The rationale for and importance of research on effects after radiation at 'low doses' are outlined. Such basic radiobiological studies on induction of repair enzymes, protective mechanisms, priming, and hypersensitivity are certainly all relevant to treatment of cancer (see Section 1, Studies at low doses - relevance to cancer treatment). Included are examples from many groups, using various endpoints to address the possibility of an induced resistance, which has been compared to the adaptive response [M.C. Joiner, P. Lambin, E.P. Malaise, T. Robson, J.E. Arrand, K.A. Skov, B. Marples, Hypersensitivity to very low single radiation doses: its relationship to the adaptive response and induced radioresistance, Mutat. Res. 358 (1996) 171-183.]. This is not intended to be an exhaustive review - rather a re-introduction of concepts such as priming and a short survey of molecular approaches to understanding induced resistance. New data on the response of HT29 cells after treatment (priming) with co-cultured activated neutrophils are included, with protection against X-rays (S1). Analysis of previously published results in various cells lines in terms of increased radioresistance (IRR)/intrinsic sensitivity are presented which complement a study on human tumour lines [P. Lambin, E.P. Malaise, M.C. Joiner, Might intrinsic radioresistance of human tumour cells be induced by radiation?, Int. Radiat. Biol. 69 (1996) 279-290]. It is not feasible to extrapolate to low doses from studies at high doses. The biological responses probably vary with dose, LET, and have variable time frames. The above approaches may lead to new types of treatment, or additional means to assess radioresponsiveness of tumours. Studies in many areas of biology would benefit from considerations of different dose regions, as the biological responses vary with dose. There may also be some implications in the fields of radiation protection and carcinogenesis, and the extensions of concepts of hyper

  4. Studies on radioresistance with HeLa cells

    International Nuclear Information System (INIS)

    Koiwai, Soichiro; Muta, Nobuyoshi

    1980-01-01

    In our previous experiments (15, 16), HeLa cells were successively irradiated with 1 kR of X-rays. After receiving 3, 5, 8, 11, 14, and 17 kR the iradiated HeLa cells showed progressively increasing radioresistance. In the present paper, the results of studies on the variation of the extrapolation number (n), mean lethal dose (D sub(o)), and quasi-threshold dose (D sub(q)) of survival curves for these radioresistant HeLa strains were reported. The values of n of the original and radioresistant strains were all found between 2 and 3, and a change of the n values with an increase in the total dose received by the cell strains was not noticed. On the other hand, the values of D sub(o) of the radioresistant strains progressively increased from 105 R in the original line to 148 R in the 17 kR strain with increasing doses of pre-irradiation. Similarly, the values of D sub(q) had a tendency to increase in the radioresistant strains. Repair of sublethal damage with the original and radioresistant strains was studied and the properties of the radioresistant strains were discussed. (author)

  5. CD133+ cells contribute to radioresistance via altered regulation of DNA repair genes in human lung cancer cells

    International Nuclear Information System (INIS)

    Desai, Amar; Webb, Bryan; Gerson, Stanton L.

    2014-01-01

    Background: Radioresistance in human tumors has been linked in part to a subset of cells termed cancer stem cells (CSCs). The prominin 1 (CD133) cell surface protein is proposed to be a marker enriching for CSCs. We explore the importance of DNA repair in contributing to radioresistance in CD133+ lung cancer cells. Materials and methods: A549 and H1299 lung cancer cell lines were used. Sorted CD133+ cells were exposed to either single 4 Gy or 8 Gy doses and clonogenic survival measured. ϒ-H2AX immunofluorescence and quantitative real time PCR was performed on sorted CD133+ cells both in the absence of IR and after two single 4 Gy doses. Lentiviral shRNA was used to silence repair genes. Results: A549 but not H1299 cells expand their CD133+ population after single 4 Gy exposure, and isolated A549 CD133+ cells demonstrate IR resistance. This resistance corresponded with enhanced repair of DNA double strand breaks (DSBs) and upregulated expression of DSB repair genes in A549 cells. Prior IR exposure of two single 4 Gy doses resulted in acquired DNA repair upregulation and improved repair proficiency in both A549 and H1299. Finally Exo1 and Rad51 silencing in A549 cells abrogated the CD133+ IR expansion phenotype and induced IR sensitivity in sorted CD133+ cells. Conclusions: CD133 identifies a population of cells within specific tumor types containing altered expression of DNA repair genes that are inducible upon exposure to chemotherapy. This altered gene expression contributes to enhanced DSB resolution and the radioresistance phenotype of these cells. We also identify DNA repair genes which may serve as promising therapeutic targets to confer radiosensitivity to CSCs

  6. Establishment of clinically relevant radioresistant cell lines and their characteristics

    International Nuclear Information System (INIS)

    Fukumoto, Manabu; Kuwahara, Yoshikazu; Suzuki, Masatoshi

    2014-01-01

    Although radiotherapy is one of the major therapeutic modalities for eradicating malignant tumors, the existence of radioresistant cells remains one of the most critical obstacles. Standard radiotherapy consists of fractionated radiation (FR) of 2-Gy X-rays once a day, 5 days a week, over 60 Gy in total. To understand the characteristics of radioresistant cells and to develop more effective radiotherapy, we have established novel radioresistant cell lines by long-term (> 5 years) exposure to moderate doses of fractionated X-rays. While all the parental human cancer cells ceased, their radioresistant derivatives continue to proliferate with daily exposure to 2-Gy FR for more than 30 days. We have coined those cells as 'clinically relevant radioresistant' (CRR) cells. Transplanted tumors into nude mice were also CRR, indicating that CRR cell lines are powerful tools to improve cancer radiotherapy. We have shown that the suppression of autophagic cell death but not apoptosis was mainly involved in cellular radioresistance. An inhibitor of the mTOR pathway which enhances autophagy was effective to overcome CRR tumors induced in nude mice. But the underlined mechanism was not through the inhibition of autophagy. Guanine nucleotide-binding protein 1 (GBP1) over expression was necessary for maintaining the CRR phenotype, but radioresistant cells were not necessarily cancer stem cells (CSCs). Targeting GBP1 positive cancer cells may be a more efficient method in conquering cancer than targeting CSCs. Slight but significant radioresistance was acquired by 0.5 Gy/12 hrs of long-term FR exposures to parental cells for more than 31 days in accordance with cyclinD1 over expression. This acquired radioresistance (ARR) was stably maintained in the tumor cells even on 31 days after the cessation of 0.5-Gy FR. Present observations give a mechanistic insight for ARR of tumor cells through long-term FR exposure, and provide novel therapeutic targets for radiosensitization

  7. Correlation of proliferative and clonogenic tumor cells in multiple myeloma

    International Nuclear Information System (INIS)

    Karp, J.E.; Burke, P.J.; Saylor, P.L.; Humphrey, R.L.

    1984-01-01

    To expand on the findings from previous clinical trials that the growth of residual tumor is increased at a predictable time following initial drug administration, malignant plasma cells from bone marrows of patients with multiple myeloma (MM) were examined for changes in proliferation and clonogenicity induced in vivo by cyclophosphamide and in vitro by drug-induced humoral stimulatory activity. Peak plasma cell [ 3 H]thymidine labeling index (LI) occurred predictably following drug and paralleled changes in agar colony formation by marrow cells obtained during therapy. Colony-forming capacity of pretreatment MM marrow populations was enhanced when those cells were cultured with humoral stimulatory activity, similar to the increased colony formation detected in Day 9 postcyclophosphamide marrows at the time of peak plasma cell LI. To further define a relationship between proliferative plasma cells and colony-forming tumor cells, MM marrows were fractionated by sedimentation on an isokinetic gradient. Enrichment of a proliferative tumor cell cohort was achieved, evidenced by [ 3 H]thymidine LI. Colony-forming cells were also enriched by isokinetic gradient sedimentation, and agar colony formation by MM marrow cell fractions correlated with the kinetic characteristics of the isolated subpopulations. These studies of whole and fractionated human MM marrow cell populations suggest that the kinetically active cells which are induced to proliferate in vivo and in vitro are closely related to the clonogenic tumor cells which produce colonies in agar and which, like those cells measured by [ 3 H]thymidine LI, respond to growth stimulation by drug-induced humoral stimulatory activity

  8. Role of endogenous substances in enhancing radioresistance background

    International Nuclear Information System (INIS)

    Goncharenko, E.N.; Gorskaya, T.G.; Gudz', T.I.; Zolotareva, L.T.; Kovaleva, Z.I.; Peshkova, E.N.

    1978-01-01

    It is shown that in Saccharomyces cerevisiae of ''wild type'' diploid cells (more radioresistant than haploid ones) are characterized by a higher content of endogenous biologically active substances, which possess a radioprotective ability (biogenous amines and SOD), and a lower level of radiosensitizing substances (hydroperoxides of higher unsaturated fatty acids). With Saccharomyces cerevisiae, bearing mutation rad 51, not all the components of the radioresistance background shoW this dependence, which is indicative of the presence of additional factors affecting radioresistance of these cells

  9. A change in radioresistance of barley under chronic irradiation

    International Nuclear Information System (INIS)

    Kal'chenko, V.A.; Shevchenko, V.A.; Fedotov, I.S.

    1981-01-01

    Mutation process and radioresistance of barley growing in an area with increased radiation background (beta radiation doss -0.6 R/day) have been analyzed. Experimental material is given with 250 kindred families where aberration level of chromosomes was analyzed in mitosis and meiosis during 4 years. It is shown that the first generations (M 1 -M 2 ) in experimental material demonstrate an increased level of structural mutations of chromosomes, while the subsequent generations (M 3 -M 4 ) demonstrate the level reduced to control. A higher radioresistance is observed in M 5 plants of chronically irradiated version as compared to nonirradiated version. It is suggested that increase in radioresistance results from the stimulation of repair systems with low doses of chronic irradiation as well as from the selection of a more radioresistant forms out of the population [ru

  10. Radioresistance-related signaling pathways in nasopharyngeal carcinoma cells

    International Nuclear Information System (INIS)

    Guo Ya; Zhu Xiaodong; Qu Song; Su Fang; Wang Qi; Zhang Wei

    2011-01-01

    Objective: To study the difference of gene expression profile between the radioresistant human nasopharyngeal carcinoma cell line CNE-2R and CNE-2, and to screen the signaling pathway associated with radioresistance of nasopharyngeal carcinoma. Methods: The radioresistant nasopharyngeal carcinoma cell line CNE-2R was constructed from the original cell line CNE-2. CNE-2R and CNE-2 cells were cultured and administered with 60 Co γ-ray irradiation at the dose of 400 cGy for 15 times. Human-6v 3.0 whole genome expression profile was used to screen the differentially expressed genes. Bioinformatic analysis was used to identify the pathways related to radioresistance. Results: The number of the differentially expressed genes that were found in these 2 experiments was 374. The Kegg pathway and Biocarta pathway analysis of the differentially expressed genes showed the biological importance of Toll-like receptor signaling pathway and IL-1 R-mediated signal transduction pathway to the radioresistance of the CNE-2R cells and the significant differences of 13 genes in these 2 pathways,including JUN, MYD88, CCL5, CXCL10, STAT1, LY96, FOS, CCL3, IL-6, IL-8, IL-1α, IL-1β, and IRAK2 (t=13.47-66.57, P<0.05). Conclusions: Toll-like receptor signaling pathway and IL-1R-mediated signal transduction pathway might be related to the occurrence of radioresistance. (authors)

  11. DNA repair mechanism in radioresistant bacteria

    International Nuclear Information System (INIS)

    Kitayama, Shigeru

    1992-01-01

    Many radiation resistant bacteria have been isolated from various sources which are not in high background field. Since Deinococcus radiodurans had been isolated first in 1956, studies on the mechanism for radioresistance were carried out mostly using this bacterium. DNA in this bacterium isn't protected against injury induced by not only ionizing radiation but also ultraviolet light. Therefore, DNA damages induced by various treatments are efficiently and accurately repaired in this cells. Damages in base and/or sugar in DNA are removed by endonucleases which, if not all, are synthesized during postirradiation incubation. Following the endonucleolytic cleavage the strand scissions in DNA are seemed to be rejoined by a process common for the repair of strand scissions induced by such as ionizing radiations. Induce protein(s) is also involved in this rejoining process of strand scissions. DNA repair genes were classified into three phenotypic groups. (1)Genes which are responsible for the endonucleolytic activities. (2) Genes involved in the rejoining of DNA strand scissions. (3) Genes which participate in genetic recombination and repair. Three genes belong to (1) and (2) were cloned onto approximately 1 kbp DNA fragments which base sequences have been determined. (author)

  12. DNA repair mechanism in radioresistant bacteria

    International Nuclear Information System (INIS)

    Kitayama, Shigeru

    1992-01-01

    Many radiation resistant bacteria have been isolated from various sources which are not in high background field. Since Deinococcus radiodurans had been isolated first in 1956, the studies on the mechanism of radioresistance were mostly carried out using this bacterium. DNA in this bacterium isn't protected against injury induced by not only ionizing radiation but also ultraviolet light. Therefore, DNA damages induced by various treatments are efficiently and accurately repaired in this cells. Damages in base and/or sugar in DNA are removed by endonucleases which, if not all, are synthesized during postirradiation incubation. Following the endonucleolytic cleavage the strand scissions in DNA are seemed to be rejoined by a process common for the repair of strand scissions induced by such as ionizing radiations. Induce protein(s) is also involved in this rejoining process of strand scissions. DNA repair genes were classified into three phenotypic groups. (1) Genes which are responsible for the endonucleolytic activities. (2) Genes involved in the rejoining of DNA strand scissions. (3) Genes which participate in genetic recombination and repair. Three genes belong to (1) and (2) were cloned onto approximately 1 kbp DNA fragments which base sequences have been determined. (author)

  13. Notch3 marks clonogenic mammary luminal progenitor cells in vivo.

    Science.gov (United States)

    Lafkas, Daniel; Rodilla, Veronica; Huyghe, Mathilde; Mourao, Larissa; Kiaris, Hippokratis; Fre, Silvia

    2013-10-14

    The identity of mammary stem and progenitor cells remains poorly understood, mainly as a result of the lack of robust markers. The Notch signaling pathway has been implicated in mammary gland development as well as in tumorigenesis in this tissue. Elevated expression of the Notch3 receptor has been correlated to the highly aggressive "triple negative" human breast cancer. However, the specific cells expressing this Notch paralogue in the mammary gland remain unknown. Using a conditionally inducible Notch3-CreERT2(SAT) transgenic mouse, we genetically marked Notch3-expressing cells throughout mammary gland development and followed their lineage in vivo. We demonstrate that Notch3 is expressed in a highly clonogenic and transiently quiescent luminal progenitor population that gives rise to a ductal lineage. These cells are capable of surviving multiple successive pregnancies, suggesting a capacity to self-renew. Our results also uncover a role for the Notch3 receptor in restricting the proliferation and consequent clonal expansion of these cells.

  14. Putative radioresistant bacterial isolate from sewage water

    International Nuclear Information System (INIS)

    Ang, April; Chua, Patricia; Perez, Kristine; Rey, April; Rivor Kristel; San Pablo, Czarina; Santos, Ernestine

    2001-01-01

    Sewage water was collected from a stagnant body of water in Balara, Quezon City. approximately 150 ml was aseptically transferred into eight Erlenmeyer flasks. Seven flasks were then subjected to different doses of radiation at the 60 Co irradiation facility, PNRI (Philippine Nuclear Research Institute) which are as follows: 0.01 kGy, 0.1 kGy, 0.5 kGy, 1 kGy, 5 kGy, 10 kGy, and 15 kGy. The remaining flask was used as the control. After irradiation, all the different treatments were subjected to colony count at the culture collection laboratory, NSRI. Results showed that the colonies from sewage water treatments irradiated at 0.01 kGy (treatment A), 0.10 kGy (treatment B), and 0.50 kGy (treatment C) exhibited a decreasing trend with colony counts 4.60 x 10 3 CFU/ml, and 1.30 x 10 3 CFU/ml, and 26 CFU/ml, respectively. Contrastingly, at 1 kGy (treatment D), high colony count of 2.95 x 10 3 CFU/ml was observed which is even higher compared to the control (1.02 x 10 3 CFU/ml). Treatment E that was irradiated at 5 kGy manifested low survival rate (25 CFU/ml) indicating the presence of few putative intermediate radioresistant bacteria. Radiation dose treatments higher than 5 kGy (i.e., 10 kGy and 15 kGy) exhibited no bacterial survival. (Author)

  15. Putative radioresistant bacterial isolate from sewage water

    Energy Technology Data Exchange (ETDEWEB)

    Ang, April; Chua, Patricia; Perez, Kristine; Rey, April; Kristel, Rivor; San Pablo, Czarina; Santos, Ernestine

    2001-01-29

    Sewage water was collected from a stagnant body of water in Balara, Quezon City. approximately 150 ml was aseptically transferred into eight Erlenmeyer flasks. Seven flasks were then subjected to different doses of radiation at the {sup 60}Co irradiation facility, PNRI (Philippine Nuclear Research Institute) which are as follows: 0.01 kGy, 0.1 kGy, 0.5 kGy, 1 kGy, 5 kGy, 10 kGy, and 15 kGy. The remaining flask was used as the control. After irradiation, all the different treatments were subjected to colony count at the culture collection laboratory, NSRI. Results showed that the colonies from sewage water treatments irradiated at 0.01 kGy (treatment A), 0.10 kGy (treatment B), and 0.50 kGy (treatment C) exhibited a decreasing trend with colony counts 4.60 x 10{sup 3} CFU/ml, and 1.30 x 10{sup 3} CFU/ml, and 26 CFU/ml, respectively. Contrastingly, at 1 kGy (treatment D), high colony count of 2.95 x 10{sup 3} CFU/ml was observed which is even higher compared to the control (1.02 x 10{sup 3} CFU/ml). Treatment E that was irradiated at 5 kGy manifested low survival rate (25 CFU/ml) indicating the presence of few putative intermediate radioresistant bacteria. Radiation dose treatments higher than 5 kGy (i.e., 10 kGy and 15 kGy) exhibited no bacterial survival. (Author)

  16. Properties of bacterial radioresistance observed in sewage sludge

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, H; Ito, H; Takehisa, M [Japan Atomic Energy Research Inst., Takasaki, Gunma. Takasaki Radiation Chemistry Research Establishment; Iizuka, H

    1981-09-01

    The changes in radiosensitivities of bacteria in sludge were investigated. The coliforms are more radioresistant in raw sludge than in cake (dewatered sludge). This radioresistance of coliforms was observed not only in raw sludge but also in the cake diluted with water. The radioresistance was independent of the difference of treatment plant, kind of sludge, and season. The oxygen effect on the radioresistance was not observed, but the resistance was changed during storage of sludge. Escherichia coli isolated from sludge was radiosensitive in buffer, but its radiosensitivity was protected by the water-extracts of sludge. On the other hand, radioresistant bacteria were present in total bacteria of sludge irradiated at 2 Mrad. However, the dominant flora in the irradiated sludge consisted of radiosensitive bacteria (mainly Pseudomonas). When a strain of radiosensitive Pseudomonas was irradiated in raw sludge and diluted cake, the radiosensitivity was remarkably protected. From these results, it is suggested that a factor affecting the radiosensitivity of bacteria is present in sludge.

  17. Survivin as a radioresistance factor in pancreatic cancer

    International Nuclear Information System (INIS)

    Asanuma, Koichi; Moriai, Ryosuke; Yajima, Tomomi; Yagihashi, Atsuhito; Yamada, Mikako; Kobayashi, Daisuke; Watanabe, Naoki

    2000-01-01

    We examined whether survivin acts as a constitutive and inducible radioresistance factor in pancreatic cancer cells. Using a quantitative TaqMan reverse transcription-polymerase chain reaction for survivin mRNA in five pancreatic cancer cell lines, we found an inverse relationship between survivin mRNA expression and radiosensitivity. PANC-1 cells, which had the highest survivin mRNA levels, were most resistant to X-irradiation; MIAPaCa-2 cells, which showed the least survivin mRNA expression, were the most sensitive to X-irradiation. Our results suggested that survivin could act as a constitutive radioresistance factor in pancreatic cancer cells. To determine whether radioresistance is enhanced by induction of survivin expression by irradiation, PANC-1 and MIAPaCa-2 cells were subjected to sublethal doses of X-irradiation followed by a lethal dose. Survivin mRNA expression was increased significantly in both PANC-1 and MIAPaCa-2 cell lines by pretreatment with a sublethal dose of X-irradiation, as was cell survival after exposure to the lethal dose. In this system, enzymatic caspase-3 activity was significantly suppressed in cells with acquired resistance. These results suggest that survivin also acts as an inducible radioresistance factor in pancreatic cancer cells. Survivin, then, appears to enhance radioresistance in pancreatic cancer cells; inhibition of survivin mRNA expression may improve the effectiveness of radiotherapy. (author)

  18. Properties of bacterial radioresistance observed in sewage sludge

    International Nuclear Information System (INIS)

    Watanabe, Hiroshi; Ito, Hitoshi; Takehisa, Masaaki; Iizuka, Hiroshi.

    1981-01-01

    The changes in radiosensitivities of bacteria in sludge were investigated. The coliforms are more radioresistant in raw sludge than in cake (dewatered sludge). This radioresistance of coliforms was observed not only in raw sludge but also in the cake diluted with water. The radioresistance was independent of the difference of treatment plant, kind of sludge, and season. The oxygen effect on the radioresistance was not observed, but the resistance was changed during storage of sludge. Escherichia coli isolated from sludge was radiosensitive in buffer, but its radiosensitivity was protected by the water-extracts of sludge. On the other hand, radioresistant bacteria were present in total bacteria of sludge irradiated at 2 Mrad. However, the dominant flora in the irradiated sludge consisted of radiosensitive bacteria (mainly Pseudomonas). When a strain of radiosensitive Pseudomonas was irradiated in raw sludge and diluted cake, the radiosensitivity was remarkably protected. From these results, it is suggested that a factor affecting the radiosensitivity of bacteria is present in sludge. (author)

  19. Establishment of a radioresistant human lung cancer cell subline and its mechanism of radioresistance

    International Nuclear Information System (INIS)

    Zhao Wei; Wang Qiong; Liu Li; Shi Xing; Ding Qian; Wu Gang

    2008-01-01

    Objective: To establish a radioresistant cell subline from a human A549 lung cancer cell line and investigate the mechanism of radioresistance. Methods: Two proposals were applied for the non-small cell lung cancer A549 cells irradiated with X-rays: A group of A549 cell line was irradiated five times, the fractionated dose was 600 cGy, and the other group was exposed 15 times, the fractionated dose was 200 cGy. After the completion of irradiation, two monoclones were obtained from the survival of cells and named the subline A549-S1 and A549-S2. The radiosensitivity and cell cycle distribution of these two clones, together with its parental A549 cells were measured by clone formation assay and flow cytometry. The mRNA and protein levels of Notchl in A549 cell line and the sublines were determined by RT-PCR and Western-blots. Results: Compared with the parental A549 cells, A549-S1 cells showed significant resistance to radiation with D 0 , D q and N values increased, and a broader initial shoulder as well as 1.38-fold increased value of SF 2 . The A549-S1 subline also showed higher percentage of cells in S phase and G 2 /M phase, but lower percentages in G 1 /G 1 phase (P 0 , D q and N values decreased and a curve initial shoulder. The ratio of cells in S and G 0 /G 1 phase ratio was lower than that in parental A549 cells, but that in G 2 /M phase ratio was higher significantly (P<0.05). The expression of Notchl had no marked change compared to A549 cell. Conclusions: The radioresistance of the A549 cell subline is correlated with the irradiation program. The cell subline shows a different cell cycle distribution from their parental line. The cell cycle distribution has a close correlaiton with the expression of Notchl. (authors)

  20. Cancer-associated adipocytes promotes breast tumor radioresistance

    Energy Technology Data Exchange (ETDEWEB)

    Bochet, Ludivine; Meulle, Aline [Universite de Toulouse, UPS, F-31077 Toulouse Cedex (France); CNRS, IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 route de Narbonne, BP 64182, F-31077 Toulouse Cedex (France); Institut National de la Sante et de la Recherche Medicale, INSERM U1048, 1 Avenue du Pr Jean Poulhes, BP 84225, F-31432 Toulouse Cedex (France); Imbert, Sandrine [CNRS, IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 route de Narbonne, BP 64182, F-31077 Toulouse Cedex (France); Salles, Bernard [Universite de Toulouse, UPS, F-31077 Toulouse Cedex (France); CNRS, IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 route de Narbonne, BP 64182, F-31077 Toulouse Cedex (France); Valet, Philippe [Universite de Toulouse, UPS, F-31077 Toulouse Cedex (France); Institut National de la Sante et de la Recherche Medicale, INSERM U1048, 1 Avenue du Pr Jean Poulhes, BP 84225, F-31432 Toulouse Cedex (France); Muller, Catherine, E-mail: muller@ipbs.fr [Universite de Toulouse, UPS, F-31077 Toulouse Cedex (France); CNRS, IPBS (Institut de Pharmacologie et de Biologie Structurale), 205 route de Narbonne, BP 64182, F-31077 Toulouse Cedex (France)

    2011-07-22

    Highlights: {yields} Tumor-surrounding adipocytes contribute to breast cancer progression. {yields} Breast tumor cells previously co-cultivated with mature adipocytes exhibit radioresistance. {yields} Increased in Chk1 phosphorylation is observed in irradiated co-cultivated tumor cells. {yields} IL-6 is over-expressed in tumor cells co-cultivated with adipocytes. {yields} IL-6 exposure confers increased Chk1 phosphorylation and radioresistance in tumor cells. -- Abstract: Mature adipocytes are excellent candidates to influence tumor behavior through heterotypic signaling processes since these cells produce hormones, growth factors, cytokines and other molecules, a heterogeneous group of molecules named adipokines. Using a 2D coculture system, we demonstrate that breast tumor cells previously co-cultivated with mature adipocytes exhibit radioresistance and an earlier and higher increase in the effector kinase Chk1, a phenotype that was associated with decreased cell death as compared to tumor cells grown alone. Interestingly, the adipocytes-induced tumor changes taking place during the coculture time preceding the exposure to IR were sufficient to confer the radioresistant effect. Notorious among the changes brought by adipocytes was the significant increase of IL-6 expression in tumor cells, whose activity may well account for the observed tumor cell protection from IR toxicity. Indeed, our data confirmed the protective role of this cytokine as tumor cells incubated after irradiation with recombinant IL-6 exhibit an increased in Chk1 phosphorylation and a radioresistant phenotype, thus far recapitulating the effects observed in the presence of adipocytes. Our current study sheds light on a new role of tumor-surrounding adipocytes in fostering a radioresistant phenotype in breast tumors, a finding that might have important clinical implications in obese patients that frequently exhibit aggressive diseases.

  1. The proliferative status of intestinal epithelial clonogenic cells: sensitivity to S phase specific cytotoxic agents.

    Science.gov (United States)

    Boarder, T A; Blackett, N M

    1976-11-01

    High concentrations of tritiated thymidine and cytosine arabinoside (Ara-C) have been used to selectively kill cells in the crypts of Lieberkuhn that are synthesizing DNA. The effect of these agents on the number of regenerating microcolonies seen 3 1/2 days after a range of radiation doses indicates that a majority of the clonogenic cells are proliferating rapidly and that the slowly proliferating cells at the base of the crypt do not represent the whole clonogenic population.

  2. Radioresistance increase in polymers at high pressures. [. gamma. rays

    Energy Technology Data Exchange (ETDEWEB)

    Milinchuk, V; KIRJUKHIN, V; KLINSHPONT, E

    1977-06-01

    The effect was studied of very high pressures ranging within 100 and 2,700 MPa on the radioresistance of polytetrafluoroethylene, polypropylene and polyethylene in gamma irradiation. For experiments industrial polymers in the shape of blocks, films and fibers were used. It is shown that in easily breakable polymers, such as polytetrafluoroethylene and polypropylene, 1.3 to 2 times less free radicals are formed as a result of gamma irradiation and a pressure of 150 MPa than at normal pressure. The considerably reduced radiation-chemical formation of radicals and the destruction suppression by cross-linking in polymers is the evidence of the polymer radioresistance in irradiation at high pressures.

  3. Effects of radiation type and delivery mode on a radioresistant eukaryote Cryptococcus neoformans

    International Nuclear Information System (INIS)

    Shuryak, Igor; Bryan, Ruth A.; Broitman, Jack; Marino, Stephen A.; Morgenstern, Alfred; Apostolidis, Christos; Dadachova, Ekaterina

    2015-01-01

    Introduction: Most research on radioresistant fungi, particularly on human pathogens such as Cryptococcus neoformans, involves sparsely-ionizing radiation. Consequently, fungal responses to densely-ionizing radiation, which can be harnessed to treat life-threatening fungal infections, remain incompletely understood. Methods: We addressed this issue by quantifying and comparing the effects of densely-ionizing α-particles (delivered either by external beam or by 213 Bi-labeled monoclonal antibodies), and sparsely-ionizing 137 Cs γ-rays, on Cryptococus neoformans. Results: The best-fit linear-quadratic parameters for clonogenic survival were the following: α = 0.24 × 10 −2 Gy −1 for γ-rays and 1.07 × 10 −2 Gy −1 for external-beam α-particles, and β = 1.44 × 10 −5 Gy −2 for both radiation types. Fungal cell killing by radiolabeled antibodies was consistent with predictions based on the α-particle dose to the cell nucleus and the linear-quadratic parameters for external-beam α-particles. The estimated RBE (for α-particles vs. γ-rays) at low doses was 4.47 for the initial portion of the α-particle track, and 7.66 for the Bragg peak. Non-radiological antibody effects accounted for up to 23% of cell death. Conclusions: These results quantify the degree of C. neoformans resistance to densely-ionizing radiations, and show how this resistance can be overcome with fungus-specific radiolabeled antibodies

  4. Cell lysis and superoxide dismutase activities of highly radioresistant bacteria

    International Nuclear Information System (INIS)

    Yoshinaka, Taeko; Yano, Keiji; Yamaguchi, Hikoyuki

    1976-01-01

    The highly radioresistant bacterium, Arthrobacter radiotolerans, has been isolated from the radioactive hot spring of Misasa, and it does not sporulate, it is Gram-positive, and its color is pink to red. This bacterium shows the highest resistance to gamma-ray among Gram-positive resistants, but the lytic enzyme capable of lysing the cells of strong radioresistants and the surface structure of the cells are little known except those about Micrococcus radiodurans. The cells of the M. radiodurans can be lysed by Achramobacter lyticus enzyme, and electron microscopic observation and chemical analysis revealed the mutilayered surface structure of the cells consisting of an inner membrane, a mucopeptide wall layer and a very outer layer. The superoxide dismutase (SOD) activity of aerobic and anaerobic bacteria was studied, and the relatively high SOD activity of the M. radiodurans was found. The SOD function acts against the threat posed by the reactive superoxide radical being generated biologically, photochemically and radiochemically in the presence of molecular oxygen. In this paper, it is reported that the lytic enzyme No.2 obtained from Cytophaga sp., containing N-acetyl-muramyl-L-alanine amidase, peptidase and endopeptidase, and showing broad lytic spectra, was able to lyse the cells of A. radiotolerans and four radioresistant micrococci, and the radioresistant bacteria showed relatively high SOD activity except M. sp. 248. It is well known that superoxide anions are generated by aerobic irradiation, and are toxic to microbial cells. (Kako, I.)

  5. Cell lysis and superoxide dismutase activities of highly radioresistant bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Yoshinaka, T; Yano, K; Yamaguchi, H [Tokyo Univ. (Japan). Faculty of Agriculture

    1976-01-01

    The highly radioresistant bacterium, Arthrobacter radiotolerans, has been isolated from the radioactive hot spring of Misasa, and it does not sporulate, it is Gram-positive, and its color is pink to red. This bacterium shows the highest resistance to gamma-ray among Gram-positive resistants, but the lytic enzyme capable of lysing the cells of strong radioresistants and the surface structure of the cells are little known except those about Micrococcus radiodurans. The cells of the M. radiodurans can be lysed by Achramobacter lyticus enzyme, and electron microscopic observation and chemical analysis revealed the mutilayered surface structure of the cells consisting of an inner membrane, a mucopeptide wall layer and a very outer layer. The superoxide dismutase (SOD) activity of aerobic and anaerobic bacteria was studied, and the relatively high SOD activity of the M. radiodurans was found. The SOD function acts against the threat posed by the reactive superoxide radical being generated biologically, photochemically and radiochemically in the presence of molecular oxygen. In this paper, it is reported that the lytic enzyme No.2 obtained from Cytophaga sp., containing N-acetyl-muramyl-L-alanine amidase, peptidase and endopeptidase, and showing broad lytic spectra, was able to lyse the cells of A. radiotolerans and four radioresistant micrococci, and the radioresistant bacteria showedrelatively high SOD activity except M. sp. 248. It is well known that superoxide anions are generated by aerobic irradiation, and are toxic to microbial cells.

  6. Transition in Survival From Low-Dose Hyper-Radiosensitivity to Increased Radioresistance Is Independent of Activation of ATM SER1981 Activity

    International Nuclear Information System (INIS)

    Krueger, Sarah A.; Collis, Spencer J.; Joiner, Michael C.; Wilson, George D.; Marples, Brian

    2007-01-01

    Purpose: The molecular basis of low-dose hyper-radiosensitivity (HRS) is only partially understood. The aim of this study was to define the roles of ataxia telangiectasia mutated (ATM) activity and the downstream ATM-dependent G 2 -phase cell cycle checkpoint in overcoming HRS and triggering radiation resistance. Methods and Materials: Survival was measured using a high-resolution clonogenic assay. ATM Ser1981 activation was measured by Western blotting. The role of ATM was determined in survival experiments after molecular (siRNA) and chemical (0.4 mM caffeine) inhibition and chemical (20 μg/mL chloroquine, 15 μM genistein) activation 4-6 h before irradiation. Checkpoint responsiveness was assessed in eight cell lines of differing HRS status using flow cytometry to quantify the progression of irradiated (0-2 Gy) G 2 -phase cells entering mitosis, using histone H3 phosphorylation analysis. Results: The dose-response pattern of ATM activation was concordant with the transition from HRS to radioresistance. However, ATM activation did not play a primary role in initiating increased radioresistance. Rather, a relationship was discovered between the function of the downstream ATM-dependent early G 2 -phase checkpoint and the prevalence and overcoming of HRS. Four cell lines that exhibited HRS failed to show low-dose ( 2 -phase checkpoint. These data suggest that clinical exploitation of HRS could be achieved by combining radiotherapy with chemotherapeutic agents that modulate this cell cycle checkpoint

  7. Reciprocal Regulation of Hypoxia-Inducible Factor 2α and GLI1 Expression Associated With the Radioresistance of Renal Cell Carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jiancheng [Department of Urology, First Affiliated Hospital of Medical School, Xi' an Jiaotong University, Xi' an (China); Department of Urology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); Wu, Kaijie [Department of Urology, First Affiliated Hospital of Medical School, Xi' an Jiaotong University, Xi' an (China); Gao, Dexuan [Department of Urology, Shandong Provincial Hospital affiliated with Shandong University, Ji' nan (China); Zhu, Guodong; Wu, Dapeng; Wang, Xinyang; Chen, Yule; Du, Yuefeng; Song, Wenbin; Ma, Zhenkun [Department of Urology, First Affiliated Hospital of Medical School, Xi' an Jiaotong University, Xi' an (China); Authement, Craig; Saha, Debabrata [Department of Urology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); Hsieh, Jer-Tsong, E-mail: jt.hsieh@utsouthwestern.edu [Department of Urology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); He, Dalin, E-mail: dalinhe@yahoo.com [Department of Urology, First Affiliated Hospital of Medical School, Xi' an Jiaotong University, Xi' an (China)

    2014-11-15

    Purpose: Renal cell carcinoma (RCC) is often considered a radioresistant tumor, but the molecular mechanism underlying its radioresistance is poorly understood. This study explored the roles of hypoxia-inducible factor 2α (HIF2α) and sonic hedgehog (SHH)-GLI1 signaling in mediating the radioresistance of RCC cells and to unveil the interaction between these 2 signaling pathways. Methods and Materials: The activities of SHH-GLI1 signaling pathway under normoxia and hypoxia in RCC cells were examined by real-time polymerase chain reaction, Western blot, and luciferase reporter assay. The expression of HIF2α and GLI1 in RCC patients was examined by immunohistochemistry, and their correlation was analyzed. Furthermore, RCC cells were treated with HIF2α-specific shRNA (sh-HIF2α), GLI1 inhibitor GANT61, or a combination to determine the effect of ionizing radiation (IR) on RCC cells based on clonogenic assay and double-strand break repair assay. Results: RCC cells exhibited elevated SHH-GLI1 activities under hypoxia, which was mediated by HIF2α. Hypoxia induced GLI1 activation through SMO-independent pathways that could be ablated by PI3K inhibitor or MEK inhibitor. Remarkably, the SHH-GLI1 pathway also upregulated HIF2α expression in normoxia. Apparently, there was a positive correlation between HIF2α and GLI1 expression in RCC patients. The combination of sh-HIF2α and GLI1 inhibitor significantly sensitized RCC cells to IR. Conclusions: Cross-talk between the HIF2α and SHH-GLI1 pathways was demonstrated in RCC. Cotargeting these 2 pathways, significantly sensitizing RCC cells to IR, provides a novel strategy for RCC treatment.

  8. Upregulation of endothelin ETB receptor-mediated vasoconstriction in rat coronary artery after organ culture

    DEFF Research Database (Denmark)

    Eskesen, Karen; Edvinsson, Lars

    2006-01-01

    The aim of this study was to examine if endothelin ET(B) receptor-mediated contraction occurred in isolated segments of rat coronary arteries during organ culture. Presence of contractile endothelin ET(B) receptors was studied by measuring the change in isometric tension in rings of left anterior......(+)-solution was not modified after 1 day in culture medium. The experiments indicate that organ culture of rat coronary arteries upregulate endothelin ET(B) receptor-mediated contraction by inducing synthesis of new protein....... descending coronary arteries isolated from hearts of rats as response to application of the selective endothelin ET(B) receptor agonist, Sarafotoxin 6c and endothelin-1. In segments cultured 1 day in serum free Dulbecco's Modified Eagle's Medium, Sarafotoxin 6c induced a concentration dependent contraction...

  9. THIP, a hypnotic and antinociceptive drug, enhances a tonic GABAA receptor mediated conductance in mouse neocortex

    DEFF Research Database (Denmark)

    Drasbek, Kim Ryun; Jensen, Kimmo

    2006-01-01

    its cellular actions in the neocortex are uncertain, we studied the effects of THIP on neurons in slices of frontoparietal neocortex of 13- to 19-day-old (P13-19) mice. Using whole-cell patch-clamp recordings, we found that the clinically relevant THIP concentration of 1 μM induced a robust tonic GABA...... suggest that THIP activates an extrasynaptic GABA(A) receptor-mediated conductance in the neocortex, which may alter the cortical network activity....

  10. The Influence of Receptor-Mediated Interactions on Reaction-Diffusion Mechanisms of Cellular Self-organisation

    KAUST Repository

    Klika, Vá clav; Baker, Ruth E.; Headon, Denis; Gaffney, Eamonn A.

    2011-01-01

    formation, motivating numerous theoretical and experimental studies, though its verification at the molecular level in biological systems has remained elusive. In this work, we consider the influence of receptor-mediated dynamics within the framework

  11. Control of ductal vs. alveolar differentiation of mammary clonogens and susceptibility to radiation-induced mammary cancer

    International Nuclear Information System (INIS)

    Kamiya, Kenji; Yokoro, Kenjiro; Clifton, K.H.; Gould, M.N.

    1991-01-01

    We have developed an in vitro-in vivo transplantation assay for measuring the concentration of clonogenic epithelial cells in cell suspensions of rat mammary tissue. Rat mammary clonogens from organoid cultures are capable of the same degree of PLDR as clonogens in vivo. The growth and differentiation of mammary clonogens to alveolar colonies or ductal colonies is regulated as follows: a) in the presence of E 2 and high prolactin (Prl), cortisol induces mammary clonogens to proliferate and differentiate to form alveolar colonies which secrete milk and begin losing clonogenic potential, b) in cortisol deficient rats, Prl and E 2 synergistically stimulate non-secretory ductal colonies, formation of which retain clonogenic potential, c) E 2 without progesterone stimulates alveolar colony formation in the presence of cortical and high Prl, d) progesterone inhibits mammary clonogen differentiation to milk-producing cells and induces ductogenesis in a dose responsive fashion in the presence of E 2 , cortisol and high Prl. High prolactin levels coupled with glucocorticoid deficiency increases the susceptibility to mammary carcinogenesis following low dose radiation exposure by increasing the number of total mammary clonogens which are the presumptive target cells and by stimulating their proliferation after exposure. (author)

  12. Neuropeptide Y restores non-receptor-mediated vasoconstrictive action in superior mesenteric arteries in portal hypertension.

    Science.gov (United States)

    Hartl, Johannes; Dietrich, Peter; Moleda, Lukas; Müller-Schilling, Martina; Wiest, Reiner

    2015-12-01

    Vascular hyporeactivity to vasoconstrictors contributes to splanchnic arterial vasodilatation and hemodynamic dysregulation in portal hypertension. Neuropeptide Y (NPY), a sympathetic cotransmitter, has been shown to improve adrenergic vascular contractility in portal hypertensive rats and markedly attenuate hyperdynamic circulation. To further characterize the NPY-effects in portal hypertension, we investigated its role for non-receptor-mediated vasoconstriction in the superior mesenteric artery (SMA) of portal vein ligated (PVL) and sham-operated rats. Ex vivo SMA perfusion of PVL and sham rats was used to analyse the effects of NPY on pressure response to non-receptor-mediated vasoconstriction. Dose-response curves to KCl (30-300 mM) were used to bypass G protein-coupled receptor mechanisms. Potential involvement of the cyclooxygenase-pathway was tested by non-selective cyclooxygenase-inhibition using indomethacin. KCl-induced vascular contractility but not vascular sensitivity was significantly attenuated in PVL rats as compared with sham rats. Administration of NPY resulted in an augmentation of KCl-evoked vascular sensitivity being not different between study groups. However, KCl-induced vascular contractility was markedly more enhanced in PVL rats, thus, vascular response was no more significantly different between PVL and sham rats after addition of NPY. Administration of indomethacin abolished the NPY-induced enhancement of vasoconstriction. Receptor-independent vascular contractility is impaired in mesenteric arteries in portal hypertension. NPY improves non-receptor mediated mesenteric vasoconstriction more effective in portal hypertension than in healthy conditions correcting splanchnic vascular hyporesponsiveness. This beneficial vasoactive action of NPY adds to its well known more pronounced effects on adrenergic vasoconstriction in portal hypertension making it a promising therapeutic agent in portal hypertension. © 2015 John Wiley & Sons A

  13. Content of endogenous thiols and radioresistance of gemmating cells of Saccharomyces ellipsoideus and Saccharomyces cerevisiale yeasts

    International Nuclear Information System (INIS)

    Simonyan, N.V.; Avakyan, Ts.M.; Dzhanpoladyan, N.L.; Stepanyan, L.G.

    1983-01-01

    It has been shown that gemmating cells of ''wild type'' yeasts are more radioresistant and contain more endogenous thiols, than resting cells. Gemmating cells of Saccharomyces cerevisial yeasts, carrying the mutation rad 51, as to radioresistance and content of SH groups do not differ from resting cells. The results obtained testify to a connec-- tion between increased radioresistance of the yeast gemmating cells and increased content of endogenous thiols in them

  14. Receptor-Mediated Drug Delivery to Macrophages in Chemotherapy of Leishmaniasis

    Science.gov (United States)

    Mukhopadhyay, Amitabha; Chaudhuri, Gautam; Arora, Sunil K.; Sehgal, Shobha; Basu, Sandip K.

    1989-05-01

    Methotrexate coupled to maleylated bovine serum albumin was taken up efficiently through the ``scavenger'' receptors present on macrophages and led to selective killing of intracellular Leishmania mexicana amazonensis amastigotes in cultured hamster peritoneal macrophages. The drug conjugate was nearly 100 times as effective as free methotrexate in eliminating the intracellular parasites. Furthermore, in a model of experimental cutaneous leishmaniasis in hamsters, the drug conjugate brought about more than 90% reduction in the size of footpad lesions within 11 days. In contrast, the free drug at a similar concentration did not significantly affect lesion size. These studies demonstrate the potential of receptor-mediated drug delivery in the therapy of macrophage-associated diseases.

  15. Understanding magnetic nanoparticle osteoblast receptor-mediated endocytosis using experiments and modeling

    International Nuclear Information System (INIS)

    Tran, Nhiem; Webster, Thomas J

    2013-01-01

    Iron oxide nanoparticles are promising candidates for controlling drug delivery through an external magnetic force to treat a wide range of diseases, including osteoporosis. Previous studies have demonstrated that in the presence of hydroxyapatite coated magnetite (Fe 3 O 4 ) nanoparticles, osteoblast (or bone forming cell) proliferation and long-term functions (such as calcium deposition) were significantly enhanced. Hydroxyapatite is the major inorganic component of bone. As a further attempt to understand why, in the current study, the uptake of such nanoparticles into osteoblasts was experimentally investigated and mathematically modeled. Magnetite nanoparticles were synthesized using a co-precipitation method and were coated with hydroxyapatite. A cellular uptake experiment at low temperatures indicated that receptor-mediated endocytosis contributed to the internalization of the magnetic nanoparticles into osteoblasts. A model was further developed to explain the uptake of magnetic nanoparticles into osteoblasts using receptor-mediated endocytosis. This model may explain the internalization of hydroxyapatite into osteoblasts to elevate intracellular calcium levels necessary to promote osteoblast functions to treat a wide range of orthopedic problems, including osteoporosis. (paper)

  16. Melanocortin MC(4) receptor-mediated feeding and grooming in rodents.

    Science.gov (United States)

    Mul, Joram D; Spruijt, Berry M; Brakkee, Jan H; Adan, Roger A H

    2013-11-05

    Decades ago it was recognized that the pharmacological profile of melanocortin ligands that stimulated grooming behavior in rats was strikingly similar to that of Xenopus laevis melanophore pigment dispersion. After cloning of the melanocortin MC1 receptor, expressed in melanocytes, and the melanocortin MC4 receptor, expressed mainly in brain, the pharmacological profiles of these receptors appeared to be very similar and it was demonstrated that these receptors mediate melanocortin-induced pigmentation and grooming respectively. Grooming is a low priority behavior that is concerned with care of body surface. Activation of central melanocortin MC4 receptors is also associated with meal termination, and continued postprandial stimulation of melanocortin MC4 receptors may stimulate natural postprandial grooming behavior as part of the behavioral satiety sequence. Indeed, melanocortins fail to suppress food intake or induce grooming behavior in melanocortin MC4 receptor-deficient rats. This review will focus on how melanocortins affect grooming behavior through the melanocortin MC4 receptor, and how melanocortin MC4 receptors mediate feeding behavior. This review also illustrates how melanocortins were the most likely candidates to mediate grooming and feeding based on the natural behaviors they induced. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. Development of radiation countermeasure using novel radioresistant bacteria

    International Nuclear Information System (INIS)

    Kumar, Raj; Singh, Shravan K.; Malhotra, Poonam; Gupta, Ashutosh K.; Singh, Praveen K.; Chhachhia, Neha

    2014-01-01

    Radioresistant bacteria sustain their lives in extreme radiation environment and have capabilities to combat radiation induced oxidative stress. Therefore, factors associated with radioresistancy in bacteria may also provide trans-species radioprotection. To test this hypothesis, present work was initiated at INMAS long back. With this background a novel radioresistant bacterium Bacillus sp. INM-1 isolated and its novel secondary metabolite i.e. Semiquinone Glucoside Derivative (SQGD) carrying radioprotective capabilities was purified. SQGD was evaluated for its free radical scavenging, protein, enzymes, plasmid and biological membranes radioprotection capabilities in vitro. SQGD was also tested for its whole body radioprotective efficacy using oral route of administration. Systemic radioprotection offered by SQGD to gastrointestinal, haematopoietic and male reproductive system was studied. Modulation in endogenous antioxidant enzymes and cytoprotective cytokines expression upon irradiation and SQGD pretreatment was determined. A laboratory process for chemical synthesis of bacterial radioprotective molecule has also been developed (Patent filed No. 2075/DEL/2014). Results of the study demonstrated that SQGD efficiently scavenge free radicals in vitro. SQGD provides excellent protection to structural and functional proteins, plasmid DNA and biomembranes against radiation induced oxidative damage. SQGD was observed to offer ∼ 83% whole body survival to lethally irradiated mice when administered 2h before irradiation by oral route. SQGD was found to ensure significant radioprotection to gastro-intestinal, hematopoietic and male reproductive system of irradiated mice. Protein expression studies revealed that SQGD pretreatment to the irradiated mice significantly increased expression of G-CSF, GM-CSF, MCSF; NFkB, IL-2, IL-12, IL-23, IL-6, PCNA and PARP. In conclusion, present study decisively justified the radioprotective potential of bacterial metabolite SQGD

  18. Dietary enhancement of intestinal radioresistance during fractionated irradiation

    International Nuclear Information System (INIS)

    Pageau, R.; St-Pierre, C.

    1978-01-01

    Rats fed laboratory chow or elemental diet 3 were given fractions of 240 rads of 60 Co γ radiation abdominally (1200 rads/week) until all animals had died. Changes in appetite, body weight, and mortality were monitored as a function of the cumulative dose received. More radiation was needed in the diet-fed group to achieve both 0 and 100% mortality, a difference of 37% at the mean lethal dose level. Both groups developed similar progressive anorexia but the diet-fed animals lost weight more slowly. Data indicate that basic intestinal radioresistance is enhanced by feeding the elemental diet

  19. Recovery and radio-resistance in mice after external irradiation

    International Nuclear Information System (INIS)

    Le Guillou, S.

    1965-01-01

    The author presents a literature study concerning recovery from external irradiation and an analysis of experimental data (which appear to suggest the idea of a radio-resistance in animals), as well as the hypotheses put forward for explaining this phenomenon. The author then describes an experiment carried out on mice whose LD 50/30 days increased from 1005 to 1380 rads and for which it was shown that an increase occurs in the number of certain anti-bodies circulating after a low dose of γ irradiation. (author) [fr

  20. Isolation and identification of a novel radio-resistant strain

    International Nuclear Information System (INIS)

    Zhang Zhidong; Mao Jun; Wang Wei; Tang Qiyong; Shi Yuhu

    2008-01-01

    A novel radio-resistant strain named RL2 was studied polyphasically, which was isolated from the soils in the Gurban-Tunggut Desert, Xinjiang. The strain is Gam-positive, sphere-shaped and pink pigmented; The DNA (G+C) contents of RL2 is 71.62mo1%; The 16S rDNA genes of RL2 and D. radiodurans type strain DSM20539 shows a high level of similarity (97.2%). According to phenotypic characteristics and phylogenetic analysis, it can be suggested that the strain RL2 has been identified as Deinococcus. sp and it may be a novel species. (authors)

  1. Investigation of ionizing sublethal doses effects on endogenous radioresistance background

    International Nuclear Information System (INIS)

    Kudryashov, Yu.B.; Goncharenko, E.N.; Antonova, S.V.; Akhalaya, M.Ya.; Bajzhumanov, A.A.; Shestakova, S.V.

    1997-01-01

    Sublethal doses of X-radiation (0.5 Gy and 1 Gy) caused the alterations in levels of main components of endogenous radioresistance background in rat tissues. There were demonstrated the decrease of serotonin content in stomach mocosa and spleen, adrenalin, noradrenalin and corticosteroids contents in adrenal glands, nonprotein thiols content in spleen and the increase of lipid peroxide level in serum on the 3-14 days after irradiation. The recovery of the investigated parameters was occurred to the 21 day after exposure. (author)

  2. Radiosensitivity of antibody responses and radioresistant secondary tetanus antitoxin responses

    International Nuclear Information System (INIS)

    Stoner, R.; Terres, G.; Cottier, H.; Hess, M.

    1976-01-01

    Primary tetanus antitoxin responses were increasingly repressed in mice when gamma radiation doses of 100 to 400 rads were delivered by whole-body exposure prior to immunization with fluid tetanus toxoid (FTT). Nearly normal secondary antitoxin responses were obtained in mice exposed to 600 rads of gamma radiation 4 days after secondary antigenic stimulation with FTT. A rapid transition from radiosensitivity of the antibody-forming system on days 1 to 3 was followed by relative radioresistance on day 4 after the booster injection of toxoid. Studies on lymphoid cellular kinetics in popliteal lymph nodes after injection of 3 H--thymidine ( 3 H--TdR) and incorporation of 3 H--L-histidine into circulating antitoxin were carried out. Analysis of tritium radioactivity in antigen--antibody precipitates of serums 2 hr after injection of the labeled amino acid revealed maximum incorporation into antibody around day 7 after the booster in nonirradiated controls and about day 12, i.e., 8 days after irradiation, in experimental mice. The shift from radiosensitivity to relative radioresistance was attributed to a marked peak of plasma-cell proliferation in the medulla of lymph nodes on day 3. Many medullary plasma cells survived and continued to proliferate after exposure to radiation. Germinal centers were destroyed by radiation within 1 day. Since antibody formation continued after exposure to radiation and after the loss of germinal centers, this supports the view that germinal-center cells were involved more in the generation of memory cells than in antibody synthesis

  3. Role of certain cellular composition in radio-resistant fungi

    International Nuclear Information System (INIS)

    Shahin, A.A.M.; Hammad, A.A.I.; Hazaa, M.M.; Swelim, M.A.; Mohamed, Y.A.

    2007-01-01

    Fifty three fungal isolates of genera Curvularia, Alternaria and Fusarium were isolated from different sources included crops, vegetables, fruits in addition to bread, chicken feed soil and air. Five isolates were selected from each genus according to the difference in the morphological characters and its source. The fifteen isolates were exposed to increasing doses of gamma rays from 0.5 to 10.0 I 10 values when irradiated in saline solution were found to be 1.92,1.25, 1.47,0.47,1.31 and 0.70 respectively while their D 10 values were 2.25, 1.56, 1.70, 1.30, 1.83 and 1.23 as the irradiation process was done in their natural sources. The values of total protein, total lipids and total nucleic acids either RNA or DNA were relatively higher in radio-resistant strains than sensitive ones. Amino acids containing sulfur (cysteine, methionine) or double bonds (histidine) and the percentage of unsaturated fatty acids were also higher in resistant strains than the sensitive ones. Exposing the six selected strains to dose level 4 kGy obviously decreased each of total protein, total amino acids and total nucleic acids especially DNA and the values of decreases were found to be higher in sensitive than the resistant strains. Dose level 12.5 kGy was quiet enough to eliminate the radioresistant fungi from the contaminated food whatever the level of contamination is

  4. Evaluation of the role of genetic factors in human radioresistance

    International Nuclear Information System (INIS)

    Telnov, Vitaliy I.; Sotnik, Natalie V.

    2002-01-01

    This study was focused on evaluation of the role of genetic factors in development of chronic radiation sickness (CRS) due to occupational exposure to external γ -rays. This study was based on results of molecular-genetic studies for 985 nuclear workers of the Mayak Production Association. CRS occurrence was related to the genetic haptoglobin (Hp) system among a number of studied genetic markers. Excess risk of CRS was revealed at similar exposure doses for individuals-carriers of Hp 2-2 (1.96) versus lower risks for carriers of Hp 1-1 and 2-1 (0.64). The contribution of genetic factors to CRS development was implemented in a rather narrow dose range, i.e. it was of a relative nature. A scheme of the relationship of affecting factor and differences in genetic radioresistance was presented in terms of deterministic effects. The obtained data did not confirm the idea that A-bomb survivors were more radioresistant, thus being not representative for radiation risk estimation

  5. Impairing the radioresistance of cancer cells by hydrogenated nanodiamonds.

    Science.gov (United States)

    Grall, Romain; Girard, Hugues; Saad, Lina; Petit, Tristan; Gesset, Céline; Combis-Schlumberger, Mathilde; Paget, Vincent; Delic, Jozo; Arnault, Jean-Charles; Chevillard, Sylvie

    2015-08-01

    Hydrogenated nanodiamonds (H-NDs) exhibit a negative electron affinity that confers a high reactivity with oxygen species and a positive charge in aqueous solutions. It allows electron emission from H-NDs following irradiation by photons and in consequence may enhance the effects of radiation on cancer cells. By using three human radioresistant cancer cell lines, we showed a potentialization of cytotoxicity after a co-exposure to H-NDs and irradiation; an event occurring through the induction of DNA damage and reactive oxygen species. This occurred together with a decrease in cell impedance, the activation of G1/S, an unlocking of G2 cell cycle check-points and early low cell death rate. At later stage of exposure, persistent increases in heterochromatinization, large γ-H2AX foci and β-galactosidase activity were detected providing evidence of cells' entrance into senescence. Similar potentialization was observed with neocarzinostatin (NCS), a radiomimetic drug. This original finding underlines a wide clinical potential of H-NDs to intensify radiation effects on radio-resistant cancer cells. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  6. HIF-1α inhibition by siRNA or chetomin in human malignant glioma cells: effects on hypoxic radioresistance and monitoring via CA9 expression

    Directory of Open Access Journals (Sweden)

    Bache Matthias

    2010-11-01

    Full Text Available Abstract Background Hypoxia induces activation of the HIF-1 pathway and is an essential characteristic of malignant gliomas. Hypoxia has been linked to tumor progression, therapy resistance and poor prognosis. However, little is known about the impact of HIF-1α inhibition on radioresistance of malignant glioma. Methods In this study, we investigated the effects of the inhibition of HIF-1α on cell survival and radiosensitivity in U251MG and U343MG glioma cells, using two different strategies. HIF-1α inhibition was achieved by siRNA targeting of HIF-1α or via chetomin, a disruptor of interactions between HIF-1α and p300. The inhibition of the HIF-1 pathway was monitored by quantitative real-time PCR and Western blot analyses of the expression levels of HIF-1α and CA9. CA9 expression was investigated as a potential indicator of the efficacy of HIF-1 inhibition and the resulting radiosensitivity of malignant glioma cell lines was determined by clonogenic assay after irradiation under normoxic (2-10 Gy or hypoxic (2-15 Gy conditions. Results Although siRNA and chetomin show distinct modes of action, both attenuated the hypoxia-induced radioresistance of malignant glioma cell lines U251MG (DMF10: 1.35 and 1.18 and U343MG (DMF10: 1.78 and 1.48. However, siRNA and chetomin showed diverse effects on radiosensitivity under normoxic conditions in U251MG (DMF10: 0.86 and 1.35 and U343MG (DMF10: 1.33 and 1.02 cells. Conclusions Results from this in vitro study suggest that inhibition of HIF-1α is a promising strategy to sensitize human malignant gliomas to radiotherapy and that CA9 could serve as an indicator of effective HIF-1-related radiosensitization.

  7. Receptor-mediated photo-cytotoxicity: synthesis of a photoactivatable psoralen derivative conjugated to insulin.

    Science.gov (United States)

    Gasparro, F P; Knobler, R M; Yemul, S S; Bisaccia, E; Edelson, R L

    1986-12-15

    4'-Aminomethyl-4,5',8-trimethylpsoralen has been chemically conjugated to insulin using a carbodiimide derivative. The psoralen moiety retains its photochemical reactivity as evidenced by its ability to crosslink DNA after exposure to long wavelength ultraviolet light (UVA, 320-400 nm). This chimeric molecule has been used to selectively kill a population of lymphocytes whose expression of insulin receptors has been stimulated with phytohemagglutinin. Insulin carries the psoralen into the cell via receptor-mediated endocytosis, where it is subsequently activated by exposure to UVA light. The UVA induced activity of AMT-insulin can be blocked by the presence of native insulin. The viability of unstimulated lymphocytes was not affected by AMT-insulin and UVA light. The hybrid insulin-psoralen molecule may be a prototype for a family of phototoxic drugs which can be selectively delivered to subsets of lymphocytes.

  8. Platelet alpha-2 adrenergic receptor-mediated phosphoinositide responses in endogenous depression

    International Nuclear Information System (INIS)

    Mori, Hideki; Koyama, Tsukasa; Yamashita, Itaru

    1991-01-01

    We have previously indicated that epinephrine stimulates phosphoinositide (PI) hydrolysis by activating alpha-2 adrenergic receptors in human platelets. This method involves the measurement of the accumulation of [ 3 H]-inositol-1-phosphate (IP-1) as an index of Pl hydrolysis; lithium is added to inhibit the metabolism of IP-1, thus giving an enhanced signal. In the present study, we assessed the platelet alpha-2 adrenergic receptor-mediated PI responses in samples from 15 unmedicated patients with endogenous depression and 15 age- and sex-matched control subjects. The responses to epinephrine in the depressed patients were significantly higher than those of the controls, whereas the basal values did not differ significantly. These results support the hypothesis that platelet alpha-2 adrenergic receptors may be supersensitive in patients with endogenous depression

  9. Bicarbonate Contributes to GABAA Receptor-Mediated Neuronal Excitation in Surgically-Resected Human Hypothalamic Hamartomas

    Science.gov (United States)

    Do-Young, Kim; Fenoglio, Kristina A.; Kerrigan, John F.; Rho, Jong M.

    2009-01-01

    SUMMARY The role of bicarbonate (HCO3-) in GABAA receptor-mediated depolarization of human hypothalamic hamartoma (HH) neurons was investigated using cellular electrophysiological and calcium imaging techniques. Activation of GABAA receptors with muscimol (30 μM) provoked neuronal excitation in over 70% of large (18-22 μM) HH neurons in HCO3- buffer. Subsequent perfusion of HCO3--free HEPES buffer produced partial suppression of muscimol-induced excitation. Additionally, 53% of large HH neurons under HCO3--free conditions exhibited reduced intracellular calcium accumulation by muscimol. These results suggest that HCO3- efflux through GABAA receptors on a subpopulation of large HH neurons may contribute to membrane depolarization and subsequent activation of L-type calcium channels. PMID:19022626

  10. Gelidium amansii promotes dendritic spine morphology and synaptogenesis, and modulates NMDA receptor-mediated postsynaptic current.

    Science.gov (United States)

    Hannan, Md Abdul; Mohibbullah, Md; Hong, Yong-Ki; Nam, Joo Hyun; Moon, Il Soo

    2014-01-01

    Neurotrophic factors are essential for the differentiation and maturation of developing neurons as well as providing survival support to the mature neurons. Moreover, therapeutically neurotrophic factors are promising to reconstruct partially damaged neuronal networks in neurodegenerative diseases. In the previous study, we reported that the ethanol extract of an edible marine alga, Gelidium amansii (GAE) had shown promising effects in the development and maturation of both axon and dendrites of hippocampal neurons. Here, we demonstrate that in primary culture of hippocampal neurons (1) GAE promotes a significant increase in the number of filopodia and dendritic spines; (2) promotes synaptogenesis; (3) enhances N-methyl-D-aspartic acid (NMDA) receptor recruitment; and (4) modulates NMDA-receptor-mediated postsynaptic current. Taken together these findings that GAE might be involved in both morphological and functional maturation of neurons suggest the possibility that GAE may constitute a promising candidate for novel compounds for the prevention and treatment of neurodegenerative diseases.

  11. Cell adhesion-mediated radioresistance (CAM-RR). Extracellular matrix-dependent improvement of cell survival in human tumor and normal cells in vitro

    International Nuclear Information System (INIS)

    Cordes, N.; Meineke, V.

    2003-01-01

    Background: Cell-extracellular matrix (ECM) contact is thought to have great impact on cellular mechanisms resulting in increased cell survival upon exposure to ionizing radiation. Several human tumor cell lines and normal human fibroblastic cell strains of different origin, all of them expressing the wide-spread and important integrin subunit β1, were irradiated, and clonogenic cell survival, β1-integrin cell surface expression, and adhesive functionality were investigated. Material and Methods: Human tumor cell lines A172 (glioblastoma), PATU8902 (pancreas carcinoma), SKMES1 (lung carcinoma), A549 (lung carcinoma), and IPC298 (melanoma) as well as normal human skin (HSF1) and lung fibroblasts (CCD32) and human keratinocytes (HaCaT) were irradiated with 0-8 Gy. Besides colony formation assays, β1-integrin cell surface expression by flow cytometry and adhesive functionality by adhesion assays were analyzed. Results: All cell lines showed improved clonogenic survival after irradiation in the presence of fibronectin as compared to plastic. Irradiated cells exhibited a significant, dose-dependent increase in β1-integrin cell surface expression following irradiation. As a parameter of the adhesive functionality of the β1-integrin, a radiation-dependent elevation of cell adhesion to fibronectin in comparison with adhesion to plastic was demonstrated. Conclusion: The in vitro cellular radiosensitivity is highly influenced by fibronectin according to the phenomenon of cell adhesion-mediated radioresistance. Additionally, our emerging data question the results of former and current in vitro cytotoxicity studies performed in the absence of an ECM. These findings might also be important for the understanding of malignant transformation, anchorage-independent cell growth, optimization of radiotherapeutic regimes and the prevention of normal tissue side effects on the basis of experimental radiobiological data. (orig.)

  12. Multifunctional pH-Responsive Folate Receptor Mediated Polymer Nanoparticles for Drug Delivery.

    Science.gov (United States)

    Cai, Xiaoqing; Yang, Xiaoye; Wang, Fang; Zhang, Chen; Sun, Deqing; Zhai, Guangxi

    2016-07-01

    Multifunctional pH-responsive folate receptor mediated targeted polymer nanoparticles (TPNps) were developed for docetaxel (DTX) delivery based on poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol)poly (β-amino ester) (P123-PAE) and poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol)-folate (P123-FA) copolymers. The DTX was loaded into the TPNps with a decent drug loading content of 15.02 ± 0.14 wt%. In vitro drug release results showed that the DTX was released from the TPNps at a pH-dependent manner. Tetrazolium dye (MTT) assay revealed that the bland polymer nanoparticles displayed almost nontoxicity at 200 μg/mL concentration. However, the DTX-loaded TPNps showed high anti-tumor activity at low IC50 (0.72 μg/mL) for MCF-7 cells following 48 h incubation. Cellular uptake experiments revealed that the TPNps had higher degree of cellular uptake than nontargeted polymer nanoparticles, indicating that the nanoparticles were internalized into the cells via FA receptor-mediated endocytosis. Moreover, the cellular uptake pathways for the FA grafted polymer were involved in energy-dependent, clathrin-mediated and caveolae-mediated endocytosis. The cell killing effect and cellular uptake of the DTX-TPNps by the MCF-7 cells were all enhanced by about two folds at pH 5.5 when compared with pH 7.4. The TPNps also significantly prolonged the in vivo retention time for the DTX. These results suggest that the biocompatible pH responsive folate-modified polymer nanoparticles present a promising safe nanosystem for intracellular targeted delivery of DTX.

  13. Receptor-mediated uptake of low density lipoprotein stimulates bile acid synthesis by cultured rat hepatocytes

    International Nuclear Information System (INIS)

    Junker, L.H.; Davis, R.A.

    1989-01-01

    The cellular mechanisms responsible for the lipoprotein-mediated stimulation of bile acid synthesis in cultured rat hepatocytes were investigated. Adding 280 micrograms/ml of cholesterol in the form of human or rat low density lipoprotein (LDL) to the culture medium increased bile acid synthesis by 1.8- and 1.6-fold, respectively. As a result of the uptake of LDL, the synthesis of [14C]cholesterol from [2-14C]acetate was decreased and cellular cholesteryl ester mass was increased. Further studies demonstrated that rat apoE-free LDL and apoE-rich high density lipoprotein (HDL) both stimulated bile acid synthesis 1.5-fold, as well as inhibited the formation of [14C]cholesterol from [2-14C]acetate. Reductive methylation of LDL blocked the inhibition of cholesterol synthesis, as well as the stimulation of bile acid synthesis, suggesting that these processes require receptor-mediated uptake. To identify the receptors responsible, competitive binding studies using 125I-labeled apoE-free LDL and 125I-labeled apoE-rich HDL were performed. Both apoE-free LDL and apoE-rich HDL displayed an equal ability to compete for binding of the other, suggesting that a receptor or a group of receptors that recognizes both apolipoproteins is involved. Additional studies show that hepatocytes from cholestyramine-treated rats displayed 2.2- and 3.4-fold increases in the binding of apoE-free LDL and apoE-rich HDL, respectively. These data show for the first time that receptor-mediated uptake of LDL by the liver is intimately linked to processes activating bile acid synthesis

  14. Characterization of GABA/sub A/ receptor-mediated 36chloride uptake in rat brain synaptoneurosomes

    International Nuclear Information System (INIS)

    Luu, M.D.; Morrow, A.L.; Paul, S.M.; Schwartz, R.D.

    1987-01-01

    γ-Aminobutyric acid (GABA) receptor-mediated 36 chloride ( 36 Cl - ) uptake was measured in synaptoneurosomes from rat brain. GABA and GABA agonists stimulated 36 Cl - uptake in a concentration-dependent manner with the following order of potency: Muscimol>GABA>piperidine-4-sulfonic acid (P4S)>4,5,6,7-tetrahydroisoxazolo-[5,4-c]pyridin-3-ol (THIP)=3-aminopropanesulfonic acid (3APS)>>taurine. Both P4S and 3APS behaved as partial agonists, while the GABA/sub B/ agonist, baclofen, was ineffective. The response to muscimol was inhibited by bicuculline and picrotoxin in a mixed competitive/non-competitive manner. Other inhibitors of GABA receptor-opened channels or non-neuronal anion channels such as penicillin, picrate, furosemide and disulfonic acid stilbenes also inhibited the response to muscimol. A regional variation in muscimol-stimulated 36 Cl - uptake was observed; the largest responses were observed in the cerebral cortex, cerebellum and hippocampus, moderate responses were obtained in the striatum and hypothalamus and the smallest response was observed in the pons-medulla. GABA receptor-mediated 36 Cl - uptake was also dependent on the anion present in the media. The muscinol response varied in media containing the following anions: Br - >Cl - ≥NO 3 - >I - ≥SCN - >>C 3 H 5 OO - ≥ClO 4 - >F - , consistent with the relative anion permeability through GABA receptor-gated anion channels and the enhancement of convulsant binding to the GABA receptor-gated Cl - channel. 43 references, 4 figures, 3 tables

  15. The role of receptor-mediated T-cells activation disorders in pulmonary tuberculosis

    Directory of Open Access Journals (Sweden)

    Irina E. Esimova

    2017-01-01

    Full Text Available Aim. To analyze the peculiarities and mechanisms of receptor-mediated T-lymphocytes disorders in different clinical forms of pulmonary tuberculosis.Materials and мethods. The study involved 116 patients with first diagnosed infiltrative and disseminated drug-sensitive and drug-resistant pulmonary tuberculosis. The key stages in receptor-mediated activation of T-lymphocytes, isolated from blood, after their CD3/CD28-induction in vitro with addition of intracellular transport blocker were analyzed. Their immunotyping was carried out with the method of two- and threecolor flow cytofluorometry. The obtained results were statistically analyzed.Results. The breach of extracellular and intracellular stages of T-lymphocytes activation, shown by reduction in total number of CD3- and CD28-positive cells, and CD3+CD28+IL2+, CD3+CD28+IL2–, CD3+NF-kB+, CD3+NFAT2+ lymphocytes, and increase in number of CD3+CTLA4+ cells, was identified with most of their manifestations in disseminated drug-resistant pulmonary tuberculosis. It was shown that the content of CD3+AP-1+ lymphocytes is variable in drug-resistant pulmonary tuberculosis: it increases in the infiltrative form and decreases in the disseminated form.Conclusion. The results showed different mechanisms leading to a deficiency of IL-2-positive lymphocytes and T-lymphocytopenia: from “functional reserve” exhaustion of T-cells in drug-sensitive pulmonary tuberculosis to immunosuppression under the influence of suppressive cytokines (in case of the infiltrative form and inhibitory protein CTLA4 (in case of the disseminated form in drug-resistant pulmonary tuberculosis. 

  16. Effects of sarcosine and N, N-dimethylglycine on NMDA receptor-mediated excitatory field potentials.

    Science.gov (United States)

    Lee, Mei-Yi; Lin, Yi-Ruu; Tu, Yi-Shu; Tseng, Yufeng Jane; Chan, Ming-Huan; Chen, Hwei-Hsien

    2017-02-28

    Sarcosine, a glycine transporter type 1 inhibitor and an N-methyl-D-aspartate (NMDA) receptor co-agonist at the glycine binding site, potentiates NMDA receptor function. Structurally similar to sarcosine, N,N-dimethylglycine (DMG) is also N-methyl glycine-derivative amino acid and commonly used as a dietary supplement. The present study compared the effects of sarcosine and DMG on NMDA receptor-mediated excitatory field potentials (EFPs) in mouse medial prefrontal cortex brain slices using a multi-electrode array system. Glycine, sarcosine and DMG alone did not alter the NMDA receptor-mediated EFPs, but in combination with glutamate, glycine and its N-methyl derivatives significantly increased the frequency and amplitude of EFPs. The enhancing effects of glycine analogs in combination with glutamate on EFPs were remarkably reduced by the glycine binding site antagonist 7-chlorokynurenate (7-CK). However, DMG, but not sarcosine, reduced the frequency and amplitude of EFPs elicited by co-application of glutamate plus glycine. D-cycloserine, a partial agonist at the glycine binding site on NMDA receptors, affected EFPs in a similar manner to DMG. Furthermore, DMG, but not sarcosine, reduced the frequencies and amplitudes of EFPs elicited by glutamate plus D-serine, another endogenous ligand for glycine binding site. These findings suggest that sarcosine acts as a full agonist, yet DMG is a partial agonist at glycine binding site of NMDA receptors. The molecular docking analysis indicated that the interactions of glycine, sarcosine, and DMG to NMDA receptors are highly similar, supporting that the glycine binding site of NMDA receptors is a critical target site for sarcosine and DMG.

  17. 31P NMR spectroscopy and HbO2 cryospectrophotometry in prediction of tumor radioresistance caused by hypoxia.

    Science.gov (United States)

    Rofstad, E K; DeMuth, P; Fenton, B M; Ceckler, T L; Sutherland, R M

    1989-04-01

    The aim of this study was to search for possible relationships between the fraction of radiobiologically hypoxic cells in tumors and their 31P NMR spectral parameters and intracapillary HbO2 saturations. Four different tumor lines, two murine sarcomas (KHT, RIF-1) and two human ovarian carcinoma xenografts (MLS, OWI), were used. When tumor volume increased from about 200 mm3 to about 2000 mm3, hypoxic fraction increased from 12 to 23% for the KHT line, from 0.9 to 1.7% for the RIF-1 line, and from 9 to 28% for the MLS line. The OWI line showed similar hypoxic fractions at 200 (17%) and 2000 mm3 (15%). Tumor bioenergetic status decreased, that is, the inorganic phosphate (Pi) resonance increased and the phosphocreatine (PCr) and nucleoside triphosphate beta (NTP beta) resonances decreased, with increasing tumor volume for the KHT, RIF-1, and MLS lines, whereas the OWI line did not show any changes in the 31P NMR spectral parameters during tumor growth. Similarly, tumor HbO2 saturation status, that is, the fraction of vessels with HbO2 saturation above 30%, decreased with increasing tumor volume for the KHT, RIF-1, and MLS lines, but remained unchanged during tumor growth for the OWI line. Although the data indicated a relationship between hypoxic fraction and tumor bioenergetic status as well as tumor HbO2 saturation status within a specific line during tumor growth, there was no correlation between hypoxic fraction and tumor bioenergetic status or tumor HbO2 saturation status across the four tumor lines. This may have occurred because cell survival time under hypoxic stress as well as fraction of non-clonogenic, but metabolically active hypoxic cells differed among the tumor lines. This indicates that 31P NMR spectroscopy and HbO2 cryospectrophotometry data have to be supplemented with other data to be useful in prediction of tumor radioresistance caused by hypoxia.

  18. The Influence of Receptor-Mediated Interactions on Reaction-Diffusion Mechanisms of Cellular Self-organisation

    Czech Academy of Sciences Publication Activity Database

    Klika, Václav; Baker, R. E.; Headon, D.; Gaffney, E. A.

    2012-01-01

    Roč. 74, č. 4 (2012), s. 935-957 ISSN 0092-8240 Institutional research plan: CEZ:AV0Z20760514 Keywords : reaction-diffusion * receptor-mediated patterning * turing models Subject RIV: BO - Biophysics Impact factor: 2.023, year: 2012 http://www.springerlink.com/content/9713544x6871w4n6/?MUD=MP

  19. Vascular endothelin ET(B) receptor-mediated contraction requires phosphorylation of ERK1/2 proteins

    DEFF Research Database (Denmark)

    Luo, Guogang; Jamali, Roya; Cao, Yong-Xiao

    2006-01-01

    In cardiovascular diseases, endothelin type B (ET(B)) receptors in arterial smooth muscle cells are upregulated. The present study revealed that organ culture of rat mesenteric artery segments enhanced endothelin ET(B) receptor-mediated contraction paralleled with increase in the receptor mRNA an...

  20. AKT-mediated enhanced aerobic glycolysis causes acquired radioresistance by human tumor cells

    International Nuclear Information System (INIS)

    Shimura, Tsutomu; Noma, Naoto; Sano, Yui; Ochiai, Yasushi; Oikawa, Toshiyuki; Fukumoto, Manabu; Kunugita, Naoki

    2014-01-01

    Background and purpose: Cellular radioresistance is a major impediment to effective radiotherapy. Here, we demonstrated that long-term exposure to fractionated radiation conferred acquired radioresistance to tumor cells due to AKT-mediated enhanced aerobic glycolysis. Material and methods: Two human tumor cell lines with acquired radioresistance were established by long-term exposure to fractionated radiation with 0.5 Gy of X-rays. Glucose uptake was inhibited using 2-deoxy-D-glucose, a non-metabolizable glucose analog. Aerobic glycolysis was assessed by measuring lactate concentrations. Cells were then used for assays of ROS generation, survival, and cell death as assessed by annexin V staining. Results: Enhanced aerobic glycolysis was shown by increased glucose transporter Glut1 expression and a high lactate production rate in acquired radioresistant cells compared with parental cells. Inhibiting the AKT pathway using the AKT inhibitor API-2 abrogated these phenomena. Moreover, we found that inhibiting glycolysis with 2-deoxy-D-glucose suppressed acquired tumor cell radioresistance. Conclusions: Long-term fractionated radiation confers acquired radioresistance to tumor cells by AKT-mediated alterations in their glucose metabolic pathway. Thus, tumor cell metabolic pathway is an attractive target to eliminate radioresistant cells and improve radiotherapy efficacy

  1. Molecular biological studies on the human radioresistance and drug resistance

    International Nuclear Information System (INIS)

    Kim, Chang Min; Hong, Weon Seon

    1992-04-01

    We irradiated the MKN45 and PC14 cell lines with 500 rads and also established the adriamycin-resistant and cis-platinum resistant cell line. The genomic DNA and total RNA were extracted and subjected to the Southern and Northern analysis using various probes including heat shock protein 70, MDR1, fos, TGFb etc. The mRNA transcript was increased 1 hour after the irradiation and sustained during the 48 hours and returned to the level of pre-irradiation. No significant change was observed with the drug resistant cell lines at the level of gene dosage. We suggest that the marked increase of the hsp70 transcript is very important finding and is believed to be a good candidate for the modulation of the cellular response to irradiation and the radioresistance. (Author)

  2. A role for survivin in radioresistance of pancreatic cancer cells

    International Nuclear Information System (INIS)

    Asanuma, Koichi; Kobayashi, Daisuke; Furuya, Daisuke; Tsuji, Naoki; Yagihashi, Atsuhito; Watanabe, Naoki

    2002-01-01

    Using gene-transduced pancreatic cancer cells, we examined whether survivin expression is directly involved in regulation of radiosensitivity. Ordinarily radiosensitive MIAPaCa-2 cells transduced with wild-type survivin gene (MS cells) proliferated more rapidly than cells transduced with control vector. MS cells were significantly less radiosensitive than control vector-transduced cells. Radiation-induced activity of caspase-3, but not caspase-7, was significantly inhibited in MS cells. On the other hand, transduction of a dominant-negative mutant survivin gene into radioresistant PANC-1 cells augmented radiosensitivity. Further, the radiation-induced increase in caspase-3 activity was enhanced, indicating that survivin function was truly inhibited. These results indicate that survivin expression directly down-regulates radiosensitivity. (author)

  3. Nuclear medicine markers of tumor oxygenation and radioresistance

    International Nuclear Information System (INIS)

    Chapman, J. Donald; Schneider, R.H.; Stobbe, C.C.; Kim, E.; Engelhardt, E.L.; Coia, L.

    1996-01-01

    Purpose/Objective: The objective of this research project was to synthesize, purify, radiolabel and characterize second-generation nuclear medicine markers of tissue oxygenation with properties superior to iodoazomycin arabinoside (IAZA) and to validate the hypoxia-marking activity of optimal compounds by independent measurements of tumor oxygenation and tumor radioresistance. Materials and Methods: Six hypoxic markers of the iodoazomycin nucleoside class with water solubilities greater than IAZA were synthesized by published procedures. The markers were purified, chemically characterized and labeled with Iodine-125 or Iodine-131. Absolute rates of marker ligation to the macromolecules of hypoxic EMT-6 tumor cells in vitro were determined as a function of marker concentration and used to establish relative marker effectiveness. Hypoxic marking activity in tumors was determined from tumor/blood (T/B) and tumor/muscle (T/M) ratios of radiolabelled marker in EMT-6 tumor-bearing C.B17/Icr scid mice. The optimal marker was administered to R3327-H and R3327-AT tumor-bearing Fischer X Copenhagen rats for estimates of tumor oxygenation by T/B and T/M ratios. Oxygen distributions in the same tumors were obtained with the Eppendorf pO 2 Histograph. The radioresistance of individual tumors was determined from in vitro plating efficiencies of cells released from tumors which had been irradiated in vivo with 20 Gy Cs-137 γ-rays. Results: Of the six iodinated azomycin nucleosides investigated, five were novel markers and all had water solubilities higher than IAZA. Iodinated azomycin xylopyranoside (β-D-IAZXP) was selected as the optimal marker of this class since it 1) exhibited the highest absolute rate of ligation to hypoxic tumor cells in vitro, 2) had the fastest plasma clearance rate in tumor-bearing mice and 3) yielded high T/B ratios in both the mouse and rat tumor models employed in this study. Planar nuclear medicine images of (I-131) β-D-IAZXP in tumor-bearing rats

  4. Superoxide dismutase in radioresistant PC-3 human prostate carcinoma cells

    International Nuclear Information System (INIS)

    Prokopovic, J.; Adzic M; Niciforovic, A.; Vucic, V.; Zaric, B.; Radojcic, M. B.

    2006-01-01

    The molecular mechanism of gamma-ionizing radiation (IR) resistance of human prostate cancer cells PC-3 is not known. Since low-LET-IR effects are primarily achieved through generation of reactive oxygen species (ROS), IR-induced expression of ROS-metabolizing antioxidant enzymes, Mn- and CuZn-superoxide dismutase (Mn- and CuZnSOD) and catalase (CAT), and their upstream regulator transcription factor NFκB was followed. Significant elevation of both SODs was found in cells irradiated with 10- and 20 Gy, while CAT and NFκB expression was unchanged. Since, such conditions lead to accumulation of H 2 O 2 , it is concluded that radioresistance of PC-3 cells may emerge from positive feed-forward vicious circle established between H 2 O 2 activation of NFκB and elevated MnSOD activity. (author)

  5. Radiobiological effects of tritiated water short-term exposure on V79 clonogenic cell survival

    DEFF Research Database (Denmark)

    Siragusa, Mattia; Fredericia, Nina Pil Møntegaard; Jensen, Mikael

    2018-01-01

    We set out to improve the accuracy of absorbed dose calculations for in-vitro measurements of the Relative Biological Effectiveness (RBE) of tritiated water (HTO) for the clonogenic cell survival assay, also considering the influence of the end-of-track Linear Energy Transfer (LET) of low-energy...... in suspension are usually comparable to those for adherent cells. RBEs calculated at the 10% survival fraction through the use of the average energy are almost similar to those obtained with the beta-spectrum. For adherent cells, an RBE of 1.6 was found when HTO cell survival curves were compared to acute γ...

  6. An in vitro clonogenic assay to assess radiation damage in rat CNS glial progenitor cells

    International Nuclear Information System (INIS)

    Maazen, R.W.M. van der; Verhagen, I.; Kogel, A.J. van der

    1990-01-01

    Normal glial progenitor cells can be isolated from the rat central nervous system (CNS) and cultured in vitro on a monolayer of type-1 astrocytes. These monolayers are able to support and stimulate explanted glial progenitor cells to proliferate. Employing these in vitro interactions of specific glial cell types, an in vivo-in vitro clonogenic assay has been developed. This method offers the possibility to study the intrinsic radiosensitivity, repair and regeneration of glial progenitor cells after in vitro or in vivo irradiation. (author)

  7. Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

    International Nuclear Information System (INIS)

    Xu, Yuan; Cardell, Lars-Olaf

    2014-01-01

    Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B 2 receptor agonist) and des-Arg 9 -bradykinin- (selective B 1 receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE 2 . The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg 9 -bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B 2 receptors, but not those on B 1 . Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in some patients with asthma

  8. Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Yuan, E-mail: yuan.xu@ki.se; Cardell, Lars-Olaf

    2014-02-15

    Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B{sub 2} receptor agonist) and des-Arg{sup 9}-bradykinin- (selective B{sub 1} receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE{sub 2}. The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg{sup 9}-bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B{sub 2} receptors, but not those on B{sub 1}. Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in

  9. Demonstration and suppression of a radioresistant host-versus-graft reaction

    International Nuclear Information System (INIS)

    Thierfelder, S.; Roessler, R. v.; Ruppelt, W.

    1975-01-01

    The possibilities of suppressing and measuring a radioresistant host-versus-graft reaction are described. According to the authors, the host-versus-graft immune reaction may outlast whole-body irradiation at high doses. (BSC/AK) [de

  10. Enhanced NMDA receptor-mediated intracellular calcium signaling in magnocellular neurosecretory neurons in heart failure rats.

    Science.gov (United States)

    Stern, Javier E; Potapenko, Evgeniy S

    2013-08-15

    An enhanced glutamate excitatory function within the hypothalamic supraoptic and paraventricluar nuclei is known to contribute to increased neurosecretory and presympathetic neuronal activity, and hence, neurohumoral activation, during heart failure (HF). Still, the precise mechanisms underlying enhanced glutamate-driven neuronal activity in HF remain to be elucidated. Here, we performed simultaneous electrophysiology and fast confocal Ca²⁺ imaging to determine whether altered N-methyl-d-aspartate (NMDA) receptor-mediated changes in intracellular Ca²⁺ levels (NMDA-ΔCa²⁺) occurred in hypothalamic magnocellular neurosecretory cells (MNCs) in HF rats. We found that activation of NMDA receptors resulted in a larger ΔCa²⁺ in MNCs from HF when compared with sham rats. The enhanced NMDA-ΔCa²⁺ was neither dependent on the magnitude of the NMDA-mediated current (voltage clamp) nor on the degree of membrane depolarization or firing activity evoked by NMDA (current clamp). Differently from NMDA receptor activation, firing activity evoked by direct membrane depolarization resulted in similar changes in intracellular Ca²⁺ in sham and HF rats. Taken together, our results support a relatively selective alteration of intracellular Ca²⁺ homeostasis and signaling following activation of NMDA receptors in MNCs during HF. The downstream functional consequences of such altered ΔCa²⁺ signaling during HF are discussed.

  11. Novel isoforms of the TFIID subunit TAF4 modulate nuclear receptor-mediated transcriptional activity

    International Nuclear Information System (INIS)

    Brunkhorst, Adrian; Neuman, Toomas; Hall, Anita; Arenas, Ernest; Bartfai, Tamas; Hermanson, Ola; Metsis, Madis

    2004-01-01

    The transcription factor TFIID consists of TATA-binding protein (TBP) and TBP-associated factors (TAFs). TAFs are essential for modulation of transcriptional activity but the regulation of TAFs is complex and many important aspects remain unclear. In this study, we have identified and characterized five novel truncated forms of the TFIID subunit TAF4 (TAF II 135). Analysis of the mouse gene structure revealed that all truncations were the results of alternative splicing and resulted in the loss of domains or parts of domains implicated in TAF4 functional interactions. Results from transcriptional assays showed that several of the TAF4 isoforms exerted dominant negative effects on TAF4 activity in nuclear receptor-mediated transcriptional activation. In addition, alternative TAF4 isoforms could be detected in specific cell types. Our results indicate an additional level of complexity in TAF4-mediated regulation of transcription and suggest context-specific roles for these new TAF4 isoforms in transcriptional regulation in vivo

  12. Plant cell surface receptor-mediated signaling - a common theme amid diversity.

    Science.gov (United States)

    He, Yunxia; Zhou, Jinggeng; Shan, Libo; Meng, Xiangzong

    2018-01-29

    Sessile plants employ a diverse array of plasma membrane-bound receptors to perceive endogenous and exogenous signals for regulation of plant growth, development and immunity. These cell surface receptors include receptor-like kinases (RLKs) and receptor-like proteins (RLPs) that harbor different extracellular domains for perception of distinct ligands. Several RLK and RLP signaling pathways converge at the somatic embryogenesis receptor kinases (SERKs), which function as shared co-receptors. A repertoire of receptor-like cytoplasmic kinases (RLCKs) associate with the receptor complexes to relay intracellular signaling. Downstream of the receptor complexes, mitogen-activated protein kinase (MAPK) cascades are among the key signaling modules at which the signals converge, and these cascades regulate diverse cellular and physiological responses through phosphorylation of different downstream substrates. In this Review, we summarize the emerging common theme that underlies cell surface receptor-mediated signaling pathways in Arabidopsis thaliana : the dynamic association of RLKs and RLPs with specific co-receptors and RLCKs for signal transduction. We further discuss how signaling specificities are maintained through modules at which signals converge, with a focus on SERK-mediated receptor signaling. © 2018. Published by The Company of Biologists Ltd.

  13. Muscarinic receptor-mediated inositol tetrakisphosphate response in bovine adrenal chromaffin cells

    International Nuclear Information System (INIS)

    Sanborn, B.B.; Schneider, A.S.

    1990-01-01

    Inositol trisphosphate (IP 3 ), a product of the phosphoinositide cycle, mobilizes intracellular Ca 2+ in many cell types. New evidence suggests that inositol tetrakisphosphate (IP 4 ), an IP 3 derivative, may act as another second messenger to further alter calcium homeostasis. However, the function and mechanism of action of IP 4 are presently unresolved. We now report evidence of muscarinic receptor-mediated accumulation of IP 4 in bovine adrenal chromaffin cells, a classic neurosecretory system in which calcium movements have been well studied. Muscarine stimulated an increase in [ 3 H]IP 4 and [ 3 H]IP 3 accumulation in chromaffin cells and this effect was completely blocked by atropine. [ 3 H]IP 4 accumulation was detectable within 15 sec, increased to a maximum by 30 sec and thereafter declined. 2,3-diphosphoglycerate, an inhibitor of IP 3 and IP 4 hydrolysis, enhanced accumulation of these inositol polyphosphates. The results provide the first evidence of a rapid inositol tetrakisphosphate response in adrenal chromaffin cells, which should facilitate the future resolution of the relationship between IP 4 and calcium homeostasis

  14. Electroacupuncture Potentiates Cannabinoid Receptor-Mediated Descending Inhibitory Control in a Mouse Model of Knee Osteoarthritis

    Directory of Open Access Journals (Sweden)

    Xiao-Cui Yuan

    2018-04-01

    Full Text Available Knee osteoarthritis (KOA is a highly prevalent, chronic joint disorder, which can lead to chronic pain. Although electroacupuncture (EA is effective in relieving chronic pain in the clinic, the involved mechanisms remain unclear. Reduced diffuse noxius inhibitory controls (DNIC function is associated with chronic pain and may be related to the action of endocannabinoids. In the present study, we determined whether EA may potentiate cannabinoid receptor-mediated descending inhibitory control and inhibit chronic pain in a mouse model of KOA. We found that the optimized parameters of EA inhibiting chronic pain were the low frequency and high intensity (2 Hz + 1 mA. EA reversed the reduced expression of CB1 receptors and the 2-arachidonoylglycerol (2-AG level in the midbrain in chronic pain. Microinjection of the CB1 receptor antagonist AM251 into the ventrolateral periaqueductal gray (vlPAG can reversed the EA effect on pain hypersensitivity and DNIC function. In addition, CB1 receptors on GABAergic but not glutamatergic neurons are involved in the EA effect on DNIC function and descending inhibitory control of 5-HT in the medulla, thus inhibiting chronic pain. Our data suggest that endocannabinoid (2-AG-CB1R-GABA-5-HT may be a novel signaling pathway involved in the effect of EA improving DNIC function and inhibiting chronic pain.

  15. Receptor-mediated endocytosis of lysozyme in renal proximal tubules of the frog Rana temporaria

    Directory of Open Access Journals (Sweden)

    E.V. Seliverstova

    2015-04-01

    Full Text Available The mechanism of protein reabsorption in the kidney of lower vertebrates remains insufficiently investigated in spite of raising interest to the amphibian and fish kidneys as a useful model for physiological and pathophysiological examinations. In the present study, we examined the renal tubular uptake and the internalization rote of lysozyme after its intravenous injection in the wintering frog Rana temporaria using immunohisto- and immunocytochemistry and specific markers for some endocytic compartments. The distinct expression of megalin and cubilin in the proximal tubule cells of lysozyme-injected frogs was revealed whereas kidney tissue of control animals showed no positive immunoreactivity. Lysozyme was detected in the apical endocytic compartment of the tubular cells and colocalized with clathrin 10 min after injection. After 20 min, lysozyme was located in the subapical compartment negative to clathrin (endosomes, and intracellular trafficking of lysozyme was coincided with the distribution of megalin and cubilin. However, internalized protein was retained in the endosomes and did not reach lysosomes within 30 min after treatment that may indicate the inhibition of intracellular trafficking in hibernating frogs. For the first time, we provided the evidence that lysozyme is filtered through the glomeruli and absorbed by receptor-mediated clathrin-dependent endocytosis in the frog proximal tubule cells. Thus, the protein uptake in the amphibian mesonephros is mediated by megalin and cubilin that confirms a critical role of endocytic receptors in the renal reabsorption of proteins in amphibians as in mammals.

  16. Killing of intracellular Mycobacterium tuberculosis by receptor-mediated drug delivery

    International Nuclear Information System (INIS)

    Majumdar, S.; Basu, S.K.

    1991-01-01

    p-Aminosalicylic acid (PAS) conjugated to maleylated bovine serum albumin (MBSA) was taken up efficiently through high-affinity MBSA-binding sites on macrophages. Binding of the radiolabeled conjugate to cultured mouse peritoneal macrophages at 4 degrees C was competed for by MBSA but not by PAS. At 37 degrees C, the radiolabeled conjugate was rapidly degraded by the macrophages, leading to release of acid-soluble degradation products in the medium. The drug conjugate was nearly 100 times as effective as free PAS in killing the intracellular mycobacteria in mouse peritoneal macrophages infected in culture with Mycobacterium tuberculosis. The killing of intracellular mycobacteria mediated by the drug conjugate was effectively prevented by simultaneous addition of excess MBSA (100 micrograms/ml) or chloroquine (3 microM) to the medium, whereas these agents did not affect the microbicidal action of free PAS. These results suggest that (i) uptake of the PAS-MBSA conjugate was mediated by cell surface receptors on macrophages which recognize MBSA and (ii) lysosomal hydrolysis of the internalized conjugate resulted in intracellular release of a pharmacologically active form of the drug, which led to selective killing of the M. tuberculosis harbored by mouse macrophages infected in culture. This receptor-mediated modality of delivering drugs to macrophages could contribute to greater therapeutic efficacy and minimization of toxic side effects in the management of tuberculosis and other intracellular mycobacterial infections

  17. GABA-A Receptors Mediate Tonic Inhibition and Neurosteroid Sensitivity in the Brain.

    Science.gov (United States)

    Reddy, Doodipala Samba

    2018-01-01

    Neurosteroids like allopregnanolone (AP) are positive allosteric modulators of synaptic and extrasynaptic GABA-A receptors. AP and related neurosteroids exhibit a greater potency for δ-containing extrasynaptic receptors. The δGABA-A receptors, which are expressed extrasynaptically in the dentate gyrus and other regions, contribute to tonic inhibition, promoting network shunting as well as reducing seizure susceptibility. Levels of endogenous neurosteroids fluctuate with ovarian cycle. Natural and synthetic neurosteroids maximally potentiate tonic inhibition in the hippocampus and provide robust protection against a variety of limbic seizures and status epilepticus. Recently, a consensus neurosteroid pharmacophore model has been proposed at extrasynaptic δGABA-A receptors based on structure-activity relationship for functional activation of tonic currents and seizure protection. Aside from anticonvulsant actions, neurosteroids have been found to be powerful anxiolytic and anesthetic agents. Neurosteroids and Zn 2+ have preferential affinity for δ-containing receptors. Thus, Zn 2+ can prevent neurosteroid activation of extrasynaptic δGABA-A receptor-mediated tonic inhibition. Recently, we demonstrated that Zn 2+ selectively inhibits extrasynaptic δGABA-A receptors and thereby fully prevents AP activation of tonic inhibition and seizure protection. We confirmed that neurosteroids exhibit greater sensitivity at extrasynaptic δGABA-A receptors. Overall, extrasynaptic GABA-A receptors are primary mediators of tonic inhibition in the brain and play a key role in the pathophysiology of epilepsy and other neurological disorders. © 2018 Elsevier Inc. All rights reserved.

  18. Beta receptor-mediated modulation of the late positive potential in humans.

    Science.gov (United States)

    de Rover, Mischa; Brown, Stephen B R E; Boot, Nathalie; Hajcak, Greg; van Noorden, Martijn S; van der Wee, Nic J A; Nieuwenhuis, Sander

    2012-02-01

    Electrophysiological studies have identified a scalp potential, the late positive potential (LPP), which is modulated by the emotional intensity of observed stimuli. Previous work has shown that the LPP reflects the modulation of activity in extrastriate visual cortical structures, but little is known about the source of that modulation. The present study investigated whether beta-adrenergic receptors are involved in the generation of the LPP. We used a genetic individual differences approach (experiment 1) and a pharmacological manipulation (experiment 2) to test the hypothesis that the LPP is modulated by the activation of β-adrenergic receptors. In experiment 1, we found that LPP amplitude depends on allelic variation in the β1-receptor gene polymorphism. In experiment 2, we found that LPP amplitude was modulated by the β-blocker propranolol in a direction dependent on subjects' level of trait anxiety: In participants with lower trait anxiety, propranolol led to a (nonsignificant) decrease in the LPP modulation; in participants with higher trait anxiety, propranolol increased the emotion-related LPP modulation. These results provide initial support for the hypothesis that the LPP reflects the downstream effects, in visual cortical areas, of β-receptor-mediated activation of the amygdala.

  19. The effect of vanadate on receptor-mediated endocytosis of asialoorosomucoid in rat liver parenchymal cells

    International Nuclear Information System (INIS)

    Kindberg, G.M.; Gudmundsen, O.; Berg, T.

    1990-01-01

    Vanadate is a phosphate analogue that inhibits enzymes involved in phosphate release and transfer reactions. Since such reactions may play important roles in endocytosis, we studied the effects of vanadate on various steps in receptor-mediated endocytosis of asialoorosomucoid labeled with 125I-tyramine-cellobiose (125I-TC-AOM). The labeled degradation products formed from 125I-TC-AOM are trapped in the lysosomes and may therefore serve as lysosomal markers in subcellular fractionation studies. Vanadate reduced the amount of active surface asialoglycoprotein receptors approximately 70%, but had no effect on the rate of internalization and retroendocytosis of ligand. The amount of surface asialoglycoprotein receptors can be reduced by lowering the incubation temperature gradually from 37 to 15 degrees C; vanadate affected only the temperature--sensitive receptors. Vanadate inhibited degradation of 125I-TC-AOM 70-80%. Degradation was much more sensitive to vanadate than binding; half-maximal effects were seen at approximately 1 mM vanadate for binding and approximately 0.1 mM vanadate for degradation. By subcellular fractionation in sucrose and Nycodenz gradients, it was shown that vanadate completely prevented the transfer of 125I-TC-AOM from endosomes to lysosomes. Therefore, the inhibition of degradation by vanadate was indirect; in the presence of vanadate, ligand did not gain access to the lysosomes. The limited degradation in the presence of vanadate took place in a prelysosomal compartment. Vanadate did not affect cell viability and ATP content

  20. Comparative radioresistance of chronically irradiated populations of Arabidopsis thaliana (L.) Heynh

    International Nuclear Information System (INIS)

    Dineva, S.B.; Abramov, V.I.; Shevchenko, V.A.

    1994-01-01

    The radioresistance of seeds of populations of Arabidopsis thaliana (L.) Heynh. growing for 5 years in the regions with different levels of radioactive contamination within 30 km zone of Chernobyl NPP was studied. The analysis of comparative radiosensitivity by root test was performed. It has been shown that plants from arabidopsis population growing under chronic irradiation did not gain an increased radioresistance. The data obtained shown that they are more radiosensitive

  1. Modification of γ-irradiation damaging effect on the seeds of radiosensitive and radioresistant plants

    International Nuclear Information System (INIS)

    Kaplan, I.S.; Tikhomirov, F.A.; Khvostova, V.V.; AN SSSR, Novosibirsk. Inst. Tsitologii i Genetiki)

    1975-01-01

    Low and high temperature treatment of seeds during irradiation has shown to result in a decrease of the general deleterious effect of radiation in both relatively radiosensitive (bean) and radioresistant (flax, mustard) species. The protective effect of the treatment is supposed to be due to its influence on short-half-life radicals and this is supportted by experiments with storage of irradiated seeds. The treatment allows to obtain high mutation frequencies in both radiosensitive and radioresistant plants

  2. A pp32-retinoblastoma protein complex modulates androgen receptor-mediated transcription and associates with components of the splicing machinery

    International Nuclear Information System (INIS)

    Adegbola, Onikepe; Pasternack, Gary R.

    2005-01-01

    We have previously shown pp32 and the retinoblastoma protein interact. pp32 and the retinoblastoma protein are nuclear receptor transcriptional coregulators: the retinoblastoma protein is a coactivator for androgen receptor, the major regulator of prostate cancer growth, while pp32, which is highly expressed in prostate cancer, is a corepressor of the estrogen receptor. We now show pp32 increases androgen receptor-mediated transcription and the retinoblastoma protein modulates this activity. Using affinity purification and mass spectrometry, we identify members of the pp32-retinoblastoma protein complex as PSF and nonO/p54nrb, proteins implicated in coordinate regulation of nuclear receptor-mediated transcription and splicing. We show that the pp32-retinoblastoma protein complex is modulated during TPA-induced K562 differentiation. Present evidence suggests that nuclear receptors assemble multiprotein complexes to coordinately regulate transcription and mRNA processing. Our results suggest that pp32 and the retinoblastoma protein may be part of a multiprotein complex that coordinately regulates nuclear receptor-mediated transcription and mRNA processing

  3. Influence of ER leak on resting cytoplasmic Ca2+ and receptor-mediated Ca2+ signalling in human macrophage.

    Science.gov (United States)

    Layhadi, Janice A; Fountain, Samuel J

    2017-06-03

    Mechanisms controlling endoplasmic reticulum (ER) Ca 2+ homeostasis are important regulators of resting cytoplasmic Ca 2+ concentration ([Ca 2+ ] cyto ) and receptor-mediated Ca 2+ signalling. Here we investigate channels responsible for ER Ca 2+ leak in THP-1 macrophage and human primary macrophage. In the absence of extracellular Ca 2+ we employ ionomycin action at the plasma membrane to stimulate ER Ca 2+ leak. Under these conditions ionomycin elevates [Ca 2+ ] cyto revealing a Ca 2+ leak response which is abolished by thapsigargin. IP 3 receptors (Xestospongin C, 2-APB), ryanodine receptors (dantrolene), and translocon (anisomycin) inhibition facilitated ER Ca 2+ leak in model macrophage, with translocon inhibition also reducing resting [Ca 2+ ] cyto . In primary macrophage, translocon inhibition blocks Ca 2+ leak but does not influence resting [Ca 2+ ] cyto . We identify a role for translocon-mediated ER Ca 2+ leak in receptor-mediated Ca 2+ signalling in both model and primary human macrophage, whereby the Ca 2+ response to ADP (P2Y receptor agonist) is augmented following anisomycin treatment. In conclusion, we demonstrate a role of ER Ca 2+ leak via the translocon in controlling resting cytoplasmic Ca 2+ in model macrophage and receptor-mediated Ca 2+ signalling in model macrophage and primary macrophage. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Increase of corneal epithelium cell radioresistance during regeneration

    International Nuclear Information System (INIS)

    Popova, M.F.; Bulyakova, N.V.; Azarova, V.S.

    1985-01-01

    A comparative study of the radiosensitivity of the normal and regenerating cornea epithelium of C 57 Bl mice was performed on the cellular level, the duration of the cell cycle being taken into account. Criteria of radiation injuries were the number of chromosome aberrations, mitotic index and duration of mitotic block. The anterior part of the head was irradiated singly with 1.75, 3.5 or 7.0 Gy and also repeatedly 3.5 + 3.5 at a 24-hours interval. The corneas were fixed 2, 4, 6, 12, 24, 48, 72 and 96 hours after irradiation. In all cases of irradiated mice the regenerating epithelium showed a shorter mitotic block and significantly lower cytogenetic injury as compared with the controls. Effects of fractionated irradiation were only shown in the regenerating epithelium. The results obtained indicate that regenerating epithelium cells of the cornea are significantly more radioresistant than normal epithelium due to activation of post-radiation recovery, and also, possibly, due to an increase in the content of endogenous radioprotectors. (author)

  5. Radiation-induced-radioresistance: mechanisms and modification radioprotection

    International Nuclear Information System (INIS)

    Bala, Madhu

    2005-01-01

    Full text: The term radiation-induced-radioresistance (RIR) has been chosen to explain a particular class of resistance against lethal doses of radiation, which is transient and is induced by pre-exposure to low doses of radiation. This is a genetically governed phenomenon and is different from adaptation which in one of its several senses, refers to evolutionary transformation into new behavioural patterns. RIR is understood to be an evolutionarily conserved fundamental cellular defense mechanism. Small doses of radiation acting as stress stimuli evoke a concerted action of molecular pathways which help the organism to cope-up with the genotoxic effects of lethal doses of radiation given subsequently. Such molecular pathways are a complex interplay of genetic and biochemical entities and are increasingly becoming the focus of research world over. Most of our information on this subject has been gathered from prokaryotes, simpler eukaryotes, human cells and the epidemiological studies. A number of genes such as GADD 45, CDKN1A, PBP74, DIR1, DDR have been reported by to participate in RIR. However, till date, the mechanism of RIR remain poorly understood. In this deliberation some of our findings on mechanisms of RIR will be presented. Further, modification of RIR by a metabolic modifier, presently under clinical investigations for tumor radiotherapy, will also be presented

  6. Inactivation of carbenicillin by some radioresistant mutant strains

    International Nuclear Information System (INIS)

    Zahiera, T.S.; Mahmoud, M.I.; Bashandy, A.A.

    1990-01-01

    Sensitivity test of five bacterial species to carbenicillin was performed microbiologically. The bacterial species were previously isolated from high level radiation environment. All the studied species could either highly decrease the antibiotic activity or even inactivate it completely. Detailed study of the inactivation of carbenicillin by the radioresistant mutant strains B. Laterosporus, B. firmus and M. roseus was performed, in the present study. Using high performace liquid chromatography technique. The gram-positive m. roseus mutant strain seemed to be the most active mutant in degrading the antibiotic. The left over of the antibiotic attained a value of 9% of the original amount after 14 day incubation of the antibiotic with this mutant strain, while the value of the left over reached 36% and 32% after the same period of incubation with the mutants B. laterosporus and B. firmus respectively. In the case of bacillus species, the degradation of the antibiotic started at the same moment when it was added to the bacterial cultures. This fact may indicate that the inactivation of the studied antibiotic by these bacillus species was due to extracellular enzymes extracted rapidly in the surrounding medium. In the case of M. roseus the inactivation process started later. after the addition of the antibiotic to the mutant culture

  7. Intratumoral heterogeneity as a confounding factor in clonogenic assays for tumour radioresponsiveness

    International Nuclear Information System (INIS)

    Britten, R.A.; Evans, A.J.; Allalunis-Turner, M.J.; Franko, A.J.; Pearcey, R.G.

    1996-01-01

    The level of intra-tumoral heterogeneity of cellular radiosensitivity within primary cultures of three carcinomas of the cervix has been established. All three cultures contained clones that varied by as much as 3-fold in their clinically relevant radiosensitivity (SF 2 ). The level of intra-tumoral heterogeneity observed in these cervical tumour cultures was sufficient to be a major confounding factor to the use of pre-treatment assessments of radiosensitivity to predict for clinical radioresponsiveness. Mathematical modeling of the relative elimination of the tumour clones during fractionated radiotherapy indicates that, in two of the three biopsy samples, the use of pre-treatment derived SF 2 values from the heterogeneous tumour sample would significantly overestimate radioresponsiveness. We conclude that assays of cellular radiosensitivity that identify the radiosensitivity of the most radioresistant clones and measure their relative abundance could potentially increase the effectiveness of SF 2 values as a predictive marker of radioresponsiveness

  8. Receptor-mediated mechanism for the transport of prolactin from blood to cerebrospinal fluid

    International Nuclear Information System (INIS)

    Walsh, R.J.; Slaby, F.J.; Posner, B.I.

    1987-01-01

    Prolactin (PRL) interacts with areas of the central nervous system which reside behind the blood-brain barrier. While vascular PRL does not cross this barrier, it is readily accessible to the cerebrospinal fluid (CSF) from which it may gain access to the PRL-responsive areas of the brain. Studies were undertaken to characterize the mechanism responsible for the translocation of PRL from blood to CSF. Rats were given external jugular vein injections of [ 125 -I]iodo-PRL in the presence or absence of an excess of unlabeled ovine PRL (oPRL), human GH, bovine GH, or porcine insulin. CSF and choroid plexus were removed 60 min later. CSF samples were electrophoresed on sodium dodecyl sulfate-polyacrylamide slab gels and resultant autoradiographs were analyzed with quantitative microdensitometry. The data revealed that unlabeled lactogenic hormones, viz. oPRL and human GH, caused a statistically significant inhibition of [ 125 I]iodo-PRL transport from blood to CSF. In contrast, nonlactogenic hormones, viz bovine GH and insulin, had no effect on [ 125 I]iodo-PRL transport into the CSF. An identical pattern of competition was observed in the binding of hormone to the choroid plexus. Furthermore, vascular injections of [ 125 I]iodo-PRL administered with a range of concentrations of unlabeled oPRL revealed a dose-response inhibition in the transport of [ 125 I]iodo-PRL from blood to CSF. The study demonstrates that PRL enters the CSF by a specific, PRL receptor-mediated transport mechanism. The data is consistent with the hypothesis that the transport mechanism resides at the choroid plexus. The existence of this transport mechanism reflects the importance of the cerebroventricular system in PRL-brain interactions

  9. Decreased α1-adrenergic receptor-mediated inositide hydrolysis in neurons from hypertensive rat brain

    International Nuclear Information System (INIS)

    Feldstein, J.B.; Gonzales, R.A.; Baker, S.P.; Sumners, C.; Crews, F.T.; Raizada, M.K.

    1986-01-01

    The expression of α 1 -adrenergic receptors and norepinephrine (NE)-stimulated hydrolysis of inositol phospholipid has been studied in neuronal cultures from the brains of normotensive (Wistar-Kyoto, WKY) and spontaneously hypertensive (SH) rats. Binding of 125 I-1-[β-(4-hydroxyphenyl)-ethyl-aminomethyl] tetralone (HEAT) to neuronal membranes was 68-85% specific and was rapid. Competition-inhibition experiments with various agonists and antagonists suggested that 125 I-HEAT bound selectively to α 1 -adrenergic receptors. Specific binding of 125 I-HEAT to neuronal membranes from SH rat brain cultures was 30-45% higher compared with binding in WKY normotensive controls. This increase was attributed to an increase in the number of α 1 -adrenergic receptors on SH rat brain neurons. Incubation of neuronal cultures of rat brain from both strains with NE resulted in a concentration-dependent stimulation of release of inositol phosphates, although neurons from SH rat brains were 40% less responsive compared with WKY controls. The decrease in responsiveness of SH rat brain neurons to NE, even though the α 1 -adrenergic receptors are increased, does not appear to be due to a general defect in membrane receptors and postreceptor signal transduction mechanisms. This is because neither the number of muscarinic-cholinergic receptors nor the carbachol-stimulated release of inositol phosphates is different in neuronal cultures from the brains of SH rats compared with neuronal cultures from the brains of WKY rats. These observations suggest that the increased expression of α 1 -adrenergic receptors does not parallel the receptor-mediated inositol phosphate hydrolysis in neuronal cultures from SH rat brain

  10. Milrinone attenuates thromboxane receptor-mediated hyperresponsiveness in hypoxic pulmonary arterial myocytes.

    Science.gov (United States)

    Santhosh, K T; Elkhateeb, O; Nolette, N; Outbih, O; Halayko, A J; Dakshinamurti, S

    2011-07-01

    Neonatal pulmonary hypertension (PPHN) is characterized by pulmonary vasoconstriction, due in part to dysregulation of the thromboxane prostanoid (TP) receptor. Hypoxia induces TP receptor-mediated hyperresponsiveness, whereas serine phosphorylation mediates desensitization of TP receptors. We hypothesized that prostacyclin (IP) receptor activity induces TP receptor phosphorylation and decreases ligand affinity; that TP receptor sensitization in hypoxic myocytes is due to IP receptor inactivation; and that this would be reversible by the cAMP-specific phosphodiesterase inhibitor milrinone. We examined functional regulation of TP receptors by serine phosphorylation and effects of IP receptor stimulation and protein kinase A (PKA) activity on TP receptor sensitivity in myocytes from neonatal porcine resistance pulmonary arteries after 72 h hypoxia in vitro. Ca(2+) response curves to U46619 (TP receptor agonist) were determined in hypoxic and normoxic myocytes incubated with or without iloprost (IP receptor agonist), forskolin (adenylyl cyclase activator), H8 (PKA inhibitor) or milrinone. TP and IP receptor saturation binding kinetics were measured in presence of iloprost or 8-bromo-cAMP. Ligand affinity for TP receptors was normalized in vitro by IP receptor signalling intermediates. However, IP receptor affinity was compromised in hypoxic myocytes, decreasing cAMP production. Milrinone normalized TP receptor sensitivity in hypoxic myocytes by restoring PKA-mediated regulatory TP receptor phosphorylation. TP receptor sensitivity and EC(50) for TP receptor agonists was regulated by PKA, as TP receptor serine phosphorylation by PKA down-regulated Ca(2+) mobilization. Hypoxia decreased IP receptor activity and cAMP generation, inducing TP receptor hyperresponsiveness, which was reversed by milrinone. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

  11. PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

    Science.gov (United States)

    Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang

    2015-02-01

    The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

  12. Receptor-mediated endocytosis generates nanomechanical force reflective of ligand identity and cellular property.

    Science.gov (United States)

    Zhang, Xiao; Ren, Juan; Wang, Jingren; Li, Shixie; Zou, Qingze; Gao, Nan

    2018-08-01

    Whether environmental (thermal, chemical, and nutrient) signals generate quantifiable, nanoscale, mechanophysical changes in the cellular plasma membrane has not been well elucidated. Assessment of such mechanophysical properties of plasma membrane may shed lights on fundamental cellular process. Atomic force microscopic (AFM) measurement of the mechanical properties of live cells was hampered by the difficulty in accounting for the effects of the cantilever motion and the associated hydrodynamic force on the mechanical measurement. These challenges have been addressed in our recently developed control-based AFM nanomechanical measurement protocol, which enables a fast, noninvasive, broadband measurement of the real-time changes in plasma membrane elasticity in live cells. Here we show using this newly developed AFM platform that the plasma membrane of live mammalian cells exhibits a constant and quantifiable nanomechanical property, the membrane elasticity. This mechanical property sensitively changes in response to environmental factors, such as the thermal, chemical, and growth factor stimuli. We demonstrate that different chemical inhibitors of endocytosis elicit distinct changes in plasma membrane elastic modulus reflecting their specific molecular actions on the lipid configuration or the endocytic machinery. Interestingly, two different growth factors, EGF and Wnt3a, elicited distinct elastic force profiles revealed by AFM at the plasma membrane during receptor-mediated endocytosis. By applying this platform to genetically modified cells, we uncovered a previously unknown contribution of Cdc42, a key component of the cellular trafficking network, to EGF-stimulated endocytosis at plasma membrane. Together, this nanomechanical AFM study establishes an important foundation that is expandable and adaptable for investigation of cellular membrane evolution in response to various key extracellular signals. © 2017 Wiley Periodicals, Inc.

  13. Decrement of GABAA receptor-mediated inhibitory postsynaptic currents in dentate granule cells in epileptic hippocampus.

    Science.gov (United States)

    Isokawa, M

    1996-05-01

    1. Inhibitory postsynaptic currents (IPSCs) were studied in hippocampal dentate granule cells (DGCs) in the pilocarpine model and human temporal lobe epilepsy, with the use of the whole cell patch-clamp recording technique in slice preparations. 2. In the pilocarpine model, hippocampal slices were prepared from rats that were allowed to experience spontaneous seizures for 2 mo. Human hippocampal specimens were obtained from epileptic patients who underwent surgical treatment for medically intractable seizures. 3. IPSCs were generated by single perforant path stimulation and recorded at a membrane potential (Vm) of 0 mV near the reversal potential of glutamate excitatory postsynaptic currents in the voltage-clamp recording. IPSCs were pharmacologically identified as gamma-aminobutyric acid-A (GABAA) IPSCs by 10 microM bicuculline methiodide. 4. During low-frequency stimulation, IPSCs were not different in amplitude among non-seizure-experienced rat hippocampi, human nonsclerotic hippocampi, seizure-experienced rat hippocampi, and human sclerotic hippocampi. In the last two groups of DGCs, current-clamp recordings indicated the presence of prolonged excitatory postsynaptic potentials (EPSPs) mediated by the N-methyl-D-aspartate (NMDA) receptor. 5. High-frequency stimulation, administered at Vm = -30 mV to activate NMDA currents, reduced GABAA IPSC amplitude specifically in seizure-experienced rat hippocampi (t = 2.5, P < 0.03) and human sclerotic hippocampi (t = 7.7, P < 0.01). This reduction was blocked by an NMDA receptor antagonist, 2-amino-5-phosphonovaleric acid (APV) (50 microM). The time for GABAA IPSCs to recover to their original amplitude was also shortened by the application of APV. 6. I conclude that, when intensively activated, NMDA receptor-mediated excitatory transmission may interact with GABAergic synaptic inhibition in DGCs in seizure-experienced hippocampus to transiently reduce GABA(A) receptor-channel function. Such interactions may contribute to

  14. Receptor-mediated transcytosis of cyclophilin B through the blood-brain barrier.

    Science.gov (United States)

    Carpentier, M; Descamps, L; Allain, F; Denys, A; Durieux, S; Fenart, L; Kieda, C; Cecchelli, R; Spik, G

    1999-07-01

    Cyclophilin B (CyPB) is a cyclosporin A (CsA)-binding protein mainly located in intracellular vesicles and secreted in biological fluids. In previous works, we demonstrated that CyPB interacts with T lymphocytes and enhances in vitro cellular incorporation and activity of CsA. In addition to its immunosuppressive activity, CsA is able to promote regeneration of damaged peripheral nerves. However, the crossing of the drug from plasma to neural tissue is restricted by the relative impermeability of the blood-brain barrier. To know whether CyPB might also participate in the delivery of CsA into the brain, we have analyzed the interactions of CyPB with brain capillary endothelial cells. First, we demonstrated that CyPB binds to two types of binding sites present at the surface of capillary endothelial cells from various species of tissues. The first type of binding sites (K(D) = 300 nM; number of sites = 3 x 10(6)) is related to interactions with negatively charged compounds such as proteoglycans. The second type of binding sites, approximately 50,000 per cell, exhibits a higher affinity for CyPB (K(D) = 15 nM) and is involved in an endocytosis process, indicating it might correspond to a functional receptor. Finally, the use of an in vitro model of blood-brain barrier allowed us to demonstrate that CyPB is transcytosed by a receptor-mediated pathway (flux = 16.5 fmol/cm2/h). In these conditions, CyPB did not significantly modify the passage of CsA, indicating that it is unlikely to provide a pathway for CsA brain delivery.

  15. NMDA receptors mediate neuron-to-glia signaling in mouse cortical astrocytes.

    Science.gov (United States)

    Lalo, Ulyana; Pankratov, Yuri; Kirchhoff, Frank; North, R Alan; Verkhratsky, Alexei

    2006-03-08

    Chemical transmission between neurons and glial cells is an important element of integration in the CNS. Here, we describe currents activated by NMDA in cortical astrocytes, identified in transgenic mice that express enhanced green fluorescent protein under control of the human glial fibrillary acidic protein promoter. Astrocytes were studied by whole-cell voltage clamp either in slices or after gentle nonenzymatic mechanical dissociation. Acutely isolated astrocytes showed a three-component response to glutamate. The initial rapid component was blocked by 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide (NBQX), which is an antagonist of AMPA receptors (IC50, 2 microM), and the NMDA receptor antagonist D-AP-5 blocked the later sustained component (IC50, 0.6 microM). The third component of glutamate application response was sensitive to D,L-threo-beta-benzyloxyaspartate, a glutamate transporter blocker. Fast application of NMDA evoked concentration-dependent inward currents (EC50, 0.3 microM); these showed use-dependent block by (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (MK-801). These NMDA-evoked currents were linearly dependent on membrane potential and were not affected by extracellular magnesium at concentrations up to 10 mM. Electrical stimulation of axons in layer IV-VI induced a complex inward current in astrocytes situated in the cortical layer II, part of which was sensitive to MK-801 at holding potential -80 mV and was not affected by the AMPA glutamate receptor antagonist NBQX. The fast miniature spontaneous currents were observed in cortical astrocytes in slices as well. These currents exhibited both AMPA and NMDA receptor-mediated components. We conclude that cortical astrocytes express functional NMDA receptors that are devoid of Mg2+ block, and these receptors are involved in neuronal-glial signal transmission.

  16. Scavenger receptors mediate the role of SUMO and Ftz-f1 in Drosophila steroidogenesis.

    Directory of Open Access Journals (Sweden)

    Ana Talamillo

    2013-04-01

    Full Text Available SUMOylation participates in ecdysteroid biosynthesis at the onset of metamorphosis in Drosophila melanogaster. Silencing the Drosophila SUMO homologue smt3 in the prothoracic gland leads to reduced lipid content, low ecdysone titers, and a block in the larval-pupal transition. Here we show that the SR-BI family of Scavenger Receptors mediates SUMO functions. Reduced levels of Snmp1 compromise lipid uptake in the prothoracic gland. In addition, overexpression of Snmp1 is able to recover lipid droplet levels in the smt3 knockdown prothoracic gland cells. Snmp1 expression depends on Ftz-f1 (an NR5A-type orphan nuclear receptor, the expression of which, in turn, depends on SUMO. Furthermore, we show by in vitro and in vivo experiments that Ftz-f1 is SUMOylated. RNAi-mediated knockdown of ftz-f1 phenocopies that of smt3 at the larval to pupal transition, thus Ftz-f1 is an interesting candidate to mediate some of the functions of SUMO at the onset of metamorphosis. Additionally, we demonstrate that the role of SUMOylation, Ftz-f1, and the Scavenger Receptors in lipid capture and mobilization is conserved in other steroidogenic tissues such as the follicle cells of the ovary. smt3 knockdown, as well as ftz-f1 or Scavenger knockdown, depleted the lipid content of the follicle cells, which could be rescued by Snmp1 overexpression. Therefore, our data provide new insights into the regulation of metamorphosis via lipid homeostasis, showing that Drosophila Smt3, Ftz-f1, and SR-BIs are part of a general mechanism for uptake of lipids such as cholesterol, required during development in steroidogenic tissues.

  17. P2X receptor-mediated ATP purinergic signaling in health and disease

    Directory of Open Access Journals (Sweden)

    Jiang LH

    2012-09-01

    Full Text Available Lin-Hua JiangSchool of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, United KingdomAbstract: Purinergic P2X receptors are plasma membrane proteins present in a wide range of mammalian cells where they act as a cellular sensor, enabling cells to detect and respond to extracellular adenosine triphosphate (ATP, an important signaling molecule. P2X receptors function as ligand-gated Ca2+-permeable cationic channels that open upon ATP binding to elevate intracellular Ca2+ concentrations and cause membrane depolarization. In response to sustained activation, P2X receptors induce formation of a pore permeable to large molecules. P2X receptors also interact with distinct functional proteins and membrane lipids to form specialized signaling complexes. Studies have provided compelling evidence to show that such P2X receptor-mediated ATP-signaling mechanisms determine and regulate a growing number and diversity of important physiological processes, including neurotransmission, muscle contraction, and cytokine release. There is accumulating evidence to support strong causative relationships of altered receptor expression and function with chronic pain, inflammatory diseases, cancers, and other pathologies or diseases. Numerous high throughput screening drug discovery programs and preclinical studies have thus far demonstrated the proof of concepts that the P2X receptors are druggable targets and selective receptor antagonism is a promising therapeutics approach. This review will discuss the recent progress in understanding the mammalian P2X receptors with respect to the ATP-signaling mechanisms, physiological and pathophysiological roles, and development and preclinical studies of receptor antagonists.Keywords: extracellular ATP, ion channel, large pore, signaling complex, chronic pain, inflammatory diseases

  18. Receptor-mediated endocytosis of low density lipoproteins in aortic endothelial cells

    International Nuclear Information System (INIS)

    Sanan, D.A.

    1986-04-01

    Lipoprotein binding and metabolism in actively-dividing (subconfluent) and quiescent (postconfluent) bovine aortic endothelial cells (ECs) were qualitatively investigated by fluorescence microscopy using dioctadecylindocarbocyanine-labelled lipoproteins and by indirect immunofluorescence microscopy. LDL and acetylated-LDL (AcLDL) were seen bound to the surfaces of subconfluent ECs (at 4 degrees C or at 37 degrees C), as a random distribution of punctate foci. ECs therefore closely resembled fibroblasts in the distribution of LDL receptors on their surfaces. No binding of LDL was seen on postconfluent EC surfaces by either direct or indirect fluorescence microscopy. The patterns of AcLDL binding on postconfluent ECs resembled those on subconfluent ECs. Intracellular LDL and AcLDL occurred as perinuclear accumulations of large fluorescent disc-shaped profiles in subconfluent ECs. These accumulations were shown to arise from surface-bound material by pulse-chase experiments. Intracellular LDL was absent in the majority of postconfluent ECs, while AcLDL accumulation was massive. 'Wounding' of cultures allowed simultaneous assessment of lipoprotein metabolism in quiescent and actively-dividing areas of the same culture. It is concluded that postconfluent quiescent bovine aortic ECs in vitro metabolise virtually no LDL via the LDL-receptor pathway due to a vanishingly low number of LDL receptors. This contrasts with the ability of postconfluent cells to metabolise relatively large amounts of AcLDL via a receptor-mediated mechanism. The significance of these conclusions is discussed with respect to the interaction of plasma lipoproteins with the endothelium in vivo. 301 refs

  19. Proteomic and transcriptomic profiling of in vitro established radiation resistant oral cancer cells for identification of radioresistance related biomarkers

    International Nuclear Information System (INIS)

    Mohd Yasser; Pawar, Sagar; Teni, Tanuja

    2016-01-01

    Radiotherapy is an integral part of oral cancer treatment, either alone or in combination with surgery. But, during radiotherapy, oral tumours of a subset of patients develop radioresistance that creates major obstruction towards its efficacy. The aim of our study was to establish radioresistant cell lines from different oral subsites using clinically admissible low dose radiation and profile them by proteomic and transcriptomic approaches to identify proteins associated with radioresistance in oral cancer

  20. Response of clonogenic cells of mice solid tumour NKLy/LL to N-methyl-N-nitrosourea

    International Nuclear Information System (INIS)

    Chan Tik Kan; Afanas'ev, G.G.; Pelevina, I.I.

    1979-01-01

    By cloning in vitro the cells of NKLy/LL solid tumour of mice it has been shown that the curve of clonogenic cell survival versus N-methyl-N-nitrosourea (MNU) dose in the 12.5-200.0 mg/kg interval is exponential and characterized by Dsub(o)=29.15 mg/kg dose value. Large size tumours (15.0-17.0 g) are characterized by higher death rate of clonogenic cells (survivor fraction approximately 0.5%) than in 1.0-7.0 g tumours (survivor fraction 2-4%). That is, evidently, related to higher sensitivity of the resting tumour cells to MNU

  1. Assessment of the Radioprotection Efficacy of Antioxidant Substances in Foods in regard to Clonogenic Cell Survival

    Energy Technology Data Exchange (ETDEWEB)

    Park, Hyosung; Lee, Minho; Kim, Eunhee [Seoul National Univ., Seoul (Korea, Republic of)

    2014-05-15

    Radiation is perceived as a hazard in spite of its diverse uses. The public are more sensitive than ever to the activities from the nuclear power plant industry since Fukushima accident and the consequential environment contamination. The never-dissolved fear from the public of the nuclear energy costs the nuclear industry too much for safety measures not to mention brings the public uneasy life. Beta carotene, oltipraz and luteolin, which are easily taken from various foods, are substances that may protect cells from damage caused by unstable molecules known as free radicals. The purpose of this study is to inform people of accessible radiation protection measures in everyday lives. Three antioxidant substances were investigated regarding their radioprotection effects counted in terms of clonogenic cell surviving fraction in vitro. The radioprotection efficacy was observed with those substances at concentrations below certain levels. At concentrations beyond those levels, the efficacy was canceled out by their inherent cytotoxicity.

  2. CD133 (Prominin negative human neural stem cells are clonogenic and tripotent.

    Directory of Open Access Journals (Sweden)

    Yirui Sun

    Full Text Available CD133 (Prominin is widely used as a marker for the identification and isolation of neural precursor cells from normal brain or tumor tissue. However, the assumption that CD133 is expressed constitutively in neural precursor cells has not been examined.In this study, we demonstrate that CD133 and a second marker CD15 are expressed heterogeneously in uniformly undifferentiated human neural stem (NS cell cultures. After fractionation by flow cytometry, clonogenic tripotent cells are found in populations negative or positive for either marker. We further show that CD133 is down-regulated at the mRNA level in cells lacking CD133 immunoreactivity. Cell cycle profiling reveals that CD133 negative cells largely reside in G1/G0, while CD133 positive cells are predominantly in S, G2, or M phase. A similar pattern is apparent in mouse NS cell lines. Compared to mouse NS cells, however, human NS cell cultures harbour an increased proportion of CD133 negative cells and display a longer doubling time. This may in part reflect a sub-population of slow- or non-cycling cells amongst human NS cells because we find that around 5% of cells do not take up BrdU over a 14-day labelling period. Non-proliferating NS cells remain undifferentiated and at least some of them are capable of re-entry into the cell cycle and subsequent continuous expansion.The finding that a significant fraction of clonogenic neural stem cells lack the established markers CD133 and CD15, and that some of these cells may be dormant or slow-cycling, has implications for approaches to identify and isolate neural stem cells and brain cancer stem cells. Our data also suggest the possibility that CD133 may be specifically down-regulated during G0/G1, and this should be considered when this marker is used to identify and isolate other tissue and cancer stem cells.

  3. Applications of high-throughput clonogenic survival assays in high-LET particle microbeams

    Directory of Open Access Journals (Sweden)

    Antonios eGeorgantzoglou

    2016-01-01

    Full Text Available Charged particle therapy is increasingly becoming a valuable tool in cancer treatment, mainly due to the favorable interaction of particle radiation with matter. Its application is still limited due, in part, to lack of data regarding the radiosensitivity of certain cell lines to this radiation type, especially to high-LET particles. From the earliest days of radiation biology, the clonogenic survival assay has been used to provide radiation response data. This method produces reliable data but it is not optimized for high-throughput microbeam studies with high-LET radiation where high levels of cell killing lead to a very low probability of maintaining cells’ clonogenic potential. A new method, therefore, is proposed in this paper, which could potentially allow these experiments to be conducted in a high-throughput fashion. Cells are seeded in special polypropylene dishes and bright-field illumination provides cell visualization. Digital images are obtained and cell detection is applied based on corner detection, generating individual cell targets as x-y points. These points in the dish are then irradiated individually by a micron field size high-LET microbeam. Post-irradiation, time-lapse imaging follows cells’ response. All irradiated cells are tracked by linking trajectories in all time-frames, based on finding their nearest position. Cell divisions are detected based on cell appearance and individual cell temporary corner density. The number of divisions anticipated is low due to the high probability of cell killing from high-LET irradiation. Survival curves are produced based on cell’s capacity to divide at least 4-5 times. The process is repeated for a range of doses of radiation. Validation shows the efficiency of the proposed cell detection and tracking method in finding cell divisions.

  4. Applications of High-Throughput Clonogenic Survival Assays in High-LET Particle Microbeams.

    Science.gov (United States)

    Georgantzoglou, Antonios; Merchant, Michael J; Jeynes, Jonathan C G; Mayhead, Natalie; Punia, Natasha; Butler, Rachel E; Jena, Rajesh

    2015-01-01

    Charged particle therapy is increasingly becoming a valuable tool in cancer treatment, mainly due to the favorable interaction of particle radiation with matter. Its application is still limited due, in part, to lack of data regarding the radiosensitivity of certain cell lines to this radiation type, especially to high-linear energy transfer (LET) particles. From the earliest days of radiation biology, the clonogenic survival assay has been used to provide radiation response data. This method produces reliable data but it is not optimized for high-throughput microbeam studies with high-LET radiation where high levels of cell killing lead to a very low probability of maintaining cells' clonogenic potential. A new method, therefore, is proposed in this paper, which could potentially allow these experiments to be conducted in a high-throughput fashion. Cells are seeded in special polypropylene dishes and bright-field illumination provides cell visualization. Digital images are obtained and cell detection is applied based on corner detection, generating individual cell targets as x-y points. These points in the dish are then irradiated individually by a micron field size high-LET microbeam. Post-irradiation, time-lapse imaging follows cells' response. All irradiated cells are tracked by linking trajectories in all time-frames, based on finding their nearest position. Cell divisions are detected based on cell appearance and individual cell temporary corner density. The number of divisions anticipated is low due to the high probability of cell killing from high-LET irradiation. Survival curves are produced based on cell's capacity to divide at least four to five times. The process is repeated for a range of doses of radiation. Validation shows the efficiency of the proposed cell detection and tracking method in finding cell divisions.

  5. Isolation of radioresistant microorganisms from a Co/sup 60/ irradiation plant

    International Nuclear Information System (INIS)

    Lezcano, Graciela

    1982-01-01

    The continuos exposition to low doses of gamma irradiation can produce changes in the microflora's radioresistance. In order to obtain information about these possible modifications, the water from the pool used as shielding of the source, as well as the air and the dust in the irradiation chamber of the semi-industrial irradiation plant existing at the Ezeiza Atomic Center were analyzed. The number of microorganisms was determined by filtration techniques and by dilutions. Radioresistance studies of the contaminating microflora were performed. The value of the D/sub 10/ dose was determined in the conditions of highest resistance. A pronounced decrease in the number of microorganisms was observed as a radiation effect in the samples of water and dust, but not in the air samples, this as a consequence of the extractors' action that continually renews the air and the flora in the chamber, thus preventing high-dose exposure. In the air samples no increase of the microorganisms' radioresistance was observed. In the pool water flora, the development of a great radioresistance was observed. A microorganism whose inactivation curve shows a shoulder of 3.2 Mrad was isolated. This high radioresistance could be the result of the continous exposure to low doses during six years. Contrarily, the microorganims of the irradiation chamber's dust did not increase their radioresistance wiht regard to the common contaminants. In the flora of the dust used as a target, two microorganims whose D/sub 10/ were in excess of 400 krad were found; these could be ocassional contaminants. The radioresistant microorganims were isolated and characterized according to Cowan's scheme, the water microorganisms being identified as belonging to the genus Corynebacterium and the earth ones to the genera Micrococcus and Corynebacterium. (author) [es

  6. Detection of Quiescent Radioresistant Epithelial Progenitors in the Adult Thymus

    Directory of Open Access Journals (Sweden)

    Maude Dumont-Lagacé

    2017-12-01

    Full Text Available Thymic aging precedes that of other organs and is initiated by the gradual loss of thymic epithelial cells (TECs. Based on in vitro culture and transplantation assays, recent studies have reported on the presence of thymic epithelial progenitor cells (TEPCs in young adult mice. However, the physiological role and properties of TEPC populations reported to date remain unclear. Using an in vivo label-retention assay, we previously identified a population of quiescent but non-senescent TECs. The goals of this study were therefore (i to evaluate the contribution of these quiescent TECs to thymic regeneration following irradiation-induced acute thymic injury and (ii to characterize their phenotypic and molecular profiles using flow cytometry, immunohistology, and transcriptome sequencing. We report that while UEA1+ cells cycle the most in steady state, they are greatly affected by irradiation, leading to cell loss and proliferative arrest following acute thymic involution. On the opposite, the UEA1– subset of quiescent TECs is radioresistant and proliferate in situ following acute thymic involution, thereby contributing to thymic regeneration in 28- to 30-week-old mice. UEA1– quiescent TECs display an undifferentiated phenotype (co-expression of K8 and K5 cytokeratins and express high levels of genes that regulate stem cell activity in different tissues (e.g., Podxl and Ptprz1. In addition, two features suggest that UEA1– quiescent TECs occupy discrete stromal niches: (i their preferential location in clusters adjacent to the cortico-medullary junction and (ii their high expression of genes involved in cross talk with mesenchymal cells. The ability of UEA1– quiescent TECs to participate to TEC regeneration qualifies them as in vivo progenitor cells particularly relevant in the context of regeneration following acute thymic injury.

  7. Fractionated irradiation-induced EMT-like phenotype conferred radioresistance in esophageal squamous cell carcinoma

    Science.gov (United States)

    Zhang, Hongfang; Luo, Honglei; Jiang, Zhenzhen; Yue, Jing; Hou, Qiang; Xie, Ruifei; Wu, Shixiu

    2016-01-01

    The efficacy of radiotherapy, one major treatment modality for esophageal squamous cell carcinoma (ESCC) is severely attenuated by radioresistance. Epithelial-to-mesenchymal transition (EMT) is a cellular process that determines therapy response and tumor progression. However, whether EMT is induced by ionizing radiation and involved in tumor radioresistance has been less studied in ESCC. Using multiple fractionated irradiation, the radioresistant esophageal squamous cancer cell line KYSE-150R had been established from its parental cell line KYSE-150. We found KYSE-150R displayed a significant EMT phenotype with an elongated spindle shape and down-regulated epithelial marker E-cadherin and up-regulated mesenchymal marker N-cadherin in comparison with KYSE-150. Furthermore, KYSE-150R also possessed some stemness-like properties characterized by density-dependent growth promotion and strong capability for sphere formation and tumorigenesis in NOD-SCID mice. Mechanical studies have revealed that WISP1, a secreted matricellular protein, is highly expressed in KYSE-150R and mediates EMT-associated radioresistance both in ESCC cells and in xenograft tumor models. Moreover, WISP1 has been demonstrated to be closely associated with the EMT phenotype observed in ESCC patients and to be an independent prognosis factor of ESCC patients treated with radiotherapy. Our study highlighted WISP1 as an attractive target to reverse EMT-associated radioresistance in ESCC and can be used as an independent prognostic factor of patients treated with radiotherapy. PMID:27125498

  8. Fractionated irradiation-induced EMT-like phenotype conferred radioresistance in esophageal squamous cell carcinoma

    International Nuclear Information System (INIS)

    Zhang, Hongfang; Luo, Honglei; Jiang, Zhenzhen; Yue, Jing; Hou, Qiang; Xie, Ruifei; Wu, Shixiu

    2016-01-01

    The efficacy of radiotherapy, one major treatment modality for esophageal squamous cell carcinoma (ESCC) is severely attenuated by radioresistance. Epithelial-to-mesenchymal transition (EMT) is a cellular process that determines therapy response and tumor progression. However, whether EMT is induced by ionizing radiation and involved in tumor radioresistance has been less studied in ESCC. Using multiple fractionated irradiation, the radioresistant esophageal squamous cancer cell line KYSE-150R had been established from its parental cell line KYSE-150. We found KYSE-150R displayed a significant EMT phenotype with an elongated spindle shape and down-regulated epithelial marker E-cadherin and up-regulated mesenchymal marker N-cadherin in comparison with KYSE-150. Furthermore, KYSE-150R also possessed some stemness-like properties characterized by density-dependent growth promotion and strong capability for sphere formation and tumorigenesis in NOD-SCID mice. Mechanical studies have revealed that WISP1, a secreted matricellular protein, is highly expressed in KYSE-150R and mediates EMT-associated radioresistance both in ESCC cells and in xenograft tumor models. Moreover, WISP1 has been demonstrated to be closely associated with the EMT phenotype observed in ESCC patients and to be an independent prognosis factor of ESCC patients treated with radiotherapy. Our study highlighted WISP1 as an attractive target to reverse EMT-associated radioresistance in ESCC and can be used as an independent prognostic factor of patients treated with radiotherapy

  9. Histamine H2 Receptor-Mediated Suppression of Intestinal Inflammation by Probiotic Lactobacillus reuteri.

    Science.gov (United States)

    Gao, Chunxu; Major, Angela; Rendon, David; Lugo, Monica; Jackson, Vanessa; Shi, Zhongcheng; Mori-Akiyama, Yuko; Versalovic, James

    2015-12-15

    -mediated suppression of colonic inflammation. The effective combination of diet, gut bacteria, and host receptor-mediated signaling may result in opportunities for therapeutic microbiology and provide clues for discovery and development of next-generation probiotics. Copyright © 2015 Gao et al.

  10. Natural killer cell lines preferentially kill clonogenic multiple myeloma cells and decrease myeloma engraftment in a bioluminescent xenograft mouse model.

    Science.gov (United States)

    Swift, Brenna E; Williams, Brent A; Kosaka, Yoko; Wang, Xing-Hua; Medin, Jeffrey A; Viswanathan, Sowmya; Martinez-Lopez, Joaquin; Keating, Armand

    2012-07-01

    Novel therapies capable of targeting drug resistant clonogenic MM cells are required for more effective treatment of multiple myeloma. This study investigates the cytotoxicity of natural killer cell lines against bulk and clonogenic multiple myeloma and evaluates the tumor burden after NK cell therapy in a bioluminescent xenograft mouse model. The cytotoxicity of natural killer cell lines was evaluated against bulk multiple myeloma cell lines using chromium release and flow cytometry cytotoxicity assays. Selected activating receptors on natural killer cells were blocked to determine their role in multiple myeloma recognition. Growth inhibition of clonogenic multiple myeloma cells was assessed in a methylcellulose clonogenic assay in combination with secondary replating to evaluate the self-renewal of residual progenitors after natural killer cell treatment. A bioluminescent mouse model was developed using the human U266 cell line transduced to express green fluorescent protein and luciferase (U266eGFPluc) to monitor disease progression in vivo and assess bone marrow engraftment after intravenous NK-92 cell therapy. Three multiple myeloma cell lines were sensitive to NK-92 and KHYG-1 cytotoxicity mediated by NKp30, NKp46, NKG2D and DNAM-1 activating receptors. NK-92 and KHYG-1 demonstrated 2- to 3-fold greater inhibition of clonogenic multiple myeloma growth, compared with killing of the bulk tumor population. In addition, the residual colonies after treatment formed significantly fewer colonies compared to the control in a secondary replating for a cumulative clonogenic inhibition of 89-99% at the 20:1 effector to target ratio. Multiple myeloma tumor burden was reduced by NK-92 in a xenograft mouse model as measured by bioluminescence imaging and reduction in bone marrow engraftment of U266eGFPluc cells by flow cytometry. This study demonstrates that NK-92 and KHYG-1 are capable of killing clonogenic and bulk multiple myeloma cells. In addition, multiple myeloma

  11. Astrocytes protect neurons against methylmercury via ATP/P2Y(1) receptor-mediated pathways in astrocytes.

    Science.gov (United States)

    Noguchi, Yusuke; Shinozaki, Youichi; Fujishita, Kayoko; Shibata, Keisuke; Imura, Yoshio; Morizawa, Yosuke; Gachet, Christian; Koizumi, Schuichi

    2013-01-01

    Methylmercury (MeHg) is a well known environmental pollutant that induces serious neuronal damage. Although MeHg readily crosses the blood-brain barrier, and should affect both neurons and glial cells, how it affects glia or neuron-to-glia interactions has received only limited attention. Here, we report that MeHg triggers ATP/P2Y1 receptor signals in astrocytes, thereby protecting neurons against MeHg via interleukin-6 (IL-6)-mediated pathways. MeHg increased several mRNAs in astrocytes, among which IL-6 was the highest. For this, ATP/P2Y1 receptor-mediated mechanisms were required because the IL-6 production was (i) inhibited by a P2Y1 receptor antagonist, MRS2179, (ii) abolished in astrocytes obtained from P2Y1 receptor-knockout mice, and (iii) mimicked by exogenously applied ATP. In addition, (iv) MeHg released ATP by exocytosis from astrocytes. As for the intracellular mechanisms responsible for IL-6 production, p38 MAP kinase was involved. MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6. As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved. Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

  12. Stable radioresistance in ataxia-telangiectasia cells containing DNA from normal human cells

    International Nuclear Information System (INIS)

    Kapp, L.N.; Painter, R.B.

    1989-01-01

    SV40-transformed ataxia-telangiectasia (AT) cells were transfected with a cosmid containing a normal human DNA library and selectable marker, the neo gene, which endows successfully transformed mammalian cells with resistance to the antibiotic G418. Cells from this line were irradiated with 50 Gy of X-rays and fused with non-transfected AT cells. Among the G418-resistant colonies recovered was one stably resistant to radiation. Resistance to ionizing radiation of both primary transfectant line and its fusion derivative was intermediate between that of AT cells and normal cells, as assayed by colony-forming ability and measurement of radiation-induced G 2 chromatic aberrations; both cell lines retained AT-like radioresistant DNA synthesis. Results suggest that, because radioresistance in transfected cells was not as great as in normal human cells, two hallmarks of AT, radiosensitivity and radioresistant DNA synthesis, may still be the result of a single defective AT gene. (author)

  13. Expressed sequence tags of differential genes in the radioresistant mice and their parental mice

    International Nuclear Information System (INIS)

    Wang Qin; Yue Jingyin; Li Jin; Song Li; Liu Qiang; Mu Chuanjie; Wu Hongying

    2009-01-01

    Objective: To explore radioresistance correlative genes in IRM-2 inbred mouse. Methods: The total RNA was extracted from spleen cells of IRM-2 and their parent 615 and ICR/JCL mouse. The mRNA differential display technique was used to analyze gene expression differences. Each differential bands were amplified by PCR, cloned and sequenced. Results: There were 75 differential expression bands appearing in IRM-2 mouse but not in 615 and ICR/JCL mouse. Fifty-two pieces of cDNA sequences were got by sequencing. Twenty-one expressed sequence tags (EST) that were not the same as known mice genes were found and registered by comparing with GenBank database. Conclusion: Twenty-one EST denote that radioresistance correlative genes may be in IRM-2 mouse, which have laid a foundation for isolating and identifying radioresistance correlative genes in further study. (authors)

  14. No impact on P-gp level in radio-resistant Mcf-7 cells

    International Nuclear Information System (INIS)

    Madhu, L.N.; Rao, Shama; Sarojini, B.K.

    2016-01-01

    Cancer has become the leading cause of human death worldwide. One possible cause for therapeutic failure is that residual tumor cells are reminiscent of stem cells, which ultimately give rise to secondary tumors or distant metastasis. The property of resistance to radiation therapy or chemotherapy might be the major clinical criterion to characterize 'cancer stem cells (CSCs)'. In the process of radiotherapy, the radiosensitive cancer will become a radioresistant one. Such radio-resistance cells might also show the characters of multi drug resistance (MRD) properties which may affect the chemotherapy process. The present study was carried out to know the expression level of P-gp, a MRD protein in radioresistance breast cancer cells. The study conducted by exposing the MCF-7 cells to 4Gy of gamma radiation

  15. Impact of anthropogenic forest contamination on radioresistance of woody plant seed

    International Nuclear Information System (INIS)

    Yushkov, P.I.

    2004-01-01

    Radioresistance of seeds of bay willow (Salix pentandra L.) and great sallow willow (Salix caprea L.) from forests chronically affected and non-affected by acidic (SO 2 , NO x , HF, etc.) industries has been studied and compared. Bay willow seeds of 6 harvests showed no difference in radioresistance. However, seeds of both species manifested strong synchronous variability in resistance to preplant exposure. Also, no influence was observed of mother plant gas content on great sallow willow seeds of different harvests. Data obtained confirm similar results of previous studies conducted by the author (1987) to identify the impact of plant gas content on radioresistance of seeds and seedlings of pine (Pinus sylvestris L.) and birch (Betula verrucosa Erh.). (author)

  16. TRAIL causes deletions at the HPRT and TK1 loci of clonogenically competent cells

    Energy Technology Data Exchange (ETDEWEB)

    Miles, Mark A.; Shekhar, Tanmay M. [Department of Biochemistry and Genetics, La Trobe University, Bundoora, Victoria (Australia); La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Victoria (Australia); Hall, Nathan E. [La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Victoria (Australia); Life Sciences Computation Centre, Victorian Life Sciences Computation Initiative, Melbourne, Victoria (Australia); Hawkins, Christine J., E-mail: c.hawkins@latrobe.edu.au [Department of Biochemistry and Genetics, La Trobe University, Bundoora, Victoria (Australia); La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Victoria (Australia)

    2016-05-15

    Highlights: • Treatment with TRAIL or EMS provokes mutations in clonogenically viable TK6 cells. • TRAIL is 2–5-fold less mutagenic than an equivalently lethal concentration of EMS. • EMS mainly causes transition mutations at the HPRT and TK1 loci of TK6 cells. • Most loss-of-function HPRT or TK1 mutations caused by TRAIL treatment are deletions. - Abstract: When chemotherapy and radiotherapy are effective, they function by inducing DNA damage in cancerous cells, which respond by undergoing apoptosis. Some adverse effects can result from collateral destruction of non-cancerous cells, via the same mechanism. Therapy-related cancers, a particularly serious adverse effect of anti-cancer treatments, develop due to oncogenic mutations created in non-cancerous cells by the DNA damaging therapies used to eliminate the original cancer. Physiologically achievable concentrations of direct apoptosis inducing anti-cancer drugs that target Bcl-2 and IAP proteins possess negligible mutagenic activity, however death receptor agonists like TRAIL/Apo2L can provoke mutations in surviving cells, probably via caspase-mediated activation of the nuclease CAD. In this study we compared the types of mutations sustained in the HPRT and TK1 loci of clonogenically competent cells following treatment with TRAIL or the alkylating agent ethyl methanesulfonate (EMS). As expected, the loss-of-function mutations in the HPRT or TK1 loci triggered by exposure to EMS were almost all transitions. In contrast, only a minority of the mutations identified in TRAIL-treated clones lacking HPRT or TK1 activity were substitutions. Almost three quarters of the TRAIL-induced mutations were partial or complete deletions of the HPRT or TK1 genes, consistent with sub-lethal TRAIL treatment provoking double strand breaks, which may be mis-repaired by non-homologous end joining (NHEJ). Mis-repair of double-strand breaks following exposure to chemotherapy drugs has been implicated in the pathogenesis of

  17. TRAIL causes deletions at the HPRT and TK1 loci of clonogenically competent cells

    International Nuclear Information System (INIS)

    Miles, Mark A.; Shekhar, Tanmay M.; Hall, Nathan E.; Hawkins, Christine J.

    2016-01-01

    Highlights: • Treatment with TRAIL or EMS provokes mutations in clonogenically viable TK6 cells. • TRAIL is 2–5-fold less mutagenic than an equivalently lethal concentration of EMS. • EMS mainly causes transition mutations at the HPRT and TK1 loci of TK6 cells. • Most loss-of-function HPRT or TK1 mutations caused by TRAIL treatment are deletions. - Abstract: When chemotherapy and radiotherapy are effective, they function by inducing DNA damage in cancerous cells, which respond by undergoing apoptosis. Some adverse effects can result from collateral destruction of non-cancerous cells, via the same mechanism. Therapy-related cancers, a particularly serious adverse effect of anti-cancer treatments, develop due to oncogenic mutations created in non-cancerous cells by the DNA damaging therapies used to eliminate the original cancer. Physiologically achievable concentrations of direct apoptosis inducing anti-cancer drugs that target Bcl-2 and IAP proteins possess negligible mutagenic activity, however death receptor agonists like TRAIL/Apo2L can provoke mutations in surviving cells, probably via caspase-mediated activation of the nuclease CAD. In this study we compared the types of mutations sustained in the HPRT and TK1 loci of clonogenically competent cells following treatment with TRAIL or the alkylating agent ethyl methanesulfonate (EMS). As expected, the loss-of-function mutations in the HPRT or TK1 loci triggered by exposure to EMS were almost all transitions. In contrast, only a minority of the mutations identified in TRAIL-treated clones lacking HPRT or TK1 activity were substitutions. Almost three quarters of the TRAIL-induced mutations were partial or complete deletions of the HPRT or TK1 genes, consistent with sub-lethal TRAIL treatment provoking double strand breaks, which may be mis-repaired by non-homologous end joining (NHEJ). Mis-repair of double-strand breaks following exposure to chemotherapy drugs has been implicated in the pathogenesis of

  18. Radioresistance of human glioma spheroids and expression of HSP70, p53 and EGFr

    International Nuclear Information System (INIS)

    Fedrigo, Carlos A; Rocha, Adriana B da; Grivicich, Ivana; Schunemann, Daniel P; Chemale, Ivan M; Santos, Daiane dos; Jacovas, Thais; Boschetti, Patryck S; Jotz, Geraldo P; Filho, Aroldo Braga

    2011-01-01

    Radiation therapy is routinely prescribed for high-grade malignant gliomas. However, the efficacy of this therapeutic modality is often limited by the occurrence of radioresistance, reflected as a diminished susceptibility of the irradiated cells to undergo cell death. Thus, cells have evolved an elegant system in response to ionizing radiation induced DNA damage, where p53, Hsp70 and/or EGFr may play an important role in the process. In the present study, we investigated whether the content of p53, Hsp70 and EGFr are associated to glioblastoma (GBM) cell radioresistance. Spheroids from U-87MG and MO59J cell lines as well as spheroids derived from primary culture of tumor tissue of one GBM patient (UGBM1) were irradiated (5, 10 and 20 Gy), their relative radioresistance were established and the p53, Hsp70 and EGFr contents were immunohistochemically determined. Moreover, we investigated whether EGFr-phospho-Akt and EGFr-MEK-ERK pathways can induce GBM radioresistance using inhibitors of activation of ERK (PD098059) and Akt (wortmannin). At 5 Gy irradiation UGBM1 and U-87MG spheroids showed growth inhibition whereas the MO59J spheroid was relatively radioresistant. Overall, no significant changes in p53 and Hsp70 expression were found following 5 Gy irradiation treatment in all spheroids studied. The only difference observed in Hsp70 content was the periphery distribution in MO59J spheroids. However, 5 Gy treatment induced a significant increase on the EGFr levels in MO59J spheroids. Furthermore, treatment with inhibitors of activation of ERK (PD098059) and Akt (wortmannin) leads to radiosensitization of MO59J spheroids. These results indicate that the PI3K-Akt and MEK-ERK pathways triggered by EGFr confer GBM radioresistance

  19. Up-regulation of integrin β3 in radioresistant pancreatic cancer impairs adenovirus-mediated gene therapy

    International Nuclear Information System (INIS)

    Egami, Takuya; Ohuchida, Kenoki; Yasui, Takaharu; Onimaru, Manabu; Toma, Hiroki; Sato, Norihiro; Tanaka, Masao; Mizumoto, Kazuhiro; Matsumoto, Kunio

    2009-01-01

    Adenovirus-mediated gene therapy is a promising approach for the treatment of pancreatic cancer. We previously reported that radiation enhanced adenovirus-mediated gene expression in pancreatic cancer, suggesting that adenoviral gene therapy might be more effective in radioresistant pancreatic cancer cells. In the present study, we compared the transduction efficiency of adenovirus-delivered genes in radiosensitive and radioresistant cells, and investigated the underlying mechanisms. We used an adenovirus expressing the hepatocyte growth factor antagonist, NK4 (Ad-NK4), as a representative gene therapy. We established two radioresistant human pancreatic cancer cell lines using fractionated irradiation. Radiosensitive and radioresistant pancreatic cancer cells were infected with Ad-NK4, and NK4 levels in the cells were measured. In order to investigate the mechanisms responsible for the differences in the transduction efficiency between these cells, we measured expression of the genes mediating adenovirus infection and endocytosis. The results revealed that NK4 levels in radioresistant cells were significantly lower (P<0.01) than those in radiosensitive cells, although there were no significant differences in adenovirus uptake between radiosensitive cells and radioresistant cells. Integrin β3 was up-regulated and the Coxsackie virus and adenovirus receptor was down-regulated in radioresistant cells, and inhibition of integrin β3 promoted adenovirus gene transfer. These results suggest that inhibition of integrin β3 in radioresistant pancreatic cancer cells could enhance adenovirus-mediated gene therapy. (author)

  20. The chalcone butein from Rhus verniciflua Stokes inhibits clonogenic growth of human breast cancer cells co-cultured with fibroblasts

    Directory of Open Access Journals (Sweden)

    Tan Jenny

    2005-03-01

    Full Text Available Abstract Background Butein (3,4,2',4'-tetrahydroxychalone, a plant polyphenol, is a major biologically active component of the stems of Rhus verniciflua Stokes. It has long been used as a food additive in Korea and as an herbal medicine throughout Asia. Recently, butein has been shown to suppress the functions of fibroblasts. Because fibroblasts are believed to play an important role in promoting the growth of breast cancer cells, we investigated the ability of butein to inhibit the clonogenic growth of small numbers of breast cancer cells co-cultured with fibroblasts in vitro. Methods We first measured the clonogenic growth of small numbers of the UACC-812 human breast cancer cell line co-cultured on monolayers of serum-activated, human fibroblasts in the presence of butein (2 μg/mL or various other modulators of fibroblast function (troglitazone-1 μg/mL; GW9662-1 μM; meloxican-1 μM; and 3,4 dehydroproline-10 μg/mL. In a subsequent experiment, we measured the dose-response effect on the clonogenic growth of UACC-812 breast cancer cells by pre-incubating the fibroblasts with varying concentrations of butein (10 μg/ml-1.25 μg/mL. Finally, we measured the clonogenic growth of primary breast cancer cells obtained from 5 clinical specimens with normal fibroblasts and with fibroblasts that had been pre-treated with a fixed dose of butein (2.5 μg/mL. Results Of the five modulators of fibroblast function that we tested, butein was by far the most potent inhibitor of clonogenic growth of UACC-812 breast cancer cells co-cultured with fibroblasts. Pre-treatment of fibroblasts with concentrations of butein as low as 2.5 μg/mL nearly abolished subsequent clonogenic growth of UACC-812 breast cancer cells co-cultured with the fibroblasts. A similar dose of butein had no effect on the clonogenic growth of breast cancer cells cultured in the absence of fibroblasts. Significantly, clonogenic growth of the primary breast cancer cells was also

  1. Assessment of Radiation-Equivalency of Favorite Foods Based on Apoptotic and Clonogenic Cell Death

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Seunghee; Park, Hyosung; Lee, Minho; Kim, Eunhee [Seoul National Univ., Seoul (Korea, Republic of)

    2013-05-15

    In this study, the cytotoxicity of favorite foods was investigated to provide chemical doses causing bio-effects comparable with those by radiation exposure. Extracts from alcohol, cigarette and coffee have been investigated for their cytotoxicity in terms of apoptosis induction and clonogenic cell killing. The comparison of familiar favorite food items with radiation in cellular toxicity might help the public have balanced judgment on the severity of radiation exposure. Future study pursues estimating dose-rate effect and the investigation will be continued with other cell lines. People fear that ionizing radiation, despite of its usefulness, bears significant risk factor. In particular, after Fukushima nuclear accident in March of 2011, people become over-sensitive to radiation exposure and nuclear power. Such fear by the public of nuclear power inevitably costs the nuclear industry a high expense for safety measures, not to mention brings the public uneasy life. It is important that the consequence of radiation exposure is conveyed to the public in a friendly way with actual data.

  2. Heterogeneity in cytokinetic, clonogenic, and radiation response induced by nutrient deprivation

    International Nuclear Information System (INIS)

    Grdina, D.J.; Williamson, K.D.; Johnson, T.S.; Sigdestad, C.P.

    1986-01-01

    A murine fibrosarcoma cell line (FSA 1233), with a capability to grow in vitro, was used to study biophysical heterogeneity induced by nutrient deprivation. Cell sub-populations were isolated by centrifuging cells through linear Renografin-density gradients. The effects of cell growth to late plateau phase on the cytokinetic, clonogenic, and radiation-survival characteristics were studied. It was observed that with increasing age of the cell culture, an increase in the proportion of relatively dense cells was associated with a decrease in plating efficiency. Cells collected at p=1.08 g/cm 3 have a plating efficiency of 0.2% compared with 0.01% for those isolated at p=1.16 g/cm 3 . Discrepancies were also seen between the tritiated thymidine labeling index (LI) and flow cytometry (FCM, % S phase) for cells collected throughout the gradient. The radiation sensitivities of selected cell populations isolated by density gradient centrifugation were determined and compared with data from a control plateau phase population. 15 refs., 4 figs., 1 tab

  3. Evolution of the clonogenic potential of human epidermal stem/progenitor cells with age

    Directory of Open Access Journals (Sweden)

    Zobiri O

    2012-02-01

    Full Text Available Olivia Zobiri, Nathalie Deshayes, Michelle Rathman-JosserandDepartment of Biological Research, L'Oréal Advanced Research, Clichy Cedex, FranceAbstract: A number of clinical observations have indicated that the regenerative potential and overall function of the epidermis is modified with age. The epidermis becomes thinner, repairs itself less efficiently after wounding, and presents modified barrier function recovery. In addition, the dermal papillae flatten out with increasing age, suggesting a modification in the interaction between epidermal and dermal compartments. As the epidermal regenerative capacity is dependent upon stem and progenitor cell function, it is naturally of interest to identify and understand age-related changes in these particular keratinocyte populations. Previous studies have indicated that the number of stem cells does not decrease with age in mouse models but little solid evidence is currently available concerning human skin. The objective of this study was to evaluate the clonogenic potential of keratinocyte populations isolated from the epidermis of over 50 human donors ranging from 18 to 71 years old. The data indicate that the number of epidermal cells presenting high regenerative potential does not dramatically decline with age in human skin. The authors believe that changes in the microenvironment controlling epidermal basal cell activity are more likely to explain the differences in epidermal function observed with increasing age.Keywords: skin, epidermal stem cells, aging, colony-forming efficiency test

  4. Upregulation of mitochondrial NAD+ levels impairs the clonogenicity of SSEA1+ glioblastoma tumor-initiating cells.

    Science.gov (United States)

    Son, Myung Jin; Ryu, Jae-Sung; Kim, Jae Yun; Kwon, Youjeong; Chung, Kyung-Sook; Mun, Seon Ju; Cho, Yee Sook

    2017-06-09

    Emerging evidence has emphasized the importance of cancer therapies targeting an abnormal metabolic state of tumor-initiating cells (TICs) in which they retain stem cell-like phenotypes and nicotinamide adenine dinucleotide (NAD + ) metabolism. However, the functional role of NAD + metabolism in regulating the characteristics of TICs is not known. In this study, we provide evidence that the mitochondrial NAD + levels affect the characteristics of glioma-driven SSEA1 + TICs, including clonogenic growth potential. An increase in the mitochondrial NAD + levels by the overexpression of the mitochondrial enzyme nicotinamide nucleotide transhydrogenase (NNT) significantly suppressed the sphere-forming ability and induced differentiation of TICs, suggesting a loss of the characteristics of TICs. In addition, increased SIRT3 activity and reduced lactate production, which are mainly observed in healthy and young cells, appeared following NNT-overexpressed TICs. Moreover, in vivo tumorigenic potential was substantially abolished by NNT overexpression. Conversely, the short interfering RNA-mediated knockdown of NNT facilitated the maintenance of TIC characteristics, as evidenced by the increased numbers of large tumor spheres and in vivo tumorigenic potential. Our results demonstrated that targeting the maintenance of healthy mitochondria with increased mitochondrial NAD + levels and SIRT3 activity could be a promising strategy for abolishing the development of TICs as a new therapeutic approach to treating aging-associated tumors.

  5. Assessment of Radiation-Equivalency of Favorite Foods Based on Apoptotic and Clonogenic Cell Death

    International Nuclear Information System (INIS)

    Lee, Seunghee; Park, Hyosung; Lee, Minho; Kim, Eunhee

    2013-01-01

    In this study, the cytotoxicity of favorite foods was investigated to provide chemical doses causing bio-effects comparable with those by radiation exposure. Extracts from alcohol, cigarette and coffee have been investigated for their cytotoxicity in terms of apoptosis induction and clonogenic cell killing. The comparison of familiar favorite food items with radiation in cellular toxicity might help the public have balanced judgment on the severity of radiation exposure. Future study pursues estimating dose-rate effect and the investigation will be continued with other cell lines. People fear that ionizing radiation, despite of its usefulness, bears significant risk factor. In particular, after Fukushima nuclear accident in March of 2011, people become over-sensitive to radiation exposure and nuclear power. Such fear by the public of nuclear power inevitably costs the nuclear industry a high expense for safety measures, not to mention brings the public uneasy life. It is important that the consequence of radiation exposure is conveyed to the public in a friendly way with actual data

  6. In vitro binding and receptor-mediated activity of terlipressin at vasopressin receptors V1 and V2.

    Science.gov (United States)

    Jamil, Khurram; Pappas, Stephen Chris; Devarakonda, Krishna R

    2018-01-01

    Terlipressin, a synthetic, systemic vasoconstrictor with selective activity at vasopressin-1 (V 1 ) receptors, is a pro-drug for the endogenous/natural porcine hormone [Lys 8 ]-vasopressin (LVP). We investigated binding and receptor-mediated cellular activities of terlipressin, LVP, and endogenous human hormone [Arg 8 ]-vasopressin (AVP) at V 1 and vasopressin-2 (V 2 ) receptors. Cell membrane homogenates of Chinese hamster ovary cells expressing human V 1 and V 2 receptors were used in competitive binding assays to measure receptor-binding activity. These cells were used in functional assays to measure receptor-mediated cellular activity of terlipressin, LVP, and AVP. Binding was measured by [ 3 H]AVP counts, and the activity was measured by fluorometric detection of intracellular calcium mobilization (V 1 ) and cyclic adenosine monophosphate (V 2 ). Binding potency at V 1 and V 2 was AVP>LVP>terlipressin. LVP and terlipressin had approximately sixfold higher affinity for V 1 than for V 2 . Cellular activity potency was also AVP>LVP>terlipressin. Terlipressin was a partial agonist at V 1 and a full agonist at V 2 ; LVP was a full agonist at both V 1 and V 2 . The in vivo response to terlipressin is likely due to the partial V 1 agonist activity of terlipressin and full V 1 agonist activity of its metabolite, LVP. These results provide supportive evidence for previous findings and further establish terlipressin pharmacology for vasopressin receptors.

  7. Receptor-mediated oral delivery of a bioencapsulated green fluorescent protein expressed in transgenic chloroplasts into the mouse circulatory system.

    Science.gov (United States)

    Limaye, Arati; Koya, Vijay; Samsam, Mohtashem; Daniell, Henry

    2006-05-01

    Oral delivery of biopharmaceutical proteins expressed in plant cells should reduce their cost of production, purification, processing, cold storage, transportation, and delivery. However, poor intestinal absorption of intact proteins is a major challenge. To overcome this limitation, we investigate here the concept of receptor-mediated oral delivery of chloroplast-expressed foreign proteins. Therefore, the transmucosal carrier cholera toxin B-subunit and green fluorescent protein (CTB-GFP), separated by a furin cleavage site, was expressed via the tobacco chloroplast genome. Polymerase chain reaction (PCR) and Southern blot analyses confirmed site-specific transgene integration and homoplasmy. Immunoblot analysis and ELISA confirmed expression of monomeric and pentameric forms of CTB-GFP, up to 21.3% of total soluble proteins. An in vitro furin cleavage assay confirmed integrity of the engineered furin cleavage site, and a GM1 binding assay confirmed the functionality of CTB-GFP pentamers. Following oral administration of CTB-GFP expressing leaf material to mice, GFP was observed in the mice intestinal mucosa, liver, and spleen in fluorescence and immunohistochemical studies, while CTB remained in the intestinal cell. This report of receptor-mediated oral delivery of a foreign protein into the circulatory system opens the door for low-cost production and delivery of human therapeutic proteins.

  8. Genomic amplification of Fanconi anemia complementation group A (FancA) in head and neck squamous cell carcinoma (HNSCC): Cellular mechanisms of radioresistance and clinical relevance.

    Science.gov (United States)

    Hess, Julia; Unger, Kristian; Orth, Michael; Schötz, Ulrike; Schüttrumpf, Lars; Zangen, Verena; Gimenez-Aznar, Igor; Michna, Agata; Schneider, Ludmila; Stamp, Ramona; Selmansberger, Martin; Braselmann, Herbert; Hieber, Ludwig; Drexler, Guido A; Kuger, Sebastian; Klein, Diana; Jendrossek, Verena; Friedl, Anna A; Belka, Claus; Zitzelsberger, Horst; Lauber, Kirsten

    2017-02-01

    Radio (chemo) therapy is a crucial treatment modality for head and neck squamous cell carcinoma (HNSCC), but relapse is frequent, and the underlying mechanisms remain largely elusive. Therefore, novel biomarkers are urgently needed. Previously, we identified gains on 16q23-24 to be associated with amplification of the Fanconi anemia A (FancA) gene and to correlate with reduced progression-free survival after radiotherapy. Here, we analyzed the effects of FancA on radiation sensitivity in vitro, characterized the underlying mechanisms, and evaluated their clinical relevance. Silencing of FancA expression in HNSCC cell lines with genomic gains on 16q23-24 resulted in significantly impaired clonogenic survival upon irradiation. Conversely, overexpression of FancA in immortalized keratinocytes conferred increased survival accompanied by improved DNA repair, reduced accumulation of chromosomal translocations, but no hyperactivation of the FA/BRCA-pathway. Downregulation of interferon signaling as identified by microarray analyses, enforced irradiation-induced senescence, and elevated production of the senescence-associated secretory phenotype (SASP) appeared to be candidate mechanisms contributing to FancA-mediated radioresistance. Data of the TCGA HNSCC cohort confirmed the association of gains on 16q24.3 with FancA overexpression and impaired overall survival. Importantly, transcriptomic alterations similar to those observed upon FancA overexpression in vitro strengthened the clinical relevance. Overall, FancA amplification and overexpression appear to be crucial for radiotherapeutic failure in HNSCC. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  9. Isolation and identification of radioprotective compound (s) from radioresistant culture of Fusarium Moniliforme

    International Nuclear Information System (INIS)

    Adam, Y.M.; Attaby, H.S.H.

    2001-01-01

    Mat extract and filtrate of the radio-resistant strain Fusarium Moniliforme have been prepared and tested for their radioprotective action on the radiosensitive strain Trichoderma Viride. Both extracts gave T. Viride a protective effect against gamma radiation. Analysis of mat extracts and filtrates of both fungi, the radioresistant F. Moniliforme and the radiosensitive T. Viride revealed a pronounced differences in amino acids quantities in addition to the presence of gibberelic acid (GA 3 ) in the filtrate of F. Moniliforme only. Generally, the radioresistant fungus F. Moniliforme was able to accumulate ten amino acids in large quantity than the radiosensitive one. These amino acids comprised, cystine, glutamic acid, serine, methionine, histidine, proline, arginine, alanine, glycine and therionine. The amino acid pool of both fungi was poor in filtrate than in the mat extract. Analysis of filtration of F. Moniliforme was characterized by high content of gibberellic acid, whereas, only traces were detected in mat extract. No gibberellic acid was detected in both mat extract and filtrate of T. Viride. The results of this investigation, revealed also that both amino acids and gibberellic acid were subjected to pronounced disturbances following irradiation compared with the control. A close correlation between the induced radioresistance of T. Viride and the amounts of amino acids and G A, added to its culture medium was observed, suggesting their participation in radioprotection

  10. Radioresistance related genes screened by protein-protein interaction network analysis in nasopharyngeal carcinoma

    International Nuclear Information System (INIS)

    Zhu Xiaodong; Guo Ya; Qu Song; Li Ling; Huang Shiting; Li Danrong; Zhang Wei

    2012-01-01

    Objective: To discover radioresistance associated molecular biomarkers and its mechanism in nasopharyngeal carcinoma by protein-protein interaction network analysis. Methods: Whole genome expression microarray was applied to screen out differentially expressed genes in two cell lines CNE-2R and CNE-2 with different radiosensitivity. Four differentially expressed genes were randomly selected for further verification by the semi-quantitative RT-PCR analysis with self-designed primers. The common differentially expressed genes from two experiments were analyzed with the SNOW online database in order to find out the central node related to the biomarkers of nasopharyngeal carcinoma radioresistance. The expression of STAT1 in CNE-2R and CNE-2 cells was measured by Western blot. Results: Compared with CNE-2 cells, 374 genes in CNE-2R cells were differentially expressed while 197 genes showed significant differences. Four randomly selected differentially expressed genes were verified by RT-PCR and had same change trend in consistent with the results of chip assay. Analysis with the SNOW database demonstrated that those 197 genes could form a complicated interaction network where STAT1 and JUN might be two key nodes. Indeed, the STAT1-α expression in CNE-2R was higher than that in CNE-2 (t=4.96, P<0.05). Conclusions: The key nodes of STAT1 and JUN may be the molecular biomarkers leading to radioresistance in nasopharyngeal carcinoma, and STAT1-α might have close relationship with radioresistance. (authors)

  11. DNA repair and cytokines: TGF-beta, IL-6, and thrombopoietin as different biomarkers of radioresistance

    Directory of Open Access Journals (Sweden)

    Francesca Bianca Aiello

    2016-07-01

    Full Text Available Double strand breaks (DSBs induced by radiotherapy are highly cytotoxic lesions, leading to chromosomal aberrations and cell death. ATM-dependent DNA-damage response, non-homologous end joining, and homologous recombination pathways coordinately contribute to repairing DSBs in higher eukaryotes. It is known that the expression of DSB repair genes is increased in tumors which is one of the main reasons for radioresistance. The inhibition of DSB repair pathways may be useful to increase tumor cell radiosensitivity and may target stem cell-like cancer cells, known to be the most radioresistant tumor components. Commonly overexpressed in neoplastic cells, cytokines confer radioresistance by promoting proliferation, survival, invasion, and angiogenesis. Unfortunately, tumor irradiation increases the expression of various cytokines displaying these effects, including transforming growth factor-beta and interlukin-6. Recently the capabilities of these cytokines to support DNA repair pathways and the ATM-dependent DNA response have been demonstrated. Thrombopoietin, essential for megakaryopoiesis and very important for hematopoietic stem cell homeostasis, has also been found to promote DNA repair in a highly selective manner. These findings reveal a novel mechanism underlying cytokine-related radioresistance, which may be clinically relevant. Therapies targeting specific cytokines may be used to improve radiosensitivity. Specific inhibitors may be chosen in consideration of different tumor microenvironments. Thrombopoietin may be useful in fending off irradiation-induced loss of hematopoietic stem cells.

  12. HIF-1 and NDRG2 contribute to hypoxia-induced radioresistance of cervical cancer Hela cells

    International Nuclear Information System (INIS)

    Liu, Junye; Zhang, Jing; Wang, Xiaowu; Li, Yan; Chen, Yongbin; Li, Kangchu; Zhang, Jian; Yao, Libo; Guo, Guozhen

    2010-01-01

    Hypoxia inducible factor 1 (HIF-1), the key mediator of hypoxia signaling pathways, has been shown involved in hypoxia-induced radioresistance. However, the underlying mechanisms are unclear. The present study demonstrated that both hypoxia and hypoxia mimetic cobalt chloride could increase the radioresistance of human cervical cancer Hela cells. Meanwhile, ectopic expression of HIF-1 could enhance the resistance of Hela cells to radiation, whereas knocking-down of HIF-1 could increase the sensitivity of Hela cells to radiation in the presence of hypoxia. N-Myc downstream-regulated gene 2 (NDRG2), a new HIF-1 target gene identified in our lab, was found to be upregulated by hypoxia and radiation in a HIF-1-dependent manner. Overexpression of NDRG2 resulted in decreased sensitivity of Hela cells to radiation while silencing NDRG2 led to radiosensitization. Moreover, NDRG2 was proved to protect Hela cells from radiation-induced apoptosis and abolish radiation-induced upregulation of Bax. Taken together, these data suggest that both HIF-1 and NDRG2 contribute to hypoxia-induced tumor radioresistance and that NDRG2 acts downstream of HIF-1 to promote radioresistance through suppressing radiation-induced Bax expression. It would be meaningful to further explore the clinical application potential of HIF-1 and NDRG2 blockade as radiosensitizer for tumor therapy.

  13. GPR56/ADGRG1 Inhibits Mesenchymal Differentiation and Radioresistance in Glioblastoma

    NARCIS (Netherlands)

    Moreno, Marta; Pedrosa, Leire; Pare, Laia; Pineda, Estela; Bejarano, Leire; Martinez, Josefina; Balasubramaniyan, Veerakumar; Ezhilarasan, Ravesanker; Kallarackal, Naveen; Kim, Sung-Hak; Wang, Jia; Audia, Alessandra; Conroy, Siobhan; Marin, Mercedes; Ribalta, Teresa; Pujol, Teresa; Herreros, Antoni; Tortosa, Avelina; Mira, Helena; Alonso, Marta M.; Gomez-Manzano, Candelaria; Graus, Francesc; Sulman, Erik P.; Piao, Xianhua; Nakano, Ichiro; Prat, Aleix; Bhat, Krishna P.; de la Iglesia, Nuria

    2017-01-01

    A mesenchymal transition occurs both during the natural evolution of glioblastoma (GBM) and in response to therapy. Here, we report that the adhesion G-protein-coupled receptor, GPR56/ADGRG1, inhibits GBM mesenchymal differentiation and radioresistance. GPR56 is enriched in proneural and

  14. Beclin1-induced autophagy abrogates radioresistance of lung cancer cells by suppressing osteopontin

    International Nuclear Information System (INIS)

    Chang, Seung-Hee; Minai-Tehrani, Arash; Shin, Ji-Young

    2012-01-01

    Osteopontin (OPN) serves as an indicator of resistance to radiotherapy. However, the role of OPN in the development of acquired radioresistance in human lung cancer cells has not yet been fully elucidated. Therefore, the potential importance of OPN as a marker of lung cancer with a potential significant role in the development of radioresistance against repeated radiotherapy has prompted us to define the pathways by which OPN regulates lung cancer cell growth. In addition, autophagy has been reported to play a key role in the radiosensitization of cancer cells. Here, we report that increased OPN expression through induction of nuclear p53 following irradiation was inhibited by exogenous beclin-1 (BECN1). Our results clearly show that BECN1 gene expression led to induction of autophagy and inhibition of cancer cell growth and angiogenesis. Our results suggest that the induction of autophagy abrogated the radioresistance of the cancer cells. Interestingly, we showed that knockdown of OPN by lentivirus-mediated shRNA induced the autophagy of human lung cancer cell. Taken together, these results suggest that OPN and BECN1 can be molecular targets for overcoming radioresistance by controlling autophagy. (author)

  15. Can cell survival parameters be deduced from non-clonogenic assays of radiation damage to normal tissue

    International Nuclear Information System (INIS)

    Michalowski, A.; Wheldon, T.E.; Kirk, J.

    1984-01-01

    The relationship between dose-response curves for large scale radiation injury to tissues and survival curves for clonogenic cells is not necessarily simple. Sterilization of clonogenic cells occurs near-instantaneously compared with the protracted lag period for gross injury to tissues. Moreover, with some types of macroscopic damage, the shapes of the dose-response curves may depend on time of assay. Changes in the area or volume of irradiated tissue may also influence the shapes of these curves. The temporal pattern of expression of large scale injury also varies between tissues, and two distinct groups can be recognized. In rapidly proliferating tissues, lag period is almost independent of dose, whilst in slowly proliferating tissues, it is inversely proportional to dose. This might be explained by invoking differences in corresponding proliferative structures of the tissues. (Three compartmental Type H versus one compartmental Type F proliferative organization). For the second group of tissues particularly, mathematical modelling suggests a systematic dissociation of the dose-response curves for clonogenic cell survival and large scale injury. In particular, it may be difficult to disentangle the contributions made to inter-fraction sparing by cellular repair processes and by proliferation-related factors. (U.K.)

  16. CD133-expressing thyroid cancer cells are undifferentiated, radioresistant and survive radioiodide therapy

    International Nuclear Information System (INIS)

    Ke, Chien-Chih; Liu, Ren-Shyan; Yang, An-Hang; Liu, Ching-Sheng; Chi, Chin-Wen; Tseng, Ling-Ming; Tsai, Yi-Fan; Ho, Jennifer H.; Lee, Chen-Hsen; Lee, Oscar K.

    2013-01-01

    131 I therapy is regularly used following surgery as a part of thyroid cancer management. Despite an overall relatively good prognosis, recurrent or metastatic thyroid cancer is not rare. CD133-expressing cells have been shown to mark thyroid cancer stem cells that possess the characteristics of stem cells and have the ability to initiate tumours. However, no studies have addressed the influence of CD133-expressing cells on radioiodide therapy of the thyroid cancer. The aim of this study was to investigate whether CD133 + cells contribute to the radioresistance of thyroid cancer and thus potentiate future recurrence and metastasis. Thyroid cancer cell lines were analysed for CD133 expression, radiosensitivity and gene expression. The anaplastic thyroid cancer cell line ARO showed a higher percentage of CD133 + cells and higher radioresistance. After γ-irradiation of the cells, the CD133 + population was enriched due to the higher apoptotic rate of CD133 - cells. In vivo 131 I treatment of ARO tumour resulted in an elevated expression of CD133, Oct4, Nanog, Lin28 and Glut1 genes. After isolation, CD133 + cells exhibited higher radioresistance and higher expression of Oct4, Nanog, Sox2, Lin28 and Glut1 in the cell line or primarily cultured papillary thyroid cancer cells, and lower expression of various thyroid-specific genes, namely NIS, Tg, TPO, TSHR, TTF1 and Pax8. This study demonstrates the existence of CD133-expressing thyroid cancer cells which show a higher radioresistance and are in an undifferentiated status. These cells possess a greater potential to survive radiotherapy and may contribute to the recurrence of thyroid cancer. A future therapeutic approach for radioresistant thyroid cancer may focus on the selective eradication of CD133 + cells. (orig.)

  17. Evidence and analysis of radioresistance induced by protracted gamma irradiation of Chlorella pyrenoidosa chick, green unicellular alga

    International Nuclear Information System (INIS)

    Santier-Riviere, S.

    1984-06-01

    Chlorella cells, unicellular green algae, are a suitable living material to study radiosensitivity of eucaryotic cells after acute or protracted gamma irradiations. Cell survival and survival curves are taken as end-points. Methods of irradiation were defined taking in account interferences of the different factors which can intervene during the experimentation. Survival curves after protracted irradiation of Chlorella cell cultures in plateau-phase have a shape that can be explained by radioresistance. The population of surviving cells becomes radioresistant in front of protracted and acute irradiations, acute irradiation allowing us to analyze radioresistance. Radioresistance increases with the total dose of protracted irradiation. The decrease of radiosensitivity with aging of cells is not able to explain the phenomenon. It is not due to selection of radioresistance cells by protracted irradiation. All the cells get radioresistance. Radioresistance decreases with the time when protracted irradiation is suppressed. It is not found in offspring. It is not a mutation but perhaps the effect of a stimulation of repair processes, but not potentially lethal damage repair [fr

  18. Halothane inhibits the cholinergic-receptor-mediated influx of calcium in primary culture of bovine adrenal medulla cells

    International Nuclear Information System (INIS)

    Yashima, N.; Wada, A.; Izumi, F.

    1986-01-01

    Adrenal medulla cells are cholinoceptive cells. Stimulation of the acetylcholine receptor causes the influx of Ca to the cells, and Ca acts as the coupler of the stimulus-secretion coupling. In this study, the authors investigated the effects of halothane on the receptor-mediated influx of 45 Ca using cultured bovine adrenal medulla cells. Halothane at clinical concentrations (0.5-2%) inhibited the influx of 45 Ca caused by carbachol, with simultaneous inhibition of catecholamine secretion. The influx of 45 Ca and the secretion of catecholamines caused by K depolarization were inhibited by a large concentration of Mg, which competes with Ca at Ca channels, but not inhibited by halothane. Inhibition of the 45 Ca influx by halothane was not overcome by increase in the carbachol concentration. Inhibition of the 45 Ca influx by halothane was examined in comparison with that caused by a large concentration of Mg by the application of Scatchard analysis as the function of the external Ca concentration. Halothane decreased the maximal influx of 45 Ca without altering the apparent kinetic constant of Ca to Ca channels. On the contrary, a large concentration of Mg increased the apparent kinetic constant without altering the maximal influx of 45 Ca. Based on these findings, the authors suggest that inhibition of the 45 Ca influx by halothane was not due to the direct competitive inhibition of Ca channels, nor to the competitive antagonism of agonist-receptor interaction. As a possibility, halothane seems to inhibit the receptor-mediated activation of Ca channels through the interference of coupling between the receptor and Ca channels

  19. Reboxetine Enhances the Olanzapine-Induced Antipsychotic-Like Effect, Cortical Dopamine Outflow and NMDA Receptor-Mediated Transmission

    Science.gov (United States)

    Marcus, Monica M; Jardemark, Kent; Malmerfelt, Anna; Björkholm, Carl; Svensson, Torgny H

    2010-01-01

    Preclinical data have shown that addition of the selective norepinephrine transporter (NET) inhibitor reboxetine increases the antipsychotic-like effect of the D2/3 antagonist raclopride and, in parallel, enhances cortical dopamine output. Subsequent clinical results suggested that adding reboxetine to stable treatments with various antipsychotic drugs (APDs) may improve positive, negative and depressive symptoms in schizophrenia. In this study, we investigated in rats the effects of adding reboxetine to the second-generation APD olanzapine on: (i) antipsychotic efficacy, using the conditioned avoidance response (CAR) test, (ii) extrapyramidal side effect (EPS) liability, using a catalepsy test, (iii) dopamine efflux in the medial prefrontal cortex and the nucleus accumbens, using in vivo microdialysis in freely moving animals and (iv) cortical N-methyl--aspartate (NMDA) receptor-mediated transmission, using intracellular electrophysiological recording in vitro. Reboxetine (6 mg/kg) enhanced the suppression of CAR induced by a suboptimal dose (1.25 mg/kg), but not an optimal (2.5 mg/kg) dose of olanzapine without any concomitant catalepsy. Addition of reboxetine to the low dose of olanzapine also markedly increased cortical dopamine outflow and facilitated prefrontal NMDA receptor-mediated transmission. Our data suggest that adjunctive treatment with a NET inhibitor may enhance the therapeutic effect of low-dose olanzapine in schizophrenia without increasing EPS liability and add an antidepressant action, thus in principle allowing for a dose reduction of olanzapine with a concomitant reduction of dose-related side effects, such as EPS and weight gain. PMID:20463659

  20. Prostaglandin E2 potentiation of P2X3 receptor mediated currents in dorsal root ganglion neurons

    Directory of Open Access Journals (Sweden)

    Huang Li-Yen

    2007-08-01

    Full Text Available Abstract Prostaglandin E2 (PGE2 is a well-known inflammatory mediator that enhances the excitability of DRG neurons. Homomeric P2X3 and heteromeric P2X2/3 receptors are abundantly expressed in dorsal root ganglia (DRG neurons and participate in the transmission of nociceptive signals. The interaction between PGE2 and P2X3 receptors has not been well delineated. We studied the actions of PGE2 on ATP-activated currents in dissociated DRG neurons under voltage-clamp conditions. PGE2 had no effects on P2X2/3 receptor-mediated responses, but significantly potentiated fast-inactivating ATP currents mediated by homomeric P2X3 receptors. PGE2 exerted its action by activating EP3 receptors. To study the mechanism underlying the action of PGE2, we found that the adenylyl cyclase activator, forskolin and the membrane-permeable cAMP analogue, 8-Br-cAMP increased ATP currents, mimicking the effect of PGE2. In addition, forskolin occluded the enhancement produced by PGE2. The protein kinase A (PKA inhibitors, H89 and PKA-I blocked the PGE2 effect. In contrast, the PKC inhibitor, bisindolymaleimide (Bis did not change the potentiating action of PGE2. We further showed that PGE2 enhanced α,β-meATP-induced allodynia and hyperalgesia and the enhancement was blocked by H89. These observations suggest that PGE2 binds to EP3 receptors, resulting in the activation of cAMP/PKA signaling pathway and leading to an enhancement of P2X3 homomeric receptor-mediated ATP responses in DRG neurons.

  1. Electroacupuncture improves cerebral blood flow and attenuates moderate ischemic injury via Angiotensin II its receptors-mediated mechanism in rats.

    Science.gov (United States)

    Li, Jing; He, Jiaojun; Du, Yuanhao; Cui, Jingjun; Ma, Ying; Zhang, Xuezhu

    2014-11-11

    To investigate the effects and potential mechanism of electroacupuncture intervention on expressions of Angiotensin II and its receptors-mediated signaling pathway in experimentally induced cerebral ischemia. Totally 126 male Wistar rats were randomly divided into control group, model group and EA group. The latter two were further divided into ten subgroups (n = 6) following Middle Cerebral Artery Occlusion (MCAO). Changes in regional cerebral blood flow (rCBF) and expressions of Angiotensin II and its receptors (AT1R, AT2R), as well as effector proteins in phosphatidyl inositol signal pathway were monitored before and at different times after MCAO. MCAO-induced decline of ipsilateral rCBF was partially suppressed by electroacupuncture, and contralateral blood flow was also superior to that of model group. Angiotensin II level was remarkably elevated immediately after MCAO, while electroacupuncture group exhibited significantly lower levels at 1 to 3 h and the value was significantly increased thereafter. The enhanced expression of AT1R was partially inhibited by electroacupuncture, while increased AT2R level was further induced. Electroacupuncture stimulation attenuated and postponed the upregulated-expressions of Gq and CaM these upregulations. ELISA results showed sharply increased expressions of DAG and IP3, which were remarkably neutralized by electroacupuncture. MCAO induced significant increases in expression of Angiotensin II and its receptor-mediated signal pathway. These enhanced expressions were significantly attenuated by electroacupuncture intervention, followed by reduced vasoconstriction and improved blood supply in ischemic region, and ultimately conferred beneficial effects on cerebral ischemia.

  2. Loss of Progesterone Receptor-Mediated Actions Induce Preterm Cellular and Structural Remodeling of the Cervix and Premature Birth

    Science.gov (United States)

    Yellon, Steven M.; Dobyns, Abigail E.; Beck, Hailey L.; Kurtzman, James T.; Garfield, Robert E.; Kirby, Michael A.

    2013-01-01

    A decline in serum progesterone or antagonism of progesterone receptor function results in preterm labor and birth. Whether characteristics of premature remodeling of the cervix after antiprogestins or ovariectomy are similar to that at term was the focus of the present study. Groups of pregnant rats were treated with vehicle, a progesterone receptor antagonist (onapristone or mifepristone), or ovariectomized on day 17 postbreeding. As expected, controls given vehicle delivered at term while rats delivered preterm after progesterone receptor antagonist treatment or ovariectomy. Similar to the cervix before term, the preterm cervix of progesterone receptor antagonist-treated rats was characterized by reduced cell nuclei density, decreased collagen content and structure, as well as a greater presence of macrophages per unit area. Thus, loss of nuclear progesterone receptor-mediated actions promoted structural remodeling of the cervix, increased census of resident macrophages, and preterm birth much like that found in the cervix at term. In contrast to the progesterone receptor antagonist-induced advance in characteristics associated with remodeling, ovariectomy-induced loss of systemic progesterone did not affect hypertrophy, extracellular collagen, or macrophage numbers in the cervix. Thus, the structure and macrophage census in the cervix appear sufficient for premature ripening and birth to occur well before term. With progesterone receptors predominantly localized on cells other than macrophages, the findings suggest that interactions between cells may facilitate the loss of progesterone receptor-mediated actions as part of a final common mechanism that remodels the cervix in certain etiologies of preterm and with parturition at term. PMID:24339918

  3. Prognostic factors for outcomes after whole-brain irradiation of brain metastases from relatively radioresistant tumors: a retrospective analysis

    NARCIS (Netherlands)

    Meyners, T.; Heisterkamp, C.; Kueter, J.D.; Veninga, T.; Stalpers, L.J.A.; Schild, S.E.; Rades, D.

    2010-01-01

    Background: This study investigated potential prognostic factors in patients treated with whole-brain irradiation (WBI) alone for brain metastases from relatively radioresistant tumors such as malignant melanoma, renal cell carcinoma, and colorectal cancer. Additionally, a potential benefit from

  4. Prognostic factors for outcomes after whole-brain irradiation of brain metastases from relatively radioresistant tumors: a retrospective analysis

    NARCIS (Netherlands)

    Meyners, Thekla; Heisterkamp, Christine; Kueter, Jan-Dirk; Veninga, Theo; Stalpers, Lukas J. A.; Schild, Steven E.; Rades, Dirk

    2010-01-01

    This study investigated potential prognostic factors in patients treated with whole-brain irradiation (WBI) alone for brain metastases from relatively radioresistant tumors such as malignant melanoma, renal cell carcinoma, and colorectal cancer. Additionally, a potential benefit from escalating the

  5. Phenobarbital but not diazepam reduces AMPA/Kainate receptor mediated currents and exerts opposite actions on initial seizures in the neonatal rat hippocampus

    Directory of Open Access Journals (Sweden)

    Romain eNardou

    2011-07-01

    Full Text Available Diazepam (DZP and phenobarbital (PB are extensively used as first and second line drugs to treat acute seizures in neonates and their actions are thought to be mediated by increasing the actions of GABAergic signals. Yet, their efficacy is variable with occasional failure or even aggravation of recurrent seizures questioning whether other mechanisms are not involved in their actions. We have now compared the effects of DZP and PB on ictal-like events (ILEs in an in vitro model of mirror focus (MF. Using the three-compartment chamber with the two immature hippocampi and their commissural fibers placed in 3 different compartments, kainate was applied to one hippocampus and PB or DZP to the contralateral one, either after one ILE or after many recurrent ILEs that produce an epileptogenic MF. We report that in contrast to PB, DZP aggravated propagating ILEs from the start and did not prevent the formation of MF. PB reduced and DZP increased the network driven Giant Depolarising Potentials suggesting that PB may exert additional actions that are not mediated by GABA signalling. In keeping with this, PB but not DZP reduced field potentials recorded in the presence of GABA and NMDA receptor antagonists. These effects are mediated by a direct action on AMPA/Kainate receptors since PB: i reduced AMPA/Kainate receptor mediated currents induced by focal applications of glutamate ; ii reduced the amplitude and the frequency of AMPA but not NMDA receptor mediated miniature EPSCs; iii augmented the number of AMPA receptor mediated EPSCs failures evoked by minimal stimulation. These effects persisted in MF. Therefore, PB exerts its anticonvulsive actions partly by reducing AMPA/Kainate receptors mediated EPSCs in addition to the pro-GABA effects. We suggest that PB may have advantage over DZP in the treatment of initial neonatal seizures since the additional reduction of glutamate receptors mediated signals may reduce the severity of neonatal seizures.

  6. The nature of variations in the radioresistance of sea urchin eggs in the early embryogenesis

    International Nuclear Information System (INIS)

    Konstantinova, M.M.; Tarasenko, A.G.; Nekrasova, I.V.; Dontsova, G.V.

    1975-01-01

    The synchronously developing sea urchin eggs (Strongylocentrotus nudus) have been used for the experiment. Radioprotective efficiency of cysteamine (applied in the concentrations ranging from 1.10 -5 to 5.10 -3 M) has been studied at two stages of mitosis: metaphase and telophase. Cysteamine (5.10 -3 M) has been found to level the variations in radiosensitivity of these phases producing the strongest effect in the radiosensitive phase (telophase) and the slightest effect in the radioresistant phase (metaphase). Determination of the relative protective effectiveness of cysteamine in relation to the original radiosensitivity of a cell, and, hence, to its thiol content, has demonstrated that the higher the radiosensitivity of a cell at the time of the exposure the higher the relative protective efficiency of cysteamine. The data obtained evidence of the idea that thiols play an essential role in the determination of radioresistance of different stages of mitosis of the developing sea urchin eggs

  7. Transmission of in-vitro radioresistance in a cancer-prone family

    International Nuclear Information System (INIS)

    Bech-Hansen, N.T.; Blattner, W.A.; Sell, B.M.; McKeen, E.A.; Lampkin, B.C.; Fraumeni, J.F. Jr.; Paterson, M.C.

    1981-01-01

    Neoplasms of possible radiogenic origin developed in two members of a family prone to a diversity of cancers. Gamma-irradiation survival studies in these two patients and three other relatives, but not their spouses, over three generations demonstrated resistance to cell killing. The D 10 value (radiation dose required to reduce survival to 10%) was significantly higher for the five radioresistant strains (491 +- 30 rad) than for control cultures (405 +- 18 rad). There was a significant correlation between individual D 10 values and D 0 survival-curve parameters, indicating that changes in the exponential slope of the survival curves accounted for much of the increase in D 10 values. This novel radiation phenotype could be a manifestation of a basic cellular defect, predisposing to a variety of tumours in family members. Thus in-vitro radioresistance, like radiosensitivity, may be a phenotype of a mechanism that increases cancer risk in man. (author)

  8. Highly radioresistant aramid fiber as a concrete-reinforcing material. Development of reinforced compound materials

    International Nuclear Information System (INIS)

    Udagawa, Akira; Moriya, Toshio.

    1997-01-01

    Nuclear installations, such as nuclear fusion reactor always receive strong influence from magnetic field. There, stray current is induced by the changes in magnetic fields among iron rods of the construction, resulting that the plasma control magnetic field might be disturbed. As the countermeasures for these troubles, iron rods mixed with non-magnetic Mn-steel have been used in JAERI, but it is insufficient to completely prevent such electromagnetic damages. Thus, aramid fiber reinforced plastics (ArFRP) was paid an attention as a concrete-reinforcing material. JAERI has been attempting to develop a radioresistant ArFRP jointly with Mitsui Construction Co., Ltd. and a highly efficient producing process of ArFRP was developed. The product had superior properties in respects of radioresistancy, heat-resistancy and durability. The properties of newly developed ArFRP rods were compared with those of the conventional ArFRP and iron rods. (M.N.)

  9. Clonogenic growth of human breast cancer cells co-cultured in direct contact with serum-activated fibroblasts

    International Nuclear Information System (INIS)

    Samoszuk, Michael; Tan, Jenny; Chorn, Guillaume

    2005-01-01

    Accumulating evidence suggests that fibroblasts play a pivotal role in promoting the growth of breast cancer cells. The objective of the present study was to characterize and validate an in vitro model of the interaction between small numbers of human breast cancer cells and human fibroblasts. We measured the clonogenic growth of small numbers of human breast cancer cells co-cultured in direct contact with serum-activated, normal human fibroblasts. Using DNA microarrays, we also characterized the gene expression profile of the serum-activated fibroblasts. In order to validate the in vivo relevance of our experiments, we then analyzed clinical samples of metastatic breast cancer for the presence of myofibroblasts expressing α-smooth muscle actin. Clonogenic growth of human breast cancer cells obtained directly from in situ and invasive tumors was dramatically and consistently enhanced when the tumor cells were co-cultured in direct contact with serum-activated fibroblasts. This effect was abolished when the cells were co-cultured in transwells separated by permeable inserts. The fibroblasts in our experimental model exhibited a gene expression signature characteristic of 'serum response' (i.e. myofibroblasts). Immunostaining of human samples of metastatic breast cancer tissue confirmed that myofibroblasts are in direct contact with breast cancer cells. Serum-activated fibroblasts promote the clonogenic growth of human breast cancer cells in vitro through a mechanism that involves direct physical contact between the cells. This model shares many important molecular and phenotypic similarities with the fibroblasts that are naturally found in breast cancers

  10. Novel histone deacetylase inhibitor CG200745 induces clonogenic cell death by modulating acetylation of p53 in cancer cells.

    Science.gov (United States)

    Oh, Eun-Taex; Park, Moon-Taek; Choi, Bo-Hwa; Ro, Seonggu; Choi, Eun-Kyung; Jeong, Seong-Yun; Park, Heon Joo

    2012-04-01

    Histone deacetylase (HDAC) plays an important role in cancer onset and progression. Therefore, inhibition of HDAC offers potential as an effective cancer treatment regimen. CG200745, (E)-N(1)-(3-(dimethylamino)propyl)-N(8)-hydroxy-2-((naphthalene-1-loxy)methyl)oct-2-enediamide, is a novel HDAC inhibitor presently undergoing a phase I clinical trial. Enhancement of p53 acetylation by HDAC inhibitors induces cell cycle arrest, differentiation, and apoptosis in cancer cells. The purpose of the present study was to investigate the role of p53 acetylation in the cancer cell death caused by CG200745. CG200745-induced clonogenic cell death was 2-fold greater in RKO cells expressing wild-type p53 than in p53-deficient RC10.1 cells. CG200745 treatment was also cytotoxic to PC-3 human prostate cancer cells, which express wild-type p53. CG200745 increased acetylation of p53 lysine residues K320, K373, and K382. CG200745 induced the accumulation of p53, promoted p53-dependent transactivation, and enhanced the expression of MDM2 and p21(Waf1/Cip1) proteins, which are encoded by p53 target genes. An examination of CG200745 effects on p53 acetylation using cells transfected with various p53 mutants showed that cells expressing p53 K382R mutants were significantly resistant to CG200745-induced clonogenic cell death compared with wild-type p53 cells. Moreover, p53 transactivation in response to CG200745 was suppressed in all cells carrying mutant forms of p53, especially K382R. Taken together, these results suggest that acetylation of p53 at K382 plays an important role in CG200745-induced p53 transactivation and clonogenic cell death.

  11. Clinically relevant radioresistant cells efficiently repair DNA double-strand breaks induced by X-rays

    International Nuclear Information System (INIS)

    Kuwahara, Yoshikazu; Baba, Taisuke; Nakagawa, Hironobu; Shimura, Tsutomu; Yamamoto, Yoichiro; Fukumoto, Manabu; Li Li; Ohkubo, Yasuhito

    2009-01-01

    Radiotherapy is one of the major therapeutic modalities for eradicating malignant tumors. However, the existence of radioresistant cells remains one of the most critical obstacles in radiotherapy and radiochemotherapy. Standard radiotherapy for tumor treatment consists of approximately 2 Gy once a day, 5 days a week, over a period of 5-8 weeks. To understand the characteristics of radioresistant cells and to develop more effective radiotherapy, we established a novel radioresistant cell line, HepG2-8960-R with clinical relevance from parental HepG2 cells by long-term fractionated exposure to 2 Gy of X-rays. HepG2-8960-R cells continued to proliferate with daily exposure to 2 Gy X-rays for more than 30 days, while all parental HepG2 cells ceased. After exposure to fractionated 2 Gy X-rays, induction frequencies of micronuclei and remaining foci of γ-H2AX in HepG2-8960-R were less than those in HepG2. Flow cytometric analysis revealed that the proportion of cells in S- and G2/M-phase of the cell cycle was higher in HepG2-8960-R than in HepG2. These suggest that the response of clinically relevant radioresistant (CRR) cells to fractionated radiation is not merely an accumulated response to each fractionated radiation. This is the first report on the establishment of a CRR cell line from an isogenic parental cell line. (author)

  12. Alterations in gene expression profiles between radioresistant and radiosensitive cell lines

    International Nuclear Information System (INIS)

    Zhou Fuxiang; Zhou Yunfeng; Xie Conghua; Dai Jing; Cao Zhen; Yu Haijun; Liao Zhengkai; Luo Zhiguo

    2007-01-01

    Objective: To study the-difference of gene expressions by the contrastive model including the cells with same pathological origin and genetic background, but definitely different radioresponse, and to find the main molecular targets related to radiosensitivity. Methods: Human larynx squamous carcinoma cell, Hep -2 was irradiated with dose of 637 cGy repeatedly to establish a radioresistant daughter cell line. The radiobiology characteristics were obtained using clone forming assay. The difference of gene expression between parent and daughter cells was detected by cDNA microarray using two different arrays including 14000 genes respectively. Results: A radioresistant cell strain Hep-2R was isolated from its parental strain Hep-2 cell. The SF 2 , D 0 , α, β for Hep-2R cell line were 0.6798, 3.24, 0.2951 and 0.0363, respectively, while 0.4148, 2.06, 0.1074 and 0.0405 for Hep-2, respectively (for SF 2 , χ 2 =63.957, P<0.001). Compared with Hep-2 cells, the expressions of 41 genes were significantly altered in the radioresistant Hep-2R cells, including 22 genes up-regulated and 19 genes down-regulated, which were involved in DNA repair, regulation of the cell cycle, cell proliferation, cytoskeleton, protein synthesis, cellular metabolism and especially apoptosis which is responsible for the different radiosensitivity between these two larynx cancer cells. The telomere protection protein gene, POT1, was the mostly up-regulated by 3.348 times. Conclusions: There is difference of gene expression between the radioresistant contrastive models. POT1 gene may be the target of radiosensitization. (authors)

  13. γ-irradiation induces radioresistant DNA synthesis in HeLa cells

    International Nuclear Information System (INIS)

    Synzynys, B.I.; Aminev, A.G.; Konstantinova, S.A.; Saenko, A.S.

    1987-01-01

    Cells of suspension HeLa culture at the logarithmic phase of growth were exposed to 60 Co-γ-rays (5 Gy), incubated in the nutritious medium, and in 4 h subjected to repeated irradiation: the dose-response function and the dynamics of DNA synthesis inhibition were determined. It was shown that DNA synthesis was inhibited to a lesser extent after preirradiation, in other words, DNA synthesis was radioresistant. A correlation between this synthesis and reproductive cell death is discussed

  14. Factors implicated to radioresistance of breast cancer and their possible roles

    International Nuclear Information System (INIS)

    Yan Weili; Huang Gang

    2006-01-01

    Radiotherapy plays an important role in the management of breast cancer. The recurrence of breast cancer after radiotherapy is considered to be related with radioresistance in breast cancer cells. Various factors, extranuclear and intranuclear, such as insulin-like growth factor-1 receptor, phosphatidylinositol 3-kinase pathway, epidermal growth factor, human epidermal growth factor receptors, p53, c-erb B2, Bcl-2, BRCA1, BRCA2, telomeres and gene expression signature, that have been implicated to influence the radiation response. (authors)

  15. Recovery and radio-resistance in mice after external irradiation; Restauration et radio-resistance chez la souris apres irradiation externe

    Energy Technology Data Exchange (ETDEWEB)

    Le Guillou, S [Commissariat a l' Energie Atomique, Fontenay-aux-Roses (France). Centre d' Etudes Nucleaires

    1965-07-01

    The author presents a literature study concerning recovery from external irradiation and an analysis of experimental data (which appear to suggest the idea of a radio-resistance in animals), as well as the hypotheses put forward for explaining this phenomenon. The author then describes an experiment carried out on mice whose LD 50/30 days increased from 1005 to 1380 rads and for which it was shown that an increase occurs in the number of certain anti-bodies circulating after a low dose of {gamma} irradiation. (author) [French] L'auteur presente une etude bibliographique de la restauration apres irradiation externe et une analyse des donnees experimentales qui paraissent suggerer la notion de radioresistance chez les animaux ainsi que les hypotheses cherchant a expliquer ce phenomene. Il relate ensuite une experience realisee sur des souris dont la DL 50/30 jours est passee de 1005 a 1380 rads et dans laquelle est montree l'augmentation de certains anticorps circulant apres une faible dose d'irradiation gamma. (auteur)

  16. Recovery and radio-resistance in mice after external irradiation; Restauration et radio-resistance chez la souris apres irradiation externe

    Energy Technology Data Exchange (ETDEWEB)

    Le Guillou, S. [Commissariat a l' Energie Atomique, Fontenay-aux-Roses (France). Centre d' Etudes Nucleaires

    1965-07-01

    The author presents a literature study concerning recovery from external irradiation and an analysis of experimental data (which appear to suggest the idea of a radio-resistance in animals), as well as the hypotheses put forward for explaining this phenomenon. The author then describes an experiment carried out on mice whose LD 50/30 days increased from 1005 to 1380 rads and for which it was shown that an increase occurs in the number of certain anti-bodies circulating after a low dose of {gamma} irradiation. (author) [French] L'auteur presente une etude bibliographique de la restauration apres irradiation externe et une analyse des donnees experimentales qui paraissent suggerer la notion de radioresistance chez les animaux ainsi que les hypotheses cherchant a expliquer ce phenomene. Il relate ensuite une experience realisee sur des souris dont la DL 50/30 jours est passee de 1005 a 1380 rads et dans laquelle est montree l'augmentation de certains anticorps circulant apres une faible dose d'irradiation gamma. (auteur)

  17. Hypoxia-inducible factor 1–mediated characteristic features of cancer cells for tumor radioresistance

    International Nuclear Information System (INIS)

    Harada, Hiroshi

    2016-01-01

    Tumor hypoxia has been attracting increasing attention in the fields of radiation biology and oncology since Thomlinson and Gray detected hypoxic cells in malignant solid tumors and showed that they exert a negative impact on the outcome of radiation therapy. This unfavorable influence has, at least partly, been attributed to cancer cells acquiring a radioresistant phenotype through the activation of the transcription factor, hypoxia-inducible factor 1 (HIF-1). On the other hand, accumulating evidence has recently revealed that, even though HIF-1 is recognized as an important regulator of cellular adaptive responses to hypoxia, it may not become active and induce tumor radioresistance under hypoxic conditions only. The mechanisms by which HIF-1 is activated in cancer cells not only under hypoxic conditions, but also under normoxic conditions, through cancer-specific genetic alterations and the resultant imbalance in intermediate metabolites have been summarized herein. The relevance of the HIF-1–mediated characteristic features of cancer cells, such as the production of antioxidants through reprogramming of the glucose metabolic pathway and cell cycle regulation, for tumor radioresistance has also been reviewed

  18. Sensitization of Radioresistant Prostate Cancer Cells by Resveratrol Isolated from Arachis hypogaea Stems.

    Science.gov (United States)

    Chen, Yu-An; Lien, Hsiu-Man; Kao, Min-Chuan; Lo, U-Ging; Lin, Li-Chiung; Lin, Chun-Jung; Chang, Sheau-Jiun; Chen, Chia-Chang; Hsieh, Jer-Tsong; Lin, Ho; Tang, Chih-Hsin; Lai, Chih-Ho

    2017-01-01

    Resveratrol (RV, 3,4',5-trihydroxystilbene) is naturally produced by a wide variety of plants including grapes and peanuts (Arachis hypogaea). However, the yield of RV from peanut stem and its potential radiosensitizing effects in prostate cancer (PCa) have not been well investigated. In this study, we characterized RV in peanut stem extract (PSE) for the first time and showed that both RV and PSE dose-dependently induced cell death in DOC-2/DAB2 interactive protein (DAB2IP)-deficient PCa cells with the radioresistant phenotype. Furthermore, the combination of radiation with either RV or PSE induced the death of radioresistant PCa cells through delayed repair of radiation-induced DNA double-strand break (DSB) and prolonged G2/M arrest, which induced apoptosis. The administration of RV and PSE effectively enhanced radiation therapy in the shDAB2IP PCa xenograft mouse model. These results demonstrate the promising synergistic effect of RV and PSE combined with radiation in the treatment of radioresistant PCa.

  19. GPR56/ADGRG1 Inhibits Mesenchymal Differentiation and Radioresistance in Glioblastoma

    Directory of Open Access Journals (Sweden)

    Marta Moreno

    2017-11-01

    Full Text Available A mesenchymal transition occurs both during the natural evolution of glioblastoma (GBM and in response to therapy. Here, we report that the adhesion G-protein-coupled receptor, GPR56/ADGRG1, inhibits GBM mesenchymal differentiation and radioresistance. GPR56 is enriched in proneural and classical GBMs and is lost during their transition toward a mesenchymal subtype. GPR56 loss of function promotes mesenchymal differentiation and radioresistance of glioma initiating cells both in vitro and in vivo. Accordingly, a low GPR56-associated signature is prognostic of a poor outcome in GBM patients even within non-G-CIMP GBMs. Mechanistically, we reveal GPR56 as an inhibitor of the nuclear factor kappa B (NF-κB signaling pathway, thereby providing the rationale by which this receptor prevents mesenchymal differentiation and radioresistance. A pan-cancer analysis suggests that GPR56 might be an inhibitor of the mesenchymal transition across multiple tumor types beyond GBM.

  20. Prenatal Alcohol Exposure Increases Histamine H3 Receptor-Mediated Inhibition of Glutamatergic Neurotransmission in Rat Dentate Gyrus.

    Science.gov (United States)

    Varaschin, Rafael K; Allen, Nyika A; Rosenberg, Martina J; Valenzuela, C Fernando; Savage, Daniel D

    2018-02-01

    We have reported that prenatal alcohol exposure (PAE)-induced deficits in dentate gyrus, long-term potentiation (LTP), and memory are ameliorated by the histamine H 3 receptor inverse agonist ABT-239. Curiously, ABT-239 did not enhance LTP or memory in control offspring. Here, we initiated an investigation of how PAE alters histaminergic neurotransmission in the dentate gyrus and other brain regions employing combined radiohistochemical and electrophysiological approaches in vitro to examine histamine H 3 receptor number and function. Long-Evans rat dams voluntarily consumed either a 0% or 5% ethanol solution 4 hours each day throughout gestation. This pattern of drinking, which produces a mean peak maternal serum ethanol concentration of 60.8 ± 5.8 mg/dl, did not affect maternal weight gain, litter size, or offspring birthweight. Radiohistochemical studies in adult offspring revealed that specific [ 3 H]-A349821 binding to histamine H 3 receptors was not different in PAE rats compared to controls. However, H 3 receptor-mediated G i /G o protein-effector coupling, as measured by methimepip-stimulated [ 35 S]-GTPγS binding, was significantly increased in cerebral cortex, cerebellum, and dentate gyrus of PAE rats compared to control. A LIGAND analysis of detailed methimepip concentration-response curves in dentate gyrus indicated that PAE significantly elevates receptor-effector coupling by a lower affinity H 3 receptor population without significantly altering the affinities of H 3 receptor subpopulations. In agreement with the [ 35 S]-GTPγS studies, a similar range of methimepip concentrations also inhibited electrically evoked field excitatory postsynaptic potential responses and increased paired-pulse ratio, a measure of decreased glutamate release, to a significantly greater extent in dentate gyrus slices from PAE rats than in controls. These results suggest that a PAE-induced elevation in H 3 receptor-mediated inhibition of glutamate release from

  1. Automatic cell cloning assay for determining the clonogenic capacity of cancer and cancer stem-like cells.

    Science.gov (United States)

    Fedr, Radek; Pernicová, Zuzana; Slabáková, Eva; Straková, Nicol; Bouchal, Jan; Grepl, Michal; Kozubík, Alois; Souček, Karel

    2013-05-01

    The clonogenic assay is a well-established in vitro method for testing the survival and proliferative capability of cells. It can be used to determine the cytotoxic effects of various treatments including chemotherapeutics and ionizing radiation. However, this approach can also characterize cells with different phenotypes and biological properties, such as stem cells or cancer stem cells. In this study, we implemented a faster and more precise method for assessing the cloning efficiency of cancer stem-like cells that were characterized and separated using a high-speed cell sorter. Cell plating onto a microplate using an automatic cell deposition unit was performed in a single-cell or dilution rank mode by the fluorescence-activated cell sorting method. We tested the new automatic cell-cloning assay (ACCA) on selected cancer cell lines and compared it with the manual approach. The obtained results were also compared with the results of the limiting dilution assay for different cell lines. We applied the ACCA to analyze the cloning capacity of different subpopulations of prostate and colon cancer cells based on the expression of the characteristic markers of stem (CD44 and CD133) and cancer stem cells (TROP-2, CD49f, and CD44). Our results revealed that the novel ACCA is a straightforward approach for determining the clonogenic capacity of cancer stem-like cells identified in both cell lines and patient samples. Copyright © 2013 International Society for Advancement of Cytometry.

  2. No supra-additive effects of goserelin and radiotherapy on clonogenic survival of prostate carcinoma cells in vitro

    Directory of Open Access Journals (Sweden)

    Thelen Paul

    2007-08-01

    Full Text Available Abstract Background Oncological results of radiotherapy for locally advanced prostate cancer (PC are significantly improved by simultaneous application of LHRH analoga (e.g. goserelin. As 85% of PC express LHRH receptors, we investigated the interaction of goserelin incubation with radiotherapy under androgen-deprived conditions in vitro. Methods LNCaP and PC-3 cells were stained for LHRH receptors. Downstream the LHRH receptor, changes in protein expression of c-fos, phosphorylated p38 and phosphorylated ERK1/2 were analyzed by means of Western blotting after incubation with goserelin and irradiation with 4 Gy. Both cell lines were incubated with different concentrations of goserelin in hormone-free medium. 12 h later cells were irradiated (0 – 4 Gy and after 12 h goserelin was withdrawn. Endpoints were clonogenic survival and cell viability (12 h, 36 h and 60 h after irradiation. Results Both tested cell lines expressed LHRH-receptors. Changes in protein expression demonstrated the functional activity of goserelin in the tested cell lines. Neither in LNCaP nor in PC-3 any significant effects of additional goserelin incubation on clonogenic survival or cell viability for all tested concentrations in comparison to radiation alone were seen. Conclusion The clinically observed increase in tumor control after combination of goserelin with radiotherapy in PC cannot be attributed to an increase in radiosensitivity of PC cells by goserelin in vitro.

  3. Acquired radioresistance of cancer and the AKT/GSK3β/cyclin D1 overexpression cycle

    International Nuclear Information System (INIS)

    Shimura, Tsutomu

    2011-01-01

    Fractionated radiotherapy (RT) is widely used in cancer therapy for its advantages in the preservation of normal tissues. However, repopulation of surviving tumor cells during fractionated RT limits the efficacy of RT. In fact, repopulating tumors often acquire radioresistance and this is the major cause of failure of RT. We have recently demonstrated that human tumor cells acquire radioresistance when exposed to fractionated radiation (FR) of X-rays every 12 hours for 1 month. The acquired radioresistance was associated with overexpression of cyclin D1, a result of a series of molecular changes; constitutive activation of DNA-PK and AKT with concomitant down-regulation of glycogen synthase kinase-3β (GSK3β) which results in suppression of cyclin D1 proteolysis. Aberrant cyclin D1 overexpression in S-phase induced DNA double strand breaks which activated DNA-PK and established the vicious cycle of cycling D1 overexpression. This overexpression of cyclin D1 is responsible for the radioresistance phenotype of long-term FR cells, since this phenotype was completely abrogated by treatment of FR cells by the AKT/PKB signaling inhibitor (API-2), an AKT inhibitor or by a Cdk4 inhibitor. Thus, targeting the AKT/GSK3β/cyclin D1/Cdk4 pathway can be an efficient modality to suppress acquired radioresistance of tumor cells. In this article, I overview the newly discovered molecular mechanisms underlying acquired radioresistance of tumor cells induced by FR, and propose a strategy for eradication of tumors using fractionated RT by overcoming tumor radioresistance. (author)

  4. The overexpressed human 46-kDa mannose 6-phosphate receptor mediates endocytosis and sorting of β-glucuronidase

    International Nuclear Information System (INIS)

    Watanabe, H.; Grubb, J.H.; Sly, W.S.

    1990-01-01

    The authors studied the function of the human small (46-kDa) mannose 6-phosphate receptor (SMPR) in transfected mouse L cells that do not express the larger insulin-like growth factor II/mannose 6-phosphate receptor. Cells overexpressing human SMPR were studied for enzyme binding to cell surface receptors, for binding to intracellular receptors in permeabilized cells, and for receptor-mediated endocytosis of recombinant human β-glucuronidase. Specific binding to human SMPR in permeabilized cells showed a pH optimum between pH 6.0 and pH 6.5. Binding was significant in the present of EDTA but was enhanced by added divalent cations. Up to 2.3% of the total functional receptor could be detected on the cell surface by enzyme binding. They present experiments showing that at very high levels of overexpression, and at pH 6.5, human SMPR mediated the endocytosis of β-glucuronidase. At pH 7.5, the rate of endocytosis was only 14% the rate seen at pH 6.5. Cells overexpressing human SMPR also showed reduced secretion of newly synthesized β-glucuronidase when compared to cells transfected with vector only, suggesting that overexpressed human SMPR can participate in sorting of newly synthesized β-glucuronidase and partially correct the sorting defect in mouse L cells that do not express the insulin-like growth factor II/mannose 6-phosphate receptor

  5. The Influence of Receptor-Mediated Interactions on Reaction-Diffusion Mechanisms of Cellular Self-organisation

    KAUST Repository

    Klika, Václav

    2011-11-10

    Understanding the mechanisms governing and regulating self-organisation in the developing embryo is a key challenge that has puzzled and fascinated scientists for decades. Since its conception in 1952 the Turing model has been a paradigm for pattern formation, motivating numerous theoretical and experimental studies, though its verification at the molecular level in biological systems has remained elusive. In this work, we consider the influence of receptor-mediated dynamics within the framework of Turing models, showing how non-diffusing species impact the conditions for the emergence of self-organisation. We illustrate our results within the framework of hair follicle pre-patterning, showing how receptor interaction structures can be constrained by the requirement for patterning, without the need for detailed knowledge of the network dynamics. Finally, in the light of our results, we discuss the ability of such systems to pattern outside the classical limits of the Turing model, and the inherent dangers involved in model reduction. © 2011 Society for Mathematical Biology.

  6. Rac1 switching at the right time and location is essential for Fcγ receptor-mediated phagosome formation.

    Science.gov (United States)

    Ikeda, Yuka; Kawai, Katsuhisa; Ikawa, Akira; Kawamoto, Kyoko; Egami, Youhei; Araki, Nobukazu

    2017-08-01

    Lamellipodia are sheet-like cell protrusions driven by actin polymerization mainly through Rac1, a GTPase molecular switch. In Fcγ receptor-mediated phagocytosis of IgG-opsonized erythrocytes (IgG-Es), Rac1 activation is required for lamellipodial extension along the surface of IgG-Es. However, the significance of Rac1 deactivation in phagosome formation is poorly understood. Our live-cell imaging and electron microscopy revealed that RAW264 macrophages expressing a constitutively active Rac1 mutant showed defects in phagocytic cup formation, while lamellipodia were formed around IgG-Es. Because activated Rac1 reduced the phosphorylation levels of myosin light chains, failure of the cup formation is probably due to inhibition of actin/myosin II contractility. Reversible photo-manipulation of the Rac1 switch in macrophages fed with IgG-Es could phenocopy two lamellipodial motilities: outward-extension and cup-constriction by Rac1 ON and OFF, respectively. In conjunction with fluorescence resonance energy transfer imaging of Rac1 activity, we provide a novel mechanistic model of phagosome formation spatiotemporally controlled by Rac1 switching within a phagocytic cup. © 2017. Published by The Company of Biologists Ltd.

  7. Ruthenium complexes with phenylterpyridine derivatives target cell membrane and trigger death receptors-mediated apoptosis in cancer cells.

    Science.gov (United States)

    Deng, Zhiqin; Gao, Pan; Yu, Lianling; Ma, Bin; You, Yuanyuan; Chan, Leung; Mei, Chaoming; Chen, Tianfeng

    2017-06-01

    Elucidation of the communication between metal complexes and cell membrane may provide useful information for rational design of metal-based anticancer drugs. Herein we synthesized a novel class of ruthenium (Ru) complexes containing phtpy derivatives (phtpy = phenylterpyridine), analyzed their structure-activity relationship and revealed their action mechanisms. The result showed that, the increase in the planarity of hydrophobic Ru complexes significantly enhanced their lipophilicity and cellular uptake. Meanwhile, the introduction of nitro group effectively improved their anticancer efficacy. Further mechanism studies revealed that, complex (2c), firstly accumulated on cell membrane and interacted with death receptors to activate extrinsic apoptosis signaling pathway. The complex was then transported into cell cytoplasm through transferrin receptor-mediated endocytosis. Most of the intracellular 2c accumulated in cell plasma, decreasing the level of cellular ROS, inducing the activation of caspase-9 and thus intensifying the apoptosis. At the same time, the residual 2c can translocate into cell nucleus to interact with DNA, induce DNA damage, activate p53 pathway and enhance apoptosis. Comparing with cisplatin, 2c possesses prolonged circulation time in blood, comparable antitumor ability and importantly, much lower toxicity in vivo. Taken together, this study uncovers the role of membrane receptors in the anticancer actions of Ru complexes, and provides fundamental information for rational design of membrane receptor targeting anticancer drugs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. ZFAT plays critical roles in peripheral T cell homeostasis and its T cell receptor-mediated response

    International Nuclear Information System (INIS)

    Doi, Keiko; Fujimoto, Takahiro; Okamura, Tadashi; Ogawa, Masahiro; Tanaka, Yoko; Mototani, Yasumasa; Goto, Motohito; Ota, Takeharu; Matsuzaki, Hiroshi; Kuroki, Masahide; Tsunoda, Toshiyuki; Sasazuki, Takehiko; Shirasawa, Senji

    2012-01-01

    Highlights: ► We generated Cd4-Cre-mediated T cell-specific Zfat-deficient mice. ► Zfat-deficiency leads to reduction in the number of the peripheral T cells. ► Impaired T cell receptor-mediated response in Zfat-deficient peripheral T cells. ► Decreased expression of IL-7Rα, IL-2Rα and IL-2 in Zfat-deficient peripheral T cells. ► Zfat plays critical roles in peripheral T cell homeostasis. -- Abstract: ZFAT, originally identified as a candidate susceptibility gene for autoimmune thyroid disease, has been reported to be involved in apoptosis, development and primitive hematopoiesis. Zfat is highly expressed in T- and B-cells in the lymphoid tissues, however, its physiological function in the immune system remains totally unknown. Here, we generated the T cell-specific Zfat-deficient mice and demonstrated that Zfat-deficiency leads to a remarkable reduction in the number of the peripheral T cells. Intriguingly, a reduced expression of IL-7Rα and the impaired responsiveness to IL-7 for the survival were observed in the Zfat-deficient T cells. Furthermore, a severe defect in proliferation and increased apoptosis in the Zfat-deficient T cells following T cell receptor (TCR) stimulation was observed with a reduced IL-2Rα expression as well as a reduced IL-2 production. Thus, our findings reveal that Zfat is a critical regulator in peripheral T cell homeostasis and its TCR-mediated response.

  9. Cryptococcus neoformans Is Internalized by Receptor-Mediated or ‘Triggered’ Phagocytosis, Dependent on Actin Recruitment

    Science.gov (United States)

    Guerra, Caroline Rezende; Seabra, Sergio Henrique; de Souza, Wanderley; Rozental, Sonia

    2014-01-01

    Cryptococcosis by the encapsulated yeast Cryptococcus neoformans affects mostly immunocompromised individuals and is a frequent neurological complication in AIDS patients. Recent studies support the idea that intracellular survival of Cryptococcus yeast cells is important for the pathogenesis of cryptococcosis. However, the initial steps of Cryptococcus internalization by host cells remain poorly understood. Here, we investigate the mechanism of Cryptococcus neoformans phagocytosis by peritoneal macrophages using confocal and electron microscopy techniques, as well as flow cytometry quantification, evaluating the importance of fungal capsule production and of host cell cytoskeletal elements for fungal phagocytosis. Electron microscopy analyses revealed that capsular and acapsular strains of C. neoformans are internalized by macrophages via both ‘zipper’ (receptor-mediated) and ‘trigger’ (membrane ruffle-dependent) phagocytosis mechanisms. Actin filaments surrounded phagosomes of capsular and acapsular yeasts, and the actin depolymerizing drugs cytochalasin D and latrunculin B inhibited yeast internalization and actin recruitment to the phagosome area. In contrast, nocodazole and paclitaxel, inhibitors of microtubule dynamics decreased internalization but did not prevent actin recruitment to the site of phagocytosis. Our results show that different uptake mechanisms, dependent on both actin and tubulin dynamics occur during yeast internalization by macrophages, and that capsule production does not affect the mode of Cryptococcus uptake by host cells. PMID:24586631

  10. Design and development of hyaluronan-functionalized polybenzofulvene nanoparticles as CD44 receptor mediated drug delivery system

    Science.gov (United States)

    Licciardi, Mariano; Scialabba, Cinzia; Giammona, Gaetano; Paolino, Marco; Razzano, Vincenzo; Grisci, Giorgio; Giuliani, Germano; Makovec, Francesco; Cappelli, Andrea

    2017-06-01

    A tri-component polymer brush (TCPB ), composed of a polybenzofulvene copolymer bearing low molecular weight hyaluronic acid (HA) on the surface of its cylindrical brush-like backbone and oligo-PEG fractions, was employed in the preparation of 350 nm nanostructured drug delivery systems capable of delivering the anticancer drug doxorubicin. The obtained drug delivery systems were characterized on the basis of drug loading and release, dimensions and zeta potential, morphology and in vitro cell activity, and uptake on three different human cell lines, namely the bronchial epithelial 16HBE, the breast adenocarcinoma MCF-7, and the colon cancer HCT116 cells. Finally, the ability of doxorubicin-loaded TCPB nanoparticles (DOXO-TCPB) to be internalized into cancer cells by CD44 receptor mediated uptake was assessed by means of uptake studies in HCT cells. These data were supported by anti-CD44-FITC staining assay. The proposed TCPB nanostructured drug delivery systems have many potential applications in nanomedicine, including cancer targeted drug delivery.

  11. Impact of flattening-filter-free radiation on the clonogenic survival of astrocytic cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Steenken, Caroline; Fleckenstein, Jens; Kegel, Stefan; Jahnke, Lennart; Simeonova, Anna; Hartmann, Linda; Kuebler, Jens; Veldwijk, Marlon R.; Wenz, Frederik; Herskind, Carsten; Giordano, Frank Anton [Universitaetsmedizin Mannheim (UMM), Medical Faculty Mannheim, Heidelberg University, Department of Radiation Oncology, Mannheim (Germany)

    2015-07-15

    Flattening-filter-free (FFF) beams are increasingly used in radiotherapy as delivery times can be substantially reduced. However, the relative biologic effectiveness (RBE) of FFF may be increased relative to conventional flattened (FLAT) beams due to differences in energy spectra. Therefore, we investigated the effects of FFF and FLAT beams on the clonogenic survival of astrocytoma cells. Three cell lines (U251, U251-MGMT, and U87) were irradiated with 6-MV and 10-MV X-rays from a linear accelerator in FFF- or FLAT-beam modes at dose rates in the range of 0.5-24 Gy/min. The surviving fraction (SF) as function of dose (2-12 Gy) was determined by the colony formation assay and fitted by the linear-quadratic model. For both beams (FFF or FLAT), the cells were pelleted in conical 15-ml centrifuge tubes and irradiated at 2-cm depth in a 1 x 1-cm{sup 2} area on the central axis of a 30 x 30-cm{sup 2} field. Dosimetry was performed with a 0.3-cm{sup 3} rigid ionization chamber. RBE was determined for FFF versus FLAT irradiation. The RBE of FFF at 7.3-11.3 Gy was 1.027 ± 0.013 and 1.063 ± 0.018 relative to FLAT beams for 6- and 10-MV beams, respectively, and was only significantly higher than 1 for 10 MV. Significantly increased survival rates were seen for lower dose rates (0.5 Gy/min FLAT vs. 5 Gy/min FLAT) at higher doses (11.9 Gy), while no differences were seen at dose rates ≥ 1.4 Gy/min (1.4 Gy/min FFF vs. 14 Gy/min FFF and 2.4 Gy/min FFF vs. 24 Gy/min FFF). FFF beams showed only a slightly increased RBE relative to FLAT beams in this experimental set-up, which is unlikely to result in clinically relevant differences in outcome. (orig.) [German] Die Flattening-Filter-freie (FFF) Bestrahlungstechnik findet zunehmend Verwendung, da sich die Applikationsdauer der einzelnen Fraktionen deutlich verkuerzen laesst. Aufgrund der Unterschiede im Spektrum koennte die relative biologische Wirksamkeit (RBW) von FFF jedoch hoeher sein als bei konventioneller Technik (d.h. bei

  12. Stimulation of accumbal GABAA receptors inhibits delta2-, but not delta1-, opioid receptor-mediated dopamine efflux in the nucleus accumbens of freely moving rats.

    Science.gov (United States)

    Aono, Yuri; Kiguchi, Yuri; Watanabe, Yuriko; Waddington, John L; Saigusa, Tadashi

    2017-11-15

    The nucleus accumbens contains delta-opioid receptors that may reduce inhibitory neurotransmission. Reduction in GABA A receptor-mediated inhibition of accumbal dopamine release due to delta-opioid receptor activation should be suppressed by stimulating accumbal GABA A receptors. As delta-opioid receptors are divided into delta2- and delta1-opioid receptors, we analysed the effects of the GABA A receptor agonist muscimol on delta2- and delta1-opioid receptor-mediated accumbal dopamine efflux in freely moving rats using in vivo microdialysis. Drugs were administered intracerebrally through the dialysis probe. Doses of compounds indicate total amount administered (mol) during 25-50min infusions. The delta2-opioid receptor agonist deltorphin II (25.0nmol)- and delta1-opioid receptor agonist DPDPE (5.0nmol)-induced increases in dopamine efflux were inhibited by the delta2-opioid receptor antagonist naltriben (1.5nmol) and the delta1-opioid receptor antagonist BNTX (150.0pmol), respectively. Muscimol (250.0pmol) inhibited deltorphin II (25.0nmol)-induced dopamine efflux. The GABA A receptor antagonist bicuculline (50.0pmol), which failed to affect deltorphin II (25.0nmol)-induced dopamine efflux, counteracted the inhibitory effect of muscimol on deltorphin II-induced dopamine efflux. Neither muscimol (250.0pmol) nor bicuculline (50.0 and 500.0pmol) altered DPDPE (5.0nmol)-induced dopamine efflux. The present results show that reduction in accumbal GABA A receptor-mediated inhibition of dopaminergic activity is necessary to produce delta2-opioid receptor-induced increase in accumbal dopamine efflux. This study indicates that activation of delta2- but not delta1-opioid receptors on the cell bodies and/or terminals of accumbal GABAergic interneurons inhibits GABA release and, accordingly, decreases GABA A receptor-mediated inhibition of dopaminergic terminals, resulting in enhanced accumbal dopamine efflux. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Additive effects of 5-Aza-2'-deoxycytidine and irradiation on clonogenic survival of human medulloblastoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Patties, Ina; Jahns, Jutta; Kortmann, Rolf-Dieter; Glasow, Annegret [Dept. of Radiotherapy and Radiooncology, Universitaetsklinikum Leipzig AoeR (Germany); Hildebrandt, Guido [Dept. of Radiotherapy and Radiooncology, Universitaetsklinikum Leipzig AoeR (Germany); Dept. of Radiotherapy, Univ. of Rostock (Germany)

    2009-05-15

    Background and purpose: in recent years, epigenetic modulators were introduced into tumor therapy. Here, the authors investigated the antitumor effect of 5-aza-2'-deoxycytidine-(5-aza-dC-)induced demethylation combined with irradiation on human medulloblastoma (MB) cells, which form the most common malignant brain tumor in children. Material and methods: three MB cell lines were treated with 5-aza-dC in a low-dose (0.1 {mu}M, 6 days) or high-dose (3/5 {mu}M, 3 days) setting and irradiated with 2, 4, 6, or 8 Gy single dose on an X-ray unit. Methylation status and mRNA expression of three candidate genes were analyzed by methylation-specific PCR (polymerase chain reaction) and quantitative real-time RT-PCR. Cell survival and mortality were determined by trypan blue exclusion test. Proliferation was analyzed by BrdU incorporation assay, and long-term cell survival was assessed by clonogenic assay. Results: 5-aza-dC treatment resulted in partial promoter demethylation and increased expression of hypermethylated candidate genes. A significant decrease of vital cell count, proliferation inhibition and increase of mortality was observed in 5-aza-dC-treated as well as in irradiated MB cells, whereby combination of both treatments led to additive effects. Although high-dose 5-aza-dC treatment was more effective in terms of demethylation, clonogenic assay revealed no differences between high- and low-dose settings indicating no relevance of 5-aza-dC-induced demethylation for decreased cell survival. MB cells pretreated with 5-aza-dC showed significantly lower plating efficiencies than untreated cells at all irradiation doses investigated. Analysis of surviving curves in irradiated MB cells, however, revealed no significant differences of {alpha}-, {beta}-values and 2-Gy surviving fraction with or without 5-aza-dC treatment. Conclusion: 5-aza-dC did not enhance radiation sensitivity of MB cells but significantly reduced the clonogenicity versus irradiation alone, which

  14. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    Energy Technology Data Exchange (ETDEWEB)

    Chow, Paik Wah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Abdul Hamid, Zariyantey, E-mail: zyantey@ukm.edu.my [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Chan, Kok Meng [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Inayat-Hussain, Salmaan Hussain [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Rajab, Nor Fadilah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia)

    2015-04-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e{sup +} cells but reduced the total counts of Sca-1{sup +}, CD11b{sup +}, Gr-1{sup +}, and CD45{sup +} cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and

  15. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    International Nuclear Information System (INIS)

    Chow, Paik Wah; Abdul Hamid, Zariyantey; Chan, Kok Meng; Inayat-Hussain, Salmaan Hussain; Rajab, Nor Fadilah

    2015-01-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e + cells but reduced the total counts of Sca-1 + , CD11b + , Gr-1 + , and CD45 + cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and progenitors. • 1,4-BQ

  16. Enhanced radiation response in radioresistant MCF-7 cells by targeting peroxiredoxin II

    Directory of Open Access Journals (Sweden)

    Diaz AJG

    2013-10-01

    Full Text Available Anthony Joseph Gomez Diaz,1 Daniel Tamae,2 Yun Yen,3 JianJian Li,4 Tieli Wang1 1Department of Chemistry and Biochemistry, California State University at Dominguez Hills, Carson, CA, 2Center of Excellence in Environmental Toxicology, Department of Pharmacology, University of Pennsylvania, Philadelphia, PA, 3Department of Clinical and Molecular Pharmacology, Beckman Research Institute of City of Hope National Medical Center, Duarte, CA, 4Department of Radiation Oncology, University of California Davis, Sacramento, CA, USA Abstract: In our previous study, we identified that a protein target, peroxiredoxin II (PrxII, is overexpressed in radioresistant MCF+FIR3 breast-cancer cells and found that its expression and function is associated with breast-cancer radiation sensitivity or resistance. Small interference RNA (siRNA targeting PrxII gene expression was able to sensitize MCF+FIR3 radioresistant breast-cancer cells to ionizing radiation. The major focus of this work was to investigate how the radiation response of MCF+FIR3 radioresistant cells was affected by the siRNA that inhibits PrxII gene expression. Our results, presented here, show that silencing PrxII gene expression increased cellular toxicity by altering cellular thiol status, inhibiting Ca2+ efflux from the cells, and perturbing the intracellular Ca2+ homeostasis. By combining radiotherapy and siRNA technology, we hope to develop new therapeutic strategies that may have potential to enhance the efficacy of chemotherapeutic agents due to this technology's property of targeting to specific cancer-related genes. Keywords: siRNA, PrxII, radiation resistance, Ca2+, MCF+FIR3

  17. Metabolism of the chemo antibiotic combination sulphamethoxazol and trimethoprim by some radioresistant strains

    International Nuclear Information System (INIS)

    Zahiera, T.S.

    1990-01-01

    The radioresistant mutants B. Firmus, B. Laterosporus, B. Megaterium, M. Roseus and M. Luteus were tested microbiologically for their sensitivity to a drug consisting of the chemo antibiotic combination of sulphamethoxazole (SMZ) and trimethoprim (TMP) in a ratio of 20:1. All the studied mutant strains were found to be insensitive to this combination of antibiotics except the radioresistant mutant strain of M.roseus. The mechanism of the inactivation of both SMZ and TMP by the radioresistant mutants of B. Laterosporus, B. Firmus, and M.Luteus was studied. The drug was incubated with each of these mutants for different periods of time to study its degradation. High performance liquid chromatography and microbiological assay techniques were used to determine the activity and the pathway of the consumption of the drug after its incubation with these three mutants for time. The results indicated that the drug acts in a synergistic effect on all the studied mutants. Only a part of both molecules of SMZ and TMP was utilized by the studied mutant strains. The ratio of the left over of SMZ/TMP were found to be 6,6 and 8 after seven days incubation periods and 12, 7.5 and 19 after two weeks incubation periods with B. Laterosporus, B. Firmus, and M. Luteus mutants respectively. The microbial activity of the extracted drug for bacilli strains isolated from clean environment had not changed significantly due to incubation with the mutants, but it was decreased for M. Luteus strain. The drug inactivation by the studied mutant strains seemed to be due to decreased affinities to SMZ and TMP, and to formation of an altered 7,8 dihydropteroate synthetase and dihydrofolate reductase

  18. The heat shock proteins at the increasing of the radioresistance of silkworm embryo Bombyx Moril

    International Nuclear Information System (INIS)

    Agaev, F.A.; Garibov, A.A.; Aliev, D.I.; Alieva, I.N.

    2002-01-01

    The aim of this study is revealing the role of Heat Shock Proteins (HSP) at the radio-modification effect of Heat Shock (HS) on the silkworm embryo. Our preliminary study was indicated that 3-daily silkworm embryo is more than 10-12 times sensitive to gamma-radiation in comparative to 7-daily embryo. Investigation of the HS effect on the radiosensitivity of the embryo indicates that thermal treatment (40 deg. C) of 3 daily embryo during 60 min before irradiation leads the increasing of its radioresistance. At the same time, the HS treatment immediately after irradiation capable increases strike affect of irradiation. LD 50 for heat treated embryo before and after irradiation consists of 46.7 and 19.0 Gy, correspondingly. For embryo irradiated without heat treatment LD 50 is 29.5 Gy. Identical effects are observed for 7-daily embryo. Increasing of the radioresistance by HS before irradiation, obviously, may be explained by the embryo cell reply to the stress factor and capable initiated by different biochemical shifts, for example, by induction of HPS synthesis. According to the results of carried out experiments HS treatment leads to the induction or increasing of the HSP synthesis into this embryo. Protein with molecular mass 70 kDa (HSP-70 kDa) has been synthesised de novo. The synthesis of the other two proteins (HSP-83 kDa and HSP-68 kDa) significantly increases at the high temperature. It is noted, that HSP-70 kDa consists of 55-60 % of whole included radioactive mark. Identical induction was observed in the experiments at the combined effect both HS and gamma-radiation on the embryo. At the post-radiation heat treatment the induction of HSP synthesis is observed, too. It was concluded that damages induced by irradiation can not prevent HSP induction into embryo. The result of comparative analysis was shown that in 3-daily embryo in spite of 7-daily embryo the synthesis of HSP is more intensive and correlated with the radio- modification effect of HS. The

  19. Rate of oxygen consumption of hamster melanoma cells as a factor influencing their radioresistance

    International Nuclear Information System (INIS)

    Pajak, S.; Subczynski, W.; Panz, T.; Lukiewicz, S.

    1980-01-01

    It has been reported in recent years that the level of radiosensitivity of neoplasmic cells in vivo and of sphaeroids in vitro can be modified by controlling their rate of oxygen consumption. Thus, an attempt was made to compare this rate in the case of the melanotic and amelanotic lines of Bomirski hamster melanoma in vitro, as it is known that these two lines distinctly differ in their reactivity to ionizing radiations. The measurements carried out by the use of a new ESR method revealed that pigmented and pigmentless cells consume oxygen at significantly different rates. This means that oxygen utilization may contribute to the overall level of radioresistance of melanoma cells. (author)

  20. Alterations in radioresistance of eucaryotic cells after the transfer of genomic wildtype DNA and metallothionein genes

    International Nuclear Information System (INIS)

    Lohrer, H.

    1987-01-01

    The presented paper describes experiments concerning the alteration of radiosensitivity of eucaryotic cells after gene transfer. Ionizing radiation (γ- or X-ray) induces DNA single- or double strand breaks, which are religated by an unknown repair system. Repair deficient cells are highly sensitive to ionizing radiation. In the experiments described, cells from a patient with the heritable disease Ataxia telangiectasia were used as well as two X-ray sensitive CHO mutant cell lines. After gene transfer of an intact human DNA repair gene or a metallothionein gene the cells should regain radioresistance. (orig.) [de

  1. Clonogenic cell line survival of a human liver cancer cell line SMMC-7721 after carbon ion irradiation with different LET

    International Nuclear Information System (INIS)

    Lei Suwen; Su Xu; Wang Jifang; Li Wenjian

    2003-01-01

    Objective: To investigate the survival fraction of a human liver cancer cell line SMMC-7721 following irradiation with carbon ions with different LET. Methods: cells of the human liver cancer cell line SMMC-7721 were irradiated with carbon ions (LET=30 and 70 keV/μm). The survival fraction was determined with clonogenic assay after 9 days incubation in a 5% CO 2 incubator at 37 degree C. Results: When the survival fractions of 70 keV/μm were D s = 0.1 and D s=0.01 absorption dose were 2.94 and 5.88 Gy respectively, and those of 30 keV/μm were 4.00 and 8.00 Gy respectively. Conclusion: For the SMMC-7721 cell line, 70 keV/μm is more effective for cell killing than 30 keV/μm

  2. Acute response of mouse kidney clonogens to fractionated irradiation in situ and then assayed in primary culture

    International Nuclear Information System (INIS)

    Yeemin Jen; Hendry, J.H.

    1991-01-01

    The radiosensitivity of mouse kidney cells after in situ single-dose, 2, 8, and 16 fraction X-irradiations was measured in primary culture using a clonogenic assay. The assay was made 12 h after single doses or 12 h after the last dose of the multifraction regimens. When analysed using the linear-quadratic model, as predicted the individual α components for all the different fractionation schedules were not significantly different, and the changes in the β values were consistent with those expected on the basis of the reciprocal fraction numbers. When all four data sets were integrated to derive a common α/β ratio, the result was 4.4±1.3 (1SE) Gy, or 2.8±0.9 Gy (a better fit) if the single-dose data set was excluded. These values fall into the range reported for kidney using assays of tissue function at long times after irradiation. (author)

  3. Test of equal effect per fraction and estimation of initial clonogen number in microcolony assays of survival after fractionated irradiation

    International Nuclear Information System (INIS)

    Thames, H.D.; Withers, H.R.

    1980-01-01

    In the use of multifraction microcolony assays to infer the low-dose response of in situ renewal systems such as intestinal crypts, the assumption of equal effect per dose fraction is required. Moreover, the construction of a cell-survival curve requires knowledge of the initial count of cells capable of repopulating each renewal structure. We describe a method of designing fractionation protocols which provides a regression estimate of the initial number of clonogens per renewal structure and a test of the hypothesis of equal effect per fraction. The essential factor in the experimental design is the use of common dose fractions (use of the same dose per fraction in series with different numbers of fractions). Applications of the method to data for which the assumption of equal effect per fraction holds (four-hour fractionation interval murine testis study) and does not hold (one-hour fractionation interval murine jejunal crypt study) are presented. (author)

  4. Critical role of zinc finger protein 521 in the control of growth, clonogenicity and tumorigenic potential of medulloblastoma cells.

    Science.gov (United States)

    Spina, Raffaella; Filocamo, Gessica; Iaccino, Enrico; Scicchitano, Stefania; Lupia, Michela; Chiarella, Emanuela; Mega, Tiziana; Bernaudo, Francesca; Pelaggi, Daniela; Mesuraca, Maria; Pazzaglia, Simonetta; Semenkow, Samantha; Bar, Eli E; Kool, Marcel; Pfister, Stefan; Bond, Heather M; Eberhart, Charles G; Steinkühler, Christian; Morrone, Giovanni

    2013-08-01

    The stem cell-associated transcription co-factor ZNF521 has been implicated in the control of hematopoietic, osteo-adipogenic and neural progenitor cells. ZNF521 is highly expressed in cerebellum and in particular in the neonatal external granule layer that contains candidate medulloblastoma cells-of-origin, and in the majority of human medulloblastomas. Here we have explored its involvement in the control of human and murine medulloblastoma cells. The effect of ZNF521 on growth and tumorigenic potential of human medulloblastoma cell lines as well as primary Ptc1-/+ mouse medulloblastoma cells was investigated in a variety of in vitro and in vivo assays, by modulating its expression using lentiviral vectors carrying the ZNF521 cDNA, or shRNAs that silence its expression. Enforced overexpression of ZNF521 in DAOY medulloblastoma cells significantly increased their proliferation, growth as spheroids and ability to generate clones in single-cell cultures and semisolid media, and enhanced their migratory ability in wound-healing assays. Importantly, ZNF521-expressing cells displayed a greatly enhanced tumorigenic potential in nude mice. All these activities required the ZNF521 N-terminal motif that recruits the nucleosome remodeling and histone deacetylase complex, which might therefore represent an appealing therapeutic target. Conversely, silencing of ZNF521 in human UW228 medulloblastoma cells that display high baseline expression decreased their proliferation, clonogenicity, sphere formation and wound-healing ability. Similarly, Zfp521 silencing in mouse Ptc1-/+ medulloblastoma cells drastically reduced their growth and tumorigenic potential. Our data strongly support the notion that ZNF521, through the recruitment of the NuRD complex, contributes to the clonogenic growth, migration and tumorigenicity of medulloblastoma cells.

  5. Ah receptor mediated suppression of the antibody response in mice is primarily dependent on the Ah phenotype of lymphoid tissue

    International Nuclear Information System (INIS)

    Silkworth, J.B.; Antrim, L.A.; Sack, G.

    1986-01-01

    Halogenated aromatic hydrocarbons act through the aromatic hydrocarbon (Ah) receptor in mice to produce a series of toxic effects of the immune system. The receptor protein is a product of the Ah gene locus. Ah responsive (Ahb/Ahb) mice express a high affinity receptor in both lymphoid and nonlymphoid tissues whereas nonresponsive Ahd/Ahd mice express a poor affinity receptor. To determine the role of the Ah receptor of lymphoid tissue relative to that of nonlymphoid tissue in the induction of immune impairment, bone marrow was used to reconstitute lethally irradiated mice of the same or opposite Ah phenotype. All mice were given 3,3',4,4'-tetrachlorobiphenyl (35 and 350 mumol/kg) ip 2 days before immunization with sheep erythrocytes (SRBC). The immune response to this T dependent antigen and organ weights were determined 5 or 7 days later in normal or chimeric mice, respectively. Monoclonal Lyt 1.1 and Lyt 1.2 antibodies were used to establish the origin of the cells which repopulated the chimeric thymuses. The immune responses of both BALB/cBy (Ahb/Ahb) and the BALB/cBy X DBA/2 hybrid, CByD2F1 (Ahb/Ahd), were significantly suppressed but DBA/2 mice were unaffected. The immune responses of chimeric BALB/cBy----BALB/cBy and BALB/cBy----DBA/2 (donor----recipient) mice were also significantly suppressed and thymic atrophy was observed in both cases. The serum anti-SRBC antibody titers of DBA/2----BALB/cBy chimeras were also significantly decreased although not to the same extent as in BALB/cBy----DBA/2 mice. Chimeric DBA/2----DBA/2 mice were not affected. These results indicate that the sensitivity to Ah receptor mediated suppression of the antibody response is primarily determined by the Ah phenotype of the lymphoid tissue

  6. Biological functionalization of drug delivery carriers to bypass size restrictions of receptor-mediated endocytosis independently from receptor targeting.

    Science.gov (United States)

    Ansar, Maria; Serrano, Daniel; Papademetriou, Iason; Bhowmick, Tridib Kumar; Muro, Silvia

    2013-12-23

    Targeting of drug carriers to cell-surface receptors involved in endocytosis is commonly used for intracellular drug delivery. However, most endocytic receptors mediate uptake via clathrin or caveolar pathways associated with ≤200-nm vesicles, restricting carrier design. We recently showed that endocytosis mediated by intercellular adhesion molecule 1 (ICAM-1), which differs from clathrin- and caveolae-mediated pathways, allows uptake of nano- and microcarriers in cell culture and in vivo due to recruitment of cellular sphingomyelinases to the plasmalemma. This leads to ceramide generation at carrier binding sites and formation of actin stress-fibers, enabling engulfment and uptake of a wide size-range of carriers. Here we adapted this paradigm to enhance uptake of drug carriers targeted to receptors associated with size-restricted pathways. We coated sphingomyelinase onto model (polystyrene) submicro- and microcarriers targeted to clathrin-associated mannose-6-phosphate receptor. In endothelial cells, this provided ceramide enrichment at the cell surface and actin stress-fiber formation, modifying the uptake pathway and enhancing carrier endocytosis without affecting targeting, endosomal transport, cell-associated degradation, or cell viability. This improvement depended on the carrier size and enzyme dose, and similar results were observed for other receptors (transferrin receptor) and cell types (epithelial cells). This phenomenon also enhanced tissue accumulation of carriers after intravenous injection in mice. Hence, it is possible to maintain targeting toward a selected receptor while bypassing natural size restrictions of its associated endocytic route by functionalization of drug carriers with biological elements mimicking the ICAM-1 pathway. This strategy holds considerable promise to enhance flexibility of design of targeted drug delivery systems.

  7. The brain cytoplasmic RNA BC1 regulates dopamine D2 receptor-mediated transmission in the striatum.

    Science.gov (United States)

    Centonze, Diego; Rossi, Silvia; Napoli, Ilaria; Mercaldo, Valentina; Lacoux, Caroline; Ferrari, Francesca; Ciotti, Maria Teresa; De Chiara, Valentina; Prosperetti, Chiara; Maccarrone, Mauro; Fezza, Filomena; Calabresi, Paolo; Bernardi, Giorgio; Bagni, Claudia

    2007-08-15

    Dopamine D(2) receptor (D(2)DR)-mediated transmission in the striatum is remarkably flexible, and changes in its efficacy have been heavily implicated in a variety of physiological and pathological conditions. Although receptor-associated proteins are clearly involved in specific forms of synaptic plasticity, the molecular mechanisms regulating the sensitivity of D(2) receptors in this brain area are essentially obscure. We have studied the physiological responses of the D(2)DR stimulations in mice lacking the brain cytoplasmic RNA BC1, a small noncoding dendritically localized RNA that is supposed to play a role in mRNA translation. We show that the efficiency of D(2)-mediated transmission regulating striatal GABA synapses is under the control of BC1 RNA, through a negative influence on D(2) receptor protein level affecting the functional pool of receptors. Ablation of the BC1 gene did not result in widespread dysregulation of synaptic transmission, because the sensitivity of cannabinoid CB(1) receptors was intact in the striatum of BC1 knock-out (KO) mice despite D(2) and CB(1) receptors mediated similar electrophysiological actions. Interestingly, the fragile X mental retardation protein FMRP, one of the multiple BC1 partners, is not involved in the BC1 effects on the D(2)-mediated transmission. Because D(2)DR mRNA is apparently equally translated in the BC1-KO and wild-type mice, whereas the protein level is higher in BC1-KO mice, we suggest that BC1 RNA controls D(2)DR indirectly, probably regulating translation of molecules involved in D(2)DR turnover and/or stability.

  8. Testin, a novel binding partner of the calcium-sensing receptor, enhances receptor-mediated Rho-kinase signalling

    International Nuclear Information System (INIS)

    Magno, Aaron L.; Ingley, Evan; Brown, Suzanne J.; Conigrave, Arthur D.; Ratajczak, Thomas; Ward, Bryan K.

    2011-01-01

    Highlights: → A yeast two-hybrid screen revealed testin bound to the calcium-sensing receptor. → The second zinc finger of LIM domain 1 of testin is critical for interaction. → Testin bound to a region of the receptor tail important for cell signalling. → Testin and receptor interaction was confirmed in mammalian (HEK293) cells. → Overexpression of testin enhanced receptor-mediated Rho signalling in HEK293 cells. -- Abstract: The calcium-sensing receptor (CaR) plays an integral role in calcium homeostasis and the regulation of other cellular functions including cell proliferation and cytoskeletal organisation. The multifunctional nature of the CaR is manifested through ligand-dependent stimulation of different signalling pathways that are also regulated by partner binding proteins. Following a yeast two-hybrid library screen using the intracellular tail of the CaR as bait, we identified several novel binding partners including the focal adhesion protein, testin. Testin has not previously been shown to interact with cell surface receptors. The sites of interaction between the CaR and testin were mapped to the membrane proximal region of the receptor tail and the second zinc-finger of LIM domain 1 of testin, the integrity of which was found to be critical for the CaR-testin interaction. The CaR-testin association was confirmed in HEK293 cells by coimmunoprecipitation and confocal microscopy studies. Ectopic expression of testin in HEK293 cells stably expressing the CaR enhanced CaR-stimulated Rho activity but had no effect on CaR-stimulated ERK signalling. These results suggest an interplay between the CaR and testin in the regulation of CaR-mediated Rho signalling with possible effects on the cytoskeleton.

  9. ZFAT plays critical roles in peripheral T cell homeostasis and its T cell receptor-mediated response

    Energy Technology Data Exchange (ETDEWEB)

    Doi, Keiko [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute of Life Sciences for the Next Generation of Women Scientists, Fukuoka University, Fukuoka (Japan); Fujimoto, Takahiro [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Okamura, Tadashi [Division of Animal Models, Department of Infectious Diseases, Research Institute, National Center for Global Health and Medicine, Tokyo (Japan); Ogawa, Masahiro [Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Tanaka, Yoko [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Mototani, Yasumasa; Goto, Motohito [Division of Animal Models, Department of Infectious Diseases, Research Institute, National Center for Global Health and Medicine, Tokyo (Japan); Ota, Takeharu; Matsuzaki, Hiroshi [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Kuroki, Masahide [Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Tsunoda, Toshiyuki [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan); Sasazuki, Takehiko [Institute for Advanced Study, Kyushu University, Fukuoka (Japan); Shirasawa, Senji, E-mail: sshirasa@fukuoka-u.ac.jp [Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka (Japan); Central Research Institute for Advanced Molecular Medicine, Fukuoka University, Fukuoka (Japan)

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer We generated Cd4-Cre-mediated T cell-specific Zfat-deficient mice. Black-Right-Pointing-Pointer Zfat-deficiency leads to reduction in the number of the peripheral T cells. Black-Right-Pointing-Pointer Impaired T cell receptor-mediated response in Zfat-deficient peripheral T cells. Black-Right-Pointing-Pointer Decreased expression of IL-7R{alpha}, IL-2R{alpha} and IL-2 in Zfat-deficient peripheral T cells. Black-Right-Pointing-Pointer Zfat plays critical roles in peripheral T cell homeostasis. -- Abstract: ZFAT, originally identified as a candidate susceptibility gene for autoimmune thyroid disease, has been reported to be involved in apoptosis, development and primitive hematopoiesis. Zfat is highly expressed in T- and B-cells in the lymphoid tissues, however, its physiological function in the immune system remains totally unknown. Here, we generated the T cell-specific Zfat-deficient mice and demonstrated that Zfat-deficiency leads to a remarkable reduction in the number of the peripheral T cells. Intriguingly, a reduced expression of IL-7R{alpha} and the impaired responsiveness to IL-7 for the survival were observed in the Zfat-deficient T cells. Furthermore, a severe defect in proliferation and increased apoptosis in the Zfat-deficient T cells following T cell receptor (TCR) stimulation was observed with a reduced IL-2R{alpha} expression as well as a reduced IL-2 production. Thus, our findings reveal that Zfat is a critical regulator in peripheral T cell homeostasis and its TCR-mediated response.

  10. Role of receptor-mediated endocytosis in the antiangiogenic effects of human T lymphoblastic cell-derived microparticles.

    Science.gov (United States)

    Yang, Chun; Xiong, Wei; Qiu, Qian; Shao, Zhuo; Shao, Zuo; Hamel, David; Tahiri, Houda; Leclair, Grégoire; Lachapelle, Pierre; Chemtob, Sylvain; Hardy, Pierre

    2012-04-15

    Microparticles possess therapeutic potential regarding angiogenesis. We have demonstrated the contribution of apoptotic human CEM T lymphocyte-derived microparticles (LMPs) as inhibitors of angiogenic responses in animal models of inflammation and tumor growth. In the present study, we characterized the antivascular endothelial growth factor (VEGF) effects of LMPs on pathological angiogenesis in an animal model of oxygen-induced retinopathy and explored the role of receptor-mediated endocytosis in the effects of LMPs on human retinal endothelial cells (HRECs). LMPs dramatically inhibited cell growth of HRECs, suppressed VEGF-induced cell migration in vitro experiments, and attenuated VEGF-induced retinal vascular leakage in vivo. Intravitreal injections of fluorescently labeled LMPs revealed accumulation of LMPs in retinal tissue, with more than 60% reductions of the vascular density in retinas of rats with oxygen-induced neovascularization. LMP uptake experiments demonstrated that the interaction between LMPs and HRECs is dependent on temperature. In addition, endocytosis is partially dependent on extracellular calcium. RNAi-mediated knockdown of low-density lipoprotein receptor (LDLR) reduced the uptake of LMPs and attenuated the inhibitory effects of LMPs on VEGF-A protein expression and HRECs cell growth. Intravitreal injection of lentivirus-mediated RNA interference reduced LDLR protein expression in retina by 53% and significantly blocked the antiangiogenic effects of LMPs on pathological vascularization. In summary, the potent antiangiogenic LMPs lead to a significant reduction of pathological retinal angiogenesis through modulation of VEGF signaling, whereas LDLR-mediated endocytosis plays a partial, but pivotal, role in the uptake of LMPs in HRECs.

  11. The increase in radioresistance of Chinese hamster cells cultured as spheroids is correlated to changes in nuclear morphology

    International Nuclear Information System (INIS)

    Gordon, D.J.; Milner, A.E.; Beaney, R.P.; Grdina, D.J.; Vaughan, A.T.

    1990-01-01

    Chinese hamster V79 cells grown as spheroids in roller culture are more radioresistant than those grown as monolayers. The supercoiled structure of chromatin, as salt-extracted nucleoids, has been examined using flow cytometry. Irradiated viable cells from spheroid culture contain restraints to supercoil relaxation that are absent in monolayer cells. Further analysis of the chromatin organization from each growth form shows that the radioresistant spheroid cells contain a DNA-protein matrix that is more resistant to detergent-induced degradation. The increase in structural integrity may be due to the retention of a 55-60 kDa protein that is apparent in the nucleoids of spheroid, but not monolayer cells. The increase in structural integrity of the spheroid cells may explain their greater radioresistance by providing a more stable platform for high-fidelity DNA damage repair

  12. Radioresistant CD4+ T cells in normal, unprimed mice, with verification of the Bergonie-Tribondeau law

    International Nuclear Information System (INIS)

    Makidono, Reiko; Ito, Akira.

    1997-01-01

    This is the first report on radioresistant CD4+ T cells found in normal, unprimed mice. After sublethal whole body irradiation, regular CD4+ as well as primitive NK1.1+ CD4+ T cells were enriched in the spleen. Since it has been well established that virgin T and B cells are highly radiosensitive, these cells were once assumed to be a unique lymphocyte population for which radiosensitivity does not follow the general law of radiation sensitivity for mammalian cells (Bergonie-Tribondeau law). These cells exhibited higher proliferative response to accessory cells than the non-irradiated control cells in the syngeneic mixed leukocyte reaction (SMLR). This indicated that virgin CD4+ T cells sensitized to, and readily respond to self-MHC class II molecules are radioresistant, and that their radioresistance, as activated cells, is consistent with the Bergonie-Tribondeau law. (author)

  13. Sister chromatid exchanges in X-ray irradiated blood lymphocytes from patients with hereditary diseases with radioresistant DNA synthesis

    International Nuclear Information System (INIS)

    Pleskach, N.M.; Andriadze, M.I.; Mikhel'son, V.M.; Zhestyanikov, V.D.

    1988-01-01

    X-ray irradiation induced sister chromatid exchanges (SCE) in blood lymphocytes from patient with Down's syndrome and adult progeria (in both the cases radioresistant DNA synthesis takes place). In normal lymphocytes (in which ionizing radiation inhibits the replicative synthesis of DNA) the rate of SCE rises with the rise of radiation dose. Thus, the rate of SCE in X-ray irradiated lymphocytes is in reverse dependence with radioresistance of replicative synthesis of DNA. The data obtained are explained in accordance with the replicative hypothesis of the SCE nature (Painter, 1980a): in cells of patients with Down's syndrome, xeroderma pigmentosum from 2 and progeria of adults the time of existence of partly replicated clusters of replicons is decreased due to radioresistant replicative synthesis of DNA, but the presence of partly replicated clusters of replicons in necessary for SCE formation. Therefore the rate of SCF in X-irradiated cells of these patients decreases

  14. Breast cancer stem cell-like cells generated during TGFβ-induced EMT are radioresistant.

    Science.gov (United States)

    Konge, Julie; Leteurtre, François; Goislard, Maud; Biard, Denis; Morel-Altmeyer, Sandrine; Vaurijoux, Aurélie; Gruel, Gaetan; Chevillard, Sylvie; Lebeau, Jérôme

    2018-05-04

    Failure of conventional antitumor therapy is commonly associated with cancer stem cells (CSCs), which are often defined as inherently resistant to radiation and chemotherapeutic agents. However, controversy about the mechanisms involved in the radiation response remains and the inherent intrinsic radioresistance of CSCs has also been questioned. These discrepancies observed in the literature are strongly associated with the cell models used. In order to clarify these contradictory observations, we studied the radiosensitivity of breast CSCs using purified CD24 -/low /CD44 + CSCs and their corresponding CD24 + /CD44 low non-stem cells. These cells were generated after induction of the epithelial-mesenchymal transition (EMT) by transforming growth factor β (TGFβ) in immortalized human mammary epithelial cells (HMLE). Consequently, these 2 cellular subpopulations have an identical genetic background, their differences being related exclusively to TGFβ-induced cell reprogramming. We showed that mesenchymal CD24 -/low /CD44 + CSCs are more resistant to radiation compared with CD24 + /CD44 low parental cells. Cell cycle distribution and free radical scavengers, but not DNA repair efficiency, appeared to be intrinsic determinants of cellular radiosensitivity. Finally, for the first time, we showed that reduced radiation-induced activation of the death receptor pathways (FasL, TRAIL and TNF-α) at the transcriptional level was a key causal event in the radioresistance of CD24 -/low / CD44+ cells acquired during EMT.

  15. The effect of caffeine on x-ray repair of radioresistant HeLa cells

    International Nuclear Information System (INIS)

    Kubo, Kihei; Koiwai, Soichiro; Morita, Kazuo

    1985-01-01

    The contribution of caffeine-modifiable repair process to the radiosensitivity of a radioresistant HeLa strain (RC-355) has been investigated in comparison with control HeLa strain (CC-24). Both the final slope and the shoulder of X-ray survival curve for log-phase cells were affected by caffeine posttreatment. When the treatment with 10 mM caffeine delayed, an increase in survival was observed with increasing interval between irradiation and the treatment. During first several hours of the repair interval, the steepness of the final slope of survival curve decreased rapidly, and rate of the decrease was found to be higher in RC-355 than in CC-24 cells. Longer time (24 hours or more) before the initiation of caffeine treatment was required for the complete recovery of the shoulder. When the cells were incubated in plateau-phase after irradiation, an appreciable increase in survival was observed in comparison with when plated immediately following X-ray. The increase was found to be greater for RC-355 than for CC-24. The results suggest that the radioresistant RC-355 cells repaired more X-ray-induced PLD than CC-24 cells did. (author)

  16. Isolation and characterization of radioresistant mutants in Bacillus subtilis and Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Kalinin, V.L.; Petrov, V.N.; Petrova, T.M.

    1981-01-01

    Vegetative cells of Bac. thuringiensis var. galleriae (the wild-type strain 351) are much more sensitive to lethal effects of UV light and 60 Co-γ-rays than those of Bac. subtilis (the wild-type strain 168). This difference is less pronounced for spores of these strains. By means of repeated γ-irradiation-regrowth cycles radioresistant mutants Bac. thuringiensis Gamsup(r) 14 and Bac. subtilis Gamsup(r) 9 were selected. The vegetative cells of these mutants are correspondingly 19 times and 3.9 times more resistant to lethal effects of γ-radiation than the cells of the parental strains. The resistance of the Gamsup(r) mutant cells to lethal effects of UV light and H 2 O 2 is also increased. The spores of the Gamsup(r) 14 mutant are 1.5-1.7 times more resistant to γ-radiation and UV light than the wild-type spores. The radioresistant mutants and the parental strains do not vary in their capacity for host-cell reactivation of UV- or γ-irradiated phages Tg13 and 105

  17. Potentially lethal damage repair in cell lines of radioresistant human tumours and normal skin fibroblasts

    International Nuclear Information System (INIS)

    Marchese, M.J.; Minarik, L.; Hall, E.J.; Zaider, M.

    1985-01-01

    Radiation cell survival data were obtained in vitro for three cell lines isolated from human tumours traditionally considered to be radioresistant-two melanomas and one osteosarcoma-as well as from a diploid skin fibroblast cell line. One melanoma cell line was much more radioresistant than the other, while the osteosarcoma and fibroblast cell lines were more radiosensitive than either. For cells growing exponentially, little potentially lethal damage repair (PLDR) could be demonstrated by comparing survival data for cells in which subculture was delayed by 6 h with those sub-cultured immediately after treatment. For the malignant cells in plateau phase, which in these cells might be better termed 'slowed growth phase', since an appreciable fraction of the cells are still cycling, a small amount of PLDR was observed, but not as much as reported by other investigators in the literature. The normal fibroblasts, which achieved a truer plateau phase in terms of noncycling cells, showed a significantly larger amount of PLDR than the tumour cells. (author)

  18. 9-β-arabinofuranosyladenine preferentially sensitizes radioresistant squamous cell carcinoma cell lines to x-rays

    International Nuclear Information System (INIS)

    Heaton, D.

    1992-06-01

    The effect of 9-β-arabinofuranosyladenine (ara-A) on sensitivity to the deleterious effects of x-rays was studied in six squamous cell carcinoma cell lines. Three lines were relatively radioresistant, having D 0 values of 2.31 to 2.89 Gy, and the other three lines were relatively radiosensitive, having D 0 values of between 1.07 and 1.45 Gy. Ara-A (50 or 500 μM) was added to cultures 30 min prior to irradiation and removed 30 min after irradiation, and sensitivity was measured in terms of cell survival. The radiosensitizing effect of ara-A was very dependent on the inherent radiosensitivity of the tumor cell line. Fifty micromolar concentrations of ara-A sensitized only the two most radioresistant lines, SCC-12B.2 and JSQ-3. Five hundred micromolar concentrations of ara-A sensitized the more sensitive cell lines, SQ-20B and SQ-9G, but failed to have any effect on the radiation response of the two most sensitive cell lines, SQ-38 and SCC-61. Concentrations of ara-A as low as 10 μM were equally efficient in inhibiting DNA synthesis in all six cell lines. These results suggest that the target for the radiosensitizing effect of ara-A is probably related to the factor controlling the inherent radiosensitivity of human tumor cells. Therefore, ara-A might be useful in overcoming radiation resistance in vivo

  19. 9-{beta}-arabinofuranosyladenine preferentially sensitizes radioresistant squamous cell carcinoma cell lines to x-rays

    Energy Technology Data Exchange (ETDEWEB)

    Heaton, D. [Rush Univ. Medical Center, Chicago, IL (United States). Therapeutic Radiology; Mustafi, R. [Chicago Univ., IL (United States). Dept. of Radiation and Cellular Oncology; Schwartz, J.L. [Chicago Univ., IL (United States). Dept. of Radiation and Cellular Oncology]|[Argonne National Lab., IL (United States)

    1992-06-01

    The effect of 9-{beta}-arabinofuranosyladenine (ara-A) on sensitivity to the deleterious effects of x-rays was studied in six squamous cell carcinoma cell lines. Three lines were relatively radioresistant, having D{sub 0} values of 2.31 to 2.89 Gy, and the other three lines were relatively radiosensitive, having D{sub 0} values of between 1.07 and 1.45 Gy. Ara-A (50 or 500 {mu}M) was added to cultures 30 min prior to irradiation and removed 30 min after irradiation, and sensitivity was measured in terms of cell survival. The radiosensitizing effect of ara-A was very dependent on the inherent radiosensitivity of the tumor cell line. Fifty micromolar concentrations of ara-A sensitized only the two most radioresistant lines, SCC-12B.2 and JSQ-3. Five hundred micromolar concentrations of ara-A sensitized the more sensitive cell lines, SQ-20B and SQ-9G, but failed to have any effect on the radiation response of the two most sensitive cell lines, SQ-38 and SCC-61. Concentrations of ara-A as low as 10 {mu}M were equally efficient in inhibiting DNA synthesis in all six cell lines. These results suggest that the target for the radiosensitizing effect of ara-A is probably related to the factor controlling the inherent radiosensitivity of human tumor cells. Therefore, ara-A might be useful in overcoming radiation resistance in vivo.

  20. Radioresistant DNA synthesis in cells of patients showing increased chromosomal sensitivity to ionizing radiation

    International Nuclear Information System (INIS)

    Barenfeld, L.S.; Pleskach, N.M.; Bildin, V.N.; Prokofjeva, V.V.; Mikhelson, V.M.

    1986-01-01

    The rate of DNA synthesis after γ-irradiation was studied either by analysis of the steady-state distribution of daughter [ 3 H]DNA in alkaline sucrose gradients or by direct assay of the amount of [ 3 H]thymidine incorporated into DNA of fibroblasts derived from a normal donor (LCH882) and from Down's syndrome (LCH944), Werner's syndrome (WS1LE) and xeroderma pigmentosum (XP2LE) patients with chromosomal sensitivity to ionizing radiation. Doses of γ-irradiation that markedly inhibited the rate of DNA synthesis in normal human cells caused almost no inhibition of DNA synthesis in the cells from the affected individuals. The radioresistant DNA synthesis in Down's syndrome cells was mainly due to a much lower inhibition of replicon initiation than that in normal cells; these cells were also more resistant to damage that inhibited replicon elongation. Our data suggest that radioresistant DNA synthesis may be an intrinsic feature of all genetic disorders showing increased radiosensitivity in terms of chromosome aberrations. (orig.)

  1. Selection of radioresistant cells by vitamin A deficiency in a small cell lung cancer cell line

    International Nuclear Information System (INIS)

    Terasaki, Takeo; Shimosato, Yukio; Wada, Makio; Yokota, Jun; Terada, Masaaki

    1990-01-01

    Radiation sensitivity of a human small cell lung cancer cell line, Lu-134-B cells, cultured in serum-supplemented medium and of cells transferred to and cultured in delipidized serum-supplemented (vitamin A-deficient) medium was studied. The cells cultured in serum-supplemented medium showed the phenotype of classic small cell lung cancer sensitive to radiation, while cells transferred to delipidized serum-supplemented medium showed partial squamous cell differentiation and became resistant to radiation. These results suggest that some small cell lung cancer cells in vitro change their morphology and radiosensitivity depending on the culture conditions. The change in radiosensitivity was reproducible, and was not reversible by culture of the radioresistant cells in delipidized serum-supplemented medium with addition of retinoic acid (vitamin A-sufficient medium) for two months, although squamous cells disappeared. Acquisition of radioresistancy was considered to occur as the result of clonal selective growth in delipidized medium of a minor cell population in the original cell culture, based on a study of chromosome number. It was also found that there was no association of myc-family oncogenes with the changes of radiosensitivity in this cell line. (author)

  2. Reprogramming mediated radio-resistance of 3D-grown cancer cells

    International Nuclear Information System (INIS)

    Xue Gang; Ren Zhenxin; Chen Yaxiong; Zhu Jiayun; Du Yarong; Pan Dong; Li Xiaoman; Hu Burong; Grabham, Peter W.

    2015-01-01

    In vitro 3D growth of tumors is a new cell culture model that more closely mimics the features of the in vivo environment and is being used increasingly in the field of biological and medical research. It has been demonstrated that cancer cells cultured in 3D matrices are more radio-resistant compared with cells in monolayers. However, the mechanisms causing this difference remain unclear. Here we show that cancer cells cultured in a 3D microenvironment demonstrated an increase in cells with stem cell properties. This was confirmed by the finding that cells in 3D cultures upregulated the gene and protein expression of the stem cell reprogramming factors such as OCT4, SOX2, NANOG, LIN28 and miR-302a, compared with cells in monolayers. Moreover, the expression of β-catenin, a regulating molecule of reprogramming factors, also increased in 3D-grown cancer cells. These findings suggest that cancer cells were reprogrammed to become stem cell-like cancer cells in a 3D growth culture microenvironment. Since cancer stem cell-like cells demonstrate an increased radio-resistance and chemo-resistance, our results offer a new perspective as to why. Our findings shed new light on understanding the features of the 3D growth cell model and its application in basic research into clinical radiotherapy and medicine. (author)

  3. Receptor-mediated endocytosis and intracellular trafficking of insulin and low-density lipoprotein by retinal vascular endothelial cells.

    Science.gov (United States)

    Stitt, A W; Anderson, H R; Gardiner, T A; Bailie, J R; Archer, D B

    1994-08-01

    The authors investigated the receptor-mediated endocytosis (RME) and intracellular trafficking of insulin and low-density lipoprotein (LDL) in cultured retinal vascular endothelial cells (RVECs). Low-density lipoprotein and insulin were conjugated to 10 nm colloidal gold, and these ligands were added to cultured bovine RVECs for 20 minutes at 4 degrees C. The cultures were then warmed to 37 degrees C and fixed after incubation times between 30 seconds and 1 hour. Control cells were incubated with unconjugated gold colloid at times and concentrations similar to those of the ligands. Additional control cells were exposed to several concentrations of anti-insulin receptor antibody or a saturating solution of unconjugated insulin before incubation with gold insulin. Using transmission electron microscopy, insulin gold and LDL gold were both observed at various stages of RME. Insulin-gold particles were first seen to bind to the apical plasma membrane (PM) before clustering in clathrin-coated pits and internalization in coated vesicles. Gold was later visualized in uncoated cytoplasmic vesicles, corresponding to early endosomes and multivesicular bodies (MVBs) or late endosomes. In several instances, localized regions of the limiting membrane of the MVBs appeared coated, a feature of endosomal membranes not previously described. After RME at the apical PM and passage through the endosomal system, the greater part of both insulin- and LDL-gold conjugates was seen to accumulate in large lysosome-like compartments. However, a small but significant proportion of the internalized ligands was transcytosed and released as discrete membrane-associated quanta at the basal cell surface. The uptake of LDL gold was greatly increased in highly vacuolated, late-passage RVECs. In controls, anti-insulin receptor antibody and excess unconjugated insulin caused up to 89% inhibition in gold-insulin binding and internalization. These results illustrate the internalization and intracellular

  4. Dopamine D2 receptors mediate two-odor discrimination and reversal learning in C57BL/6 mice

    Directory of Open Access Journals (Sweden)

    Grandy David K

    2004-04-01

    Full Text Available Abstract Background Dopamine modulation of neuronal signaling in the frontal cortex, midbrain, and striatum is essential for processing and integrating diverse external sensory stimuli and attaching salience to environmental cues that signal causal relationships, thereby guiding goal-directed, adaptable behaviors. At the cellular level, dopamine signaling is mediated through D1-like or D2-like receptors. Although a role for D1-like receptors in a variety of goal-directed behaviors has been identified, an explicit involvement of D2 receptors has not been clearly established. To determine whether dopamine D2 receptor-mediated signaling contributes to associative and reversal learning, we compared C57Bl/6J mice that completely lack functional dopamine D2 receptors to wild-type mice with respect to their ability to attach appropriate salience to external stimuli (stimulus discrimination and disengage from inappropriate behavioral strategies when reinforcement contingencies change (e.g. reversal learning. Results Mildly food-deprived female wild-type and dopamine D2 receptor deficient mice rapidly learned to retrieve and consume visible food reinforcers from a small plastic dish. Furthermore, both genotypes readily learned to dig through the same dish filled with sterile sand in order to locate a buried food pellet. However, the dopamine D2 receptor deficient mice required significantly more trials than wild-type mice to discriminate between two dishes, each filled with a different scented sand, and to associate one of the two odors with the presence of a reinforcer (food. In addition, the dopamine D2 receptor deficient mice repeatedly fail to alter their response patterns during reversal trials where the reinforcement rules were inverted. Conclusions Inbred C57Bl/6J mice that develop in the complete absence of functional dopamine D2 receptors are capable of olfaction but display an impaired ability to acquire odor-driven reinforcement contingencies

  5. Hypersensitivity to thromboxane receptor mediated cerebral vasomotion and CBF oscillations during acute NO-deficiency in rats.

    Directory of Open Access Journals (Sweden)

    Béla Horváth

    Full Text Available BACKGROUND: Low frequency (4-12 cpm spontaneous fluctuations of the cerebrovascular tone (vasomotion and oscillations of the cerebral blood flow (CBF have been reported in diseases associated with endothelial dysfunction. Since endothelium-derived nitric oxide (NO suppresses constitutively the release and vascular effects of thromboxane A(2 (TXA(2, NO-deficiency is often associated with activation of thromboxane receptors (TP. In the present study we hypothesized that in the absence of NO, overactivation of the TP-receptor mediated cerebrovascular signaling pathway contributes to the development of vasomotion and CBF oscillations. METHODOLOGY/PRINCIPAL FINDINGS: Effects of pharmacological modulation of TP-receptor activation and its downstream signaling pathway have been investigated on CBF oscillations (measured by laser-Doppler flowmetry in anesthetized rats and vasomotion (measured by isometric tension recording in isolated rat middle cerebral arteries, MCAs both under physiological conditions and after acute inhibition of NO synthesis. Administration of the TP-receptor agonist U-46619 (1 µg/kg i.v. to control animals failed to induce any changes of the systemic or cerebral circulatory parameters. Inhibition of the NO synthesis by nitro-L-arginine methyl ester (L-NAME, 100 mg/kg i.v. resulted in increased mean arterial blood pressure and a decreased CBF accompanied by appearance of CBF-oscillations with a dominant frequency of 148±2 mHz. U-46619 significantly augmented the CBF-oscillations induced by L-NAME while inhibition of endogenous TXA(2 synthesis by ozagrel (10 mg/kg i.v. attenuated it. In isolated MCAs U-46619 in a concentration of 100 nM, which induced weak and stable contraction under physiological conditions, evoked sustained vasomotion in the absence of NO, which effect could be completely reversed by inhibition of Rho-kinase by 10 µM Y-27632. CONCLUSION/SIGNIFICANCE: These results suggest that hypersensitivity of the TP

  6. Preservation of differentiation and clonogenic potential of human hematopoietic stem and progenitor cells during lyophilization and ambient storage.

    Science.gov (United States)

    Buchanan, Sandhya S; Pyatt, David W; Carpenter, John F

    2010-09-01

    Progenitor cell therapies show great promise, but their potential for clinical applications requires improved storage and transportation. Desiccated cells stored at ambient temperature would provide economic and practical advantages over approaches employing cell freezing and subzero temperature storage. The objectives of this study were to assess a method for loading the stabilizing sugar, trehalose, into hematopoietic stem and progenitor cells (HPC) and to evaluate the effects of subsequent freeze-drying and storage at ambient temperature on differentiation and clonogenic potential. HPC were isolated from human umbilical cord blood and loaded with trehalose using an endogenous cell surface receptor, termed P2Z. Solution containing trehalose-loaded HPC was placed into vials, which were transferred to a tray freeze-dryer and removed during each step of the freeze-drying process to assess differentiation and clonogenic potential. Control groups for these experiments were freshly isolated HPC. Control cells formed 1450+/-230 CFU-GM, 430+/-140 BFU-E, and 50+/-40 CFU-GEMM per 50 microL. Compared to the values for the control cells, there was no statistical difference observed for cells removed at the end of the freezing step or at the end of primary drying. There was a gradual decrease in the number of CFU-GM and BFU-E for cells removed at different temperatures during secondary drying; however, there were no significant differences in the number of CFU-GEMM. To determine storage stability of lyophilized HPC, cells were stored for 4 weeks at 25 degrees C in the dark. Cells reconstituted immediately after lyophilization produced 580+/-90 CFU-GM ( approximately 40%, relative to unprocessed controls pGM (approximately 35%, relative to unprocessed controls, p<0.0001), 112+/-68 BFU-E (approximately 26%, p<0.0001), and 36+/-17 CFU-GEMM ( approximately 82%, p = 0.2164) These studies are the first to document high level retention of CFU-GEMM following lyophilization and

  7. Preservation of differentiation and clonogenic potential of human hematopoietic stem and progenitor cells during lyophilization and ambient storage.

    Directory of Open Access Journals (Sweden)

    Sandhya S Buchanan

    2010-09-01

    Full Text Available Progenitor cell therapies show great promise, but their potential for clinical applications requires improved storage and transportation. Desiccated cells stored at ambient temperature would provide economic and practical advantages over approaches employing cell freezing and subzero temperature storage. The objectives of this study were to assess a method for loading the stabilizing sugar, trehalose, into hematopoietic stem and progenitor cells (HPC and to evaluate the effects of subsequent freeze-drying and storage at ambient temperature on differentiation and clonogenic potential. HPC were isolated from human umbilical cord blood and loaded with trehalose using an endogenous cell surface receptor, termed P2Z. Solution containing trehalose-loaded HPC was placed into vials, which were transferred to a tray freeze-dryer and removed during each step of the freeze-drying process to assess differentiation and clonogenic potential. Control groups for these experiments were freshly isolated HPC. Control cells formed 1450+/-230 CFU-GM, 430+/-140 BFU-E, and 50+/-40 CFU-GEMM per 50 microL. Compared to the values for the control cells, there was no statistical difference observed for cells removed at the end of the freezing step or at the end of primary drying. There was a gradual decrease in the number of CFU-GM and BFU-E for cells removed at different temperatures during secondary drying; however, there were no significant differences in the number of CFU-GEMM. To determine storage stability of lyophilized HPC, cells were stored for 4 weeks at 25 degrees C in the dark. Cells reconstituted immediately after lyophilization produced 580+/-90 CFU-GM ( approximately 40%, relative to unprocessed controls p<0.0001, 170+/-70 BFU-E (approximately 40%, p<0.0001, and 41+/-22 CFU-GEMM (approximately 82%, p = 0.4171, and cells reconstituted after 28 days at room temperature produced 513+/-170 CFU-GM (approximately 35%, relative to unprocessed controls, p<0

  8. The repair/repopulation effects ratio in the postirradiation recovery of hexaploid wheat varieties contrast by their radioresistance

    International Nuclear Information System (INIS)

    Selezneva, E.M.; Sarapul'tsev, B.I.

    1988-01-01

    The cytogenic and morphometrical distinctions between hexaploid wheat varieties contrast by their radioresistance during the postirradiation period are attributed to the differential activity of caffeine-dependent repair processes; they are not a reliable function of the rate of aberrant cell elimination

  9. Microbial radio-resistance of Salmonella Typhimurium in egg increases due to repetitive irradiation with electron beam

    International Nuclear Information System (INIS)

    Tesfai, Adiam T.; Beamer, Sarah K.; Matak, Kristen E.; Jaczynski, Jacek

    2011-01-01

    Ionizing radiation improves food safety. However, foodborne pathogens develop increased resistance in response to sub-lethal stresses such as heat, pH, antibiotics, etc. Therefore, it is hypothesized that foodborne pathogens may develop increased radio-resistance to electron beam (e-beam) radiation. The objective was to determine if D 10 -value for Salmonella Typhimurium in de-shelled raw egg (egg white and yolk mixed together) increases due to repetitive processing with e-beam at sub-lethal doses. Survivors were enumerated on non-selective (TSA) and selective (XLD) media. Survivors from the highest dose were isolated and used in subsequent e-beam cycle. This process was repeated four times for a total of five e-beam cycles. D 10 -values for S. Typhimurium enumerated on TSA and XLD following each e-beam cycle were calculated as inverse reciprocal of the slope of survivor curves. D 10 -values for the ATCC strain were 0.59±0.031 and 0.46±0.022 kGy on TSA and XLD, respectively. However, following the fifth e-beam cycle, the respective D 10 -values increased (P 0.05) to develop radio-resistance faster on selective media, likely due to facilitated selection of radio-resistant cells within microbial population following each e-beam cycle. For all five e-beam cycles, S. Typhimurium had higher (P 10 -values on non-selective media, indicating that sub-lethal injury followed by cellular repair and recovery are important for radio-resistance and inactivation of this microorganism. This study demonstrated that e-beam efficiently inactivates S. Typhimurium in raw egg; however, similar to other inactivation techniques and factors affecting microbial growth, S. Typhimurium develops increased radio-resistance if repetitively processed with e-beam at sub-lethal doses.

  10. Neutron and photon clonogenic survival curves of two chemotherapy resistant human intermediate-grade non-Hodgkin lymphoma cell lines

    International Nuclear Information System (INIS)

    Aref, Amr; Yudelev, Mark; Mohammad, Ramzi; Choudhuri, Rajani; Orton, Colin; Al-Katib, Ayad

    1999-01-01

    Background: The potential role of neutron therapy in the management of intermediate-grade non-Hodgkin lymphoma (IGNHL) has not been examined because of the belief that the anticipated radiobiological effectiveness (RBE) would be uniformly very low. Purpose: To determine the fast neutron RBE for two chemotherapy-resistant IGNHL cell lines. Methods and Materials: Conventional soft agar clonogenic survival curves following irradiation by 60 Co and fast neutron were established for two IGNHL cell lines. These cell lines, WSU-DLCL2 and SK-DHL2B, were found in previous studies to be able to repair sublethal damage, and were also resistant to L-Pam and doxorubicin chemotherapy. Results: When the surviving fraction after 2 Gy photon was chosen as the biological endpoint, the RBE for WSU-DLCL2 and SK-DHL2B measured 3.34 and 3.06. Similarly, when 10% survival was considered, the RBE for these two cell lines measured 2.54 and 2.59. The RBE, as measured by the ratios α neutron/α photon, for WSU-DLCL2, SK-DHL2B cell lines are 6.67 and 5.65, respectively. These results indicate that the RBE for these IGNHL cell lines is higher than the average RBE for cell lines of other histological types. Conclusion: Fast neutron irradiation may be of potential value in treating selected cases of IGNHL

  11. Radioresistant DNA synthesis in fibroblasts of a patient with Down's syndrome

    International Nuclear Information System (INIS)

    Barenfel'd, L.S.; Bil'din, V.N.; Pleskach, N.M.; Prokof'eva, V.V.

    1985-01-01

    Ionizing radiation effect on DNA replication on fibroblasts of a healthy donor and a patient with Down's syndrome either by direct 3 H-thymidine inclusion into DNA, or by analysis of the sizes of daughter DNA moleculs at the state of stable distribution in acid saccharose, gradients was studied. Gamma-radiation doses (5-10 Gy) suppressing DNA synthesis in normal fibroblasts practically had no effect on DNA synthesisin fibroblasts of a patient with Down's syndrome. Radioresistant DNA synthesis in Down's syndrome is conditioned by a far less supression of replicon initiation as compared with the one in normal cells. So, it is stated that in Down's disease there is no delay in DNA synthesis by ionizing radiation that enables the normal cells to repair DNA damages before replication renewal

  12. Investigations on radiosensitive and radioresistant populations of Drosophila melanogaster. Pt. 10

    International Nuclear Information System (INIS)

    Noethel, H.

    1981-01-01

    In earlier work, immature oocytes of the irradiated population RoeI 4 of Drosophila melanogaster were found to be radioresistant relative to those of the basic population RoeI and to those of the control population Berlin wild (+K). The resistance of RoeI 4 relative to RoeI was previously attributed to a hypothetical 'factor' rar-3. In the present paper, evidence is presented to show that rar-3 is a single, recessive genetic factor, located on chromosome 3 at a map position of about 49.8. The action of rar-3 is apparently independent of that of rar-1 and rar-2, the factors already present in RoeI. (orig.)

  13. Metabolic analysis of radioresistant medulloblastoma stem-like clones and potential therapeutic targets.

    Directory of Open Access Journals (Sweden)

    Lue Sun

    Full Text Available Medulloblastoma is a fatal brain tumor in children, primarily due to the presence of treatment-resistant medulloblastoma stem cells. The energy metabolic pathway is a potential target of cancer therapy because it is often different between cancer cells and normal cells. However, the metabolic properties of medulloblastoma stem cells, and whether specific metabolic pathways are essential for sustaining their stem cell-like phenotype and radioresistance, remain unclear. We have established radioresistant medulloblastoma stem-like clones (rMSLCs by irradiation of the human medulloblastoma cell line ONS-76. Here, we assessed reactive oxygen species (ROS production, mitochondria function, oxygen consumption rate (OCR, energy state, and metabolites of glycolysis and tricarboxylic acid cycle in rMSLCs and parental cells. rMSLCs showed higher lactate production and lower oxygen consumption rate than parental cells. Additionally, rMSLCs had low mitochondria mass, low endogenous ROS production, and existed in a low-energy state. Treatment with the metabolic modifier dichloroacetate (DCA resulted in mitochondria dysfunction, glycolysis inhibition, elongated mitochondria morphology, and increased ROS production. DCA also increased radiosensitivity by suppression of the DNA repair capacity through nuclear oxidization and accelerated the generation of acetyl CoA to compensate for the lack of ATP. Moreover, treatment with DCA decreased cancer stem cell-like characters (e.g., CD133 positivity and sphere-forming ability in rMSLCs. Together, our findings provide insights into the specific metabolism of rMSLCs and illuminate potential metabolic targets that might be exploited for therapeutic benefit in medulloblastoma.

  14. Cyclophilin B Expression Is Associated with In Vitro Radioresistance and Clinical Outcome after Radiotherapy

    Directory of Open Access Journals (Sweden)

    Paul D. Williams

    2011-12-01

    Full Text Available The tools for predicting clinical outcome after radiotherapy are not yet optimal. To improve on this, we applied the COXEN informatics approach to in vitro radiation sensitivity data of transcriptionally profiled human cells and gene expression data from untreated head and neck squamous cell carcinoma (HNSCC and bladder tumors to generate a multigene predictive model that is independent of histologic findings and reports on tumor radiosensitivity. The predictive ability of this 41-gene model was evaluated in patients with HNSCC and was found to stratify clinical outcome after radiotherapy. In contrast, this model was not useful in stratifying similar patients not treated with radiation. This led us to hypothesize that expression of some of the 41 genes contributes to tumor radioresistance and clinical recurrence. Hence, we evaluated the expression the 41 genes as a function of in vitro radioresistance in the NCI-60 cancer cell line panel and found cyclophilin B (PPIB, a peptidylprolyl isomerase and target of cyclosporine A (CsA, had the strongest direct correlation. Functional inhibition of PPIB by small interfering RNA depletion or CsA treatment leads to radiosensitization in cancer cells and reduced cellular DNA repair. Immunohistochemical evaluation of PPIB expression in patients with HNSCC was found to be associated with outcome after radiotherapy. This work demonstrates that a novel 41-gene expression model of radiation sensitivity developed in bladder cancer cell lines and human skin fibroblasts predicts clinical outcome after radiotherapy in head and neck cancer patients and identifies PPIB as a potential target for clinical radiosensitization.

  15. Inhibition of GRP78 abrogates radioresistance in oropharyngeal carcinoma cells after EGFR inhibition by cetuximab.

    Directory of Open Access Journals (Sweden)

    Chaonan Sun

    Full Text Available The EGFR-specific mAb cetuximab is one of the most effective treatments for oropharyngeal carcinoma, while patient responses to EGFR inhibitors given alone are modest. Combination treatment with radiation can improve the efficacy of treatment through increasing radiosensitivity, while resistance to radiation after administration of cetuximab limits its efficiency. Radiation and drugs can damage the endoplasmic reticulum (ER homeostatic state and result in ER stress (ERS, subsequently causing resistance to radiation and drugs. Whether the ERS pathway is involved in radioresistance after administration of cetuximab has not been reported. Herein, we show that cetuximab could increase the radiosensitivity of FaDu cells but not Detroit562 cells. In addition, cetuximab inhibited the radiation-induced activation of the ERS signalling pathway IRE1α/ATF6-GRP78 in FaDu cells, while this effect was absent in Detroit562 cells. Silencing GRP78 increased the radiosensitivity of oropharyngeal carcinoma cells and inhibited radiation-induced DNA double-strand-break (DSB repair and autophagy. More interestingly, silencing GRP78 abrogated resistance to cetuximab and radiation in Detroit562 cells and had a synergistic effect with cetuximab in increasing the radiosensitivity of FaDu cells. Immunohistochemistry showed that overexpression of both GRP78 and EGFR was associated with a poor prognosis in oropharyngeal carcinoma patients (P<0.05. Overall, the results of this study show that radioresistance after EGFR inhibition by cetuximab is mediated by the ERS signalling pathway IRE1α/ATF6-GRP78. This suppression was consequently unable to inhibit radiation-induced DSB repair and autophagy in oropharyngeal carcinoma cells, which conferred resistance to radiotherapy and cetuximab. These results suggest that the cooperative effects of radiotherapy and cetuximab could be further improved by inhibiting GRP78 in non-responsive oropharyngeal carcinoma patients.

  16. Mechanism of derivation of radioresistance in HeLa cell population after repeated x-irradiation

    International Nuclear Information System (INIS)

    Kubo, Kihei; Koiwai, Soichiro; Morita, Kazuo

    1982-01-01

    The Radioresistant strain (X-8-5) was obtained from HeLa-SC population X-irradiated repeatedly for five times with 800 rad. The mean lethal dose (D 0 ) was 196 rad for X-8-5 cells, while it was 166 rad for control HeLa-SC cells. The fraction of cells containing an unusually long acrocentric chromosome (LA 2) exclusively increased with increasing number of irradiation of HeLa-SC population. A clonal strain with LA 2 marker was isolated from X-8-5 population and named RC-355. Since the RC-355 cells were more resistant (D 0 = 220 rad)than parental X-8-5 cells (D 0 = 196 rad), it was suggested that the cells with LA 2 were responsible for the radioresistance of X-8-5 population. The RC-355 cells were further subjected to the analysis of Q-banded karyotypes and it was observed that 18 types of specific markers (rm 1-17 and LA 2) were included in RC-355 cells in addition to 12 types of markers observed in most of HeLa-SC cells. Since the analysis of Q-banded karyotypes of RC-355 cells showed that RC-355 specific markers were not produced by radiation-induced rearrangements of HeLa-SC chromosomes, because twelve kinds of HeLa-SC markers were presented in RC-355 cells without any change, it was concluded that a small number of cells with LA 2 marker were originally presented in the control population and the relative fraction of them occupied increased after irradiation. (author)

  17. Alpha 1-adrenergic receptor-mediated phosphoinositide hydrolysis and prostaglandin E2 formation in Madin-Darby canine kidney cells. Possible parallel activation of phospholipase C and phospholipase A2

    International Nuclear Information System (INIS)

    Slivka, S.R.; Insel, P.A.

    1987-01-01

    alpha 1-Adrenergic receptors mediate two effects on phospholipid metabolism in Madin-Darby canine kidney (MDCK-D1) cells: hydrolysis of phosphoinositides and arachidonic acid release with generation of prostaglandin E2 (PGE2). The similarity in concentration dependence for the agonist (-)-epinephrine in eliciting these two responses implies that they are mediated by a single population of alpha 1-adrenergic receptors. However, we find that the kinetics of the two responses are quite different, PGE2 production occurring more rapidly and transiently than the hydrolysis of phosphoinositides. The antibiotic neomycin selectively decreases alpha 1-receptor-mediated phosphatidylinositol 4,5-bisphosphate hydrolysis without decreasing alpha 1-receptor-mediated arachidonic acid release and PGE2 generation. In addition, receptor-mediated inositol trisphosphate formation is independent of extracellular calcium, whereas release of labeled arachidonic acid is largely calcium-dependent. Moreover, based on studies obtained with labeled arachidonic acid, receptor-mediated generation of arachidonic acid cannot be accounted for by breakdown of phosphatidylinositol monophosphate, phosphatidylinositol bisphosphate, or phosphatidic acid. Further studies indicate that epinephrine produces changes in formation or turnover of several classes of membrane phospholipids in MDCK cells. We conclude that alpha 1-adrenergic receptors in MDCK cells appear to regulate phospholipid metabolism by the parallel activation of phospholipase C and phospholipase A2. This parallel activation of phospholipases contrasts with models described in other systems which imply sequential activation of phospholipase C and diacylglycerol lipase or phospholipase A2

  18. Receptor-mediated endocytosis of polypeptide hormones is a regulated process: inhibition of [125I]iodoinsulin internalization in hypoinsulinemic diabetes of rat and man

    International Nuclear Information System (INIS)

    Carpentier, J.L.; Robert, A.; Grunberger, G.; van Obberghen, E.; Freychet, P.; Orci, L.; Gorden, P.

    1986-01-01

    Much data suggest that receptor-mediated endocytosis is regulated in states of hormone excess. Thus, in hyperinsulinemic states there is an accelerated loss of cell surface insulin receptors. In the present experiments we addressed this question in hypoinsulinemic states, in which insulin binding to cell surface receptors is generally increased. In hepatocytes obtained from hypoinsulinemic streptozotocin-induced diabetic rats, [ 125 I]iodoglucagon internalization was increased, while at the same time [ 125 I]iodoinsulin internalization was decreased. The defect in [ 125 I]iodoinsulin internalization was corrected by insulin treatment of the animal. In peripheral blood monocytes from patients with type I insulinopenic diabetes, internalization of [ 125 I]iodoinsulin was impaired; this defect was not present in insulin-treated patients. These data in the hypoinsulinemic rat and human diabetes suggest that receptor-mediated endocytosis is regulated in states of insulin deficiency as well as insulin excess. Delayed or reduced internalization of the insulin-receptor complex could amplify the muted signal caused by deficient hormone secretion

  19. Nitric oxide inhibits the bradykinin B2 receptor-mediated adrenomedullary catecholamine release but has no effect on adrenal blood flow response in vivo.

    Science.gov (United States)

    Bouallegue, Ali; Yamaguchi, Nobuharu

    2005-06-01

    The role of nitric oxide (NO) in bradykinin (BK)-induced adrenal catecholamine secretion still remains obscure. The present study was to investigate whether an inhibition of NO synthase with N(omega)-nitro-L-arginine methyl ester (L-NAME) would modulate BK-induced adrenal catecholamine secretion (ACS) and adrenal vasodilating response (AVR) in anesthetized dogs. Plasma catecholamine concentrations were determined with an HPLC coupled with an electrochemical detector. All drugs were locally administered to the left adrenal gland via intra-arterial infusion. BK dose-dependently increased both ACS and AVR. Hoe-140, a selective B(2) antagonist, significantly blocked the BK-induced increases in both ACS and AVR. In the presence of L-NAME, the BK-induced ACS was significantly enhanced, while the simultaneous AVR remained unaffected. These results suggest that the both BK-induced ACS and AVR are primarily mediated by B(2) receptors in the canine adrenal gland. Our results also suggest that the enhanced ACS in response to BK in the presence of L-NAME may have resulted from a specific inhibition of NO formation in the adrenal gland. It is concluded that the BK-induced NO may play an inhibitory role in the B(2)-receptor-mediated mechanisms regulating ACS, while it may not be implicated in the B(2)-receptor-mediated AVR under in vivo conditions.

  20. USP1 targeting impedes GBM growth by inhibiting stem cell maintenance and radioresistance.

    Science.gov (United States)

    Lee, Jin-Ku; Chang, Nakho; Yoon, Yeup; Yang, Heekyoung; Cho, Heejin; Kim, Eunhee; Shin, Yongjae; Kang, Wonyoung; Oh, Young Taek; Mun, Gyeong In; Joo, Kyeung Min; Nam, Do-Hyun; Lee, Jeongwu

    2016-01-01

    Clinical benefits from standard therapies against glioblastoma (GBM) are limited in part due to intrinsic radio- and chemoresistance of GBM and inefficient targeting of GBM stem-like cells (GSCs). Novel therapeutic approaches that overcome treatment resistance and diminish stem-like properties of GBM are needed. We determined the expression levels of ubiquitination-specific proteases (USPs) by transcriptome analysis and found that USP1 is highly expressed in GBM. Using the patient GBM-derived primary tumor cells, we inhibited USP1 by shRNA-mediated knockdown or its specific inhibitor pimozide and evaluated the effects on stem cell marker expression, proliferation, and clonogenic growth of tumor cells. USP1 was highly expressed in gliomas relative to normal brain tissues and more preferentially in GSC enrichment marker (CD133 or CD15) positive cells. USP1 positively regulated the protein stability of the ID1 and CHEK1, critical regulators of DNA damage response and stem cell maintenance. Targeting USP1 by RNA interference or treatment with a chemical USP1 inhibitor attenuated clonogenic growth and survival of GSCs and enhanced radiosensitivity of GBM cells. Finally, USP1 inhibition alone or in combination with radiation significantly prolonged the survival of tumor-bearing mice. USP1-mediated protein stabilization promotes GSC maintenance and treatment resistance, thereby providing a rationale for USP1 inhibition as a potential therapeutic approach against GBM. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  1. Effect of interleukin-9 on clonogenic maturation and cell-cycle status of fetal and adult hematopoietic progenitors

    Energy Technology Data Exchange (ETDEWEB)

    Holbrook, S.T.; Ohls, R.K.; Schibler, K.R.; Yang, Y.C.; Christensen, R.D. (Univ. of Utah School of Medicine, Salt Lake City (USA))

    1991-05-15

    We assessed the effect of interleukin-9 (IL-9) on clonogenic maturation and cell-cycle status of hematopoietic progenitors of fetal (umbilical cord blood) and adult (bone marrow) origin. As a single agent IL-9 supported, in a concentration-dependent fashion, maturation of burst-forming units-erythroid (BFU-E) of adult and fetal origin. However, only 1/3 the number of adult BFU-E colonies developed, as did in response to granulocyte-macrophage colony-stimulating factor (GM-CSF), and only 1/6 the number developed as did in response to IL-3. In contrast, the effect of IL-9 on fetal BFU-E colonies was equal to that of GM-CSF and IL-3. Synergistic effects of IL-9 with low concentrations (0.1 ng/mL) of GM-CSF and IL-3 were seen on adult BFU-E colony formation, but no effect was apparent at higher concentrations (1.0 ng/mL). In contrast, using fetal cells, synergistic effects of IL-9 with low and high concentrations of GM-CSF and IL-3 were apparent. Addition of IL-9 to plates containing fetal cells plus GM-CSF and IL-3 not only resulted in more BFU-E colonies, but also in more multicentered (greater than or equal to 10 individual centers) colonies, and more cells per colony. IL-9 had a wider spectrum of action on progenitors of fetal origin than on progenitors of adult origin, supporting the generation of fetal multipotent colony-forming unit (CFU)-Mix and CFU-GM colonies. Incubation with IL-9 did not accelerate cycling of adult or fetal BFU-E, CFU-Mix, or CFU-GM to the extent observed after incubation with IL-6.

  2. Effect of interleukin-9 on clonogenic maturation and cell-cycle status of fetal and adult hematopoietic progenitors

    International Nuclear Information System (INIS)

    Holbrook, S.T.; Ohls, R.K.; Schibler, K.R.; Yang, Y.C.; Christensen, R.D.

    1991-01-01

    We assessed the effect of interleukin-9 (IL-9) on clonogenic maturation and cell-cycle status of hematopoietic progenitors of fetal (umbilical cord blood) and adult (bone marrow) origin. As a single agent IL-9 supported, in a concentration-dependent fashion, maturation of burst-forming units-erythroid (BFU-E) of adult and fetal origin. However, only 1/3 the number of adult BFU-E colonies developed, as did in response to granulocyte-macrophage colony-stimulating factor (GM-CSF), and only 1/6 the number developed as did in response to IL-3. In contrast, the effect of IL-9 on fetal BFU-E colonies was equal to that of GM-CSF and IL-3. Synergistic effects of IL-9 with low concentrations (0.1 ng/mL) of GM-CSF and IL-3 were seen on adult BFU-E colony formation, but no effect was apparent at higher concentrations (1.0 ng/mL). In contrast, using fetal cells, synergistic effects of IL-9 with low and high concentrations of GM-CSF and IL-3 were apparent. Addition of IL-9 to plates containing fetal cells plus GM-CSF and IL-3 not only resulted in more BFU-E colonies, but also in more multicentered (greater than or equal to 10 individual centers) colonies, and more cells per colony. IL-9 had a wider spectrum of action on progenitors of fetal origin than on progenitors of adult origin, supporting the generation of fetal multipotent colony-forming unit (CFU)-Mix and CFU-GM colonies. Incubation with IL-9 did not accelerate cycling of adult or fetal BFU-E, CFU-Mix, or CFU-GM to the extent observed after incubation with IL-6

  3. Genome-wide analyses of long noncoding RNA expression profiles correlated with radioresistance in nasopharyngeal carcinoma via next-generation deep sequencing.

    Science.gov (United States)

    Li, Guo; Liu, Yong; Liu, Chao; Su, Zhongwu; Ren, Shuling; Wang, Yunyun; Deng, Tengbo; Huang, Donghai; Tian, Yongquan; Qiu, Yuanzheng

    2016-09-06

    Radioresistance is one of the major factors limiting the therapeutic efficacy and prognosis of patients with nasopharyngeal carcinoma (NPC). Accumulating evidence has suggested that aberrant expression of long noncoding RNAs (lncRNAs) contributes to cancer progression. Therefore, here we identified lncRNAs associated with radioresistance in NPC. The differential expression profiles of lncRNAs associated with NPC radioresistance were constructed by next-generation deep sequencing by comparing radioresistant NPC cells with their parental cells. LncRNA-related mRNAs were predicted and analyzed using bioinformatics algorithms compared with the mRNA profiles related to radioresistance obtained in our previous study. Several lncRNAs and associated mRNAs were validated in established NPC radioresistant cell models and NPC tissues. By comparison between radioresistant CNE-2-Rs and parental CNE-2 cells by next-generation deep sequencing, a total of 781 known lncRNAs and 2054 novel lncRNAs were annotated. The top five upregulated and downregulated known/novel lncRNAs were detected using quantitative real-time reverse transcription-polymerase chain reaction, and 7/10 known lncRNAs and 3/10 novel lncRNAs were demonstrated to have significant differential expression trends that were the same as those predicted by deep sequencing. From the prediction process, 13 pairs of lncRNAs and their associated genes were acquired, and the prediction trends of three pairs were validated in both radioresistant CNE-2-Rs and 6-10B-Rs cell lines, including lncRNA n373932 and SLITRK5, n409627 and PRSS12, and n386034 and RIMKLB. LncRNA n373932 and its related SLITRK5 showed dramatic expression changes in post-irradiation radioresistant cells and a negative expression correlation in NPC tissues (R = -0.595, p < 0.05). Our study provides an overview of the expression profiles of radioresistant lncRNAs and potentially related mRNAs, which will facilitate future investigations into the

  4. Microarray analysis of DNA damage repair gene expression profiles in cervical cancer cells radioresistant to 252Cf neutron and X-rays

    International Nuclear Information System (INIS)

    Qing, Yi; Wang, Ge; Wang, Dong; Yang, Xue-Qin; Zhong, Zhao-Yang; Lei, Xin; Xie, Jia-Yin; Li, Meng-Xia; Xiang, De-Bing; Li, Zeng-Peng; Yang, Zhen-Zhou

    2010-01-01

    The aim of the study was to obtain stable radioresistant sub-lines from the human cervical cancer cell line HeLa by prolonged exposure to 252 Cf neutron and X-rays. Radioresistance mechanisms were investigated in the resulting cells using microarray analysis of DNA damage repair genes. HeLa cells were treated with fractionated 252 Cf neutron and X-rays, with a cumulative dose of 75 Gy each, over 8 months, yielding the sub-lines HeLaNR and HeLaXR. Radioresistant characteristics were detected by clone formation assay, ultrastructural observations, cell doubling time, cell cycle distribution, and apoptosis assay. Gene expression patterns of the radioresistant sub-lines were studied through microarray analysis and verified by Western blotting and real-time PCR. The radioresistant sub-lines HeLaNR and HeLaXR were more radioresisitant to 252 Cf neutron and X-rays than parental HeLa cells by detecting their radioresistant characteristics, respectively. Compared to HeLa cells, the expression of 24 genes was significantly altered by at least 2-fold in HeLaNR cells. Of these, 19 genes were up-regulated and 5 down-regulated. In HeLaXR cells, 41 genes were significantly altered by at least 2-fold; 38 genes were up-regulated and 3 down-regulated. Chronic exposure of cells to ionizing radiation induces adaptive responses that enhance tolerance of ionizing radiation and allow investigations of cellular radioresistance mechanisms. The insights gained into the molecular mechanisms activated by these 'radioresistance' genes will lead to new therapeutic targets for cervical cancer

  5. Differential modulation of expression of nuclear receptor mediated genes by tris(2-butoxyethyl) phosphate (TBOEP) on early life stages of zebrafish (Danio rerio)

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Zhiyuan, E-mail: zhiyuan_nju@163.com [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Yu, Yijun, E-mail: yjun.yu@gmail.com [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Tang, Song [School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Liu, Hongling, E-mail: hlliu@nju.edu.cn [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Su, Guanyong; Xie, Yuwei [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Giesy, John P. [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China); Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Department of Biology and Chemistry, City University of Hong Kong, Kowloon, Hong Kong Special Administrative Region (Hong Kong); Hecker, Markus [School of Environment and Sustainability, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3 (Canada); Yu, Hongxia [State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing, Jiangsu 210023 (China)

    2015-12-15

    Highlights: • Effects of TBOEP on expression of genes of several nuclear hormone receptors and their relationship with adverse effect pathways in zebrafish. • TBOEP was neither an agonist nor antagonist of AR or AhR as determined by use of in vitro mammalian cell-based receptor transactivation assays. • Modulation of ER- and MR-dependent pathways allowed for development of feasible receptor-mediated, critical mechanisms of toxic action. - Abstract: As one substitute for phased-out brominated flame retardants (BFRs), tris(2-butoxyethyl) phosphate (TBOEP) is frequently detected in aquatic organisms. However, knowledge about endocrine disrupting mechanisms associated with nuclear receptors caused by TBOEP remained restricted to results from in vitro studies with mammalian cells. In the study, results of which are presented here, embryos/larvae of zebrafish (Danio rerio) were exposed to 0.02, 0.1 or 0.5 μM TBOEP to investigate expression of genes under control of several nuclear hormone receptors (estrogen receptors (ERs), androgen receptor (AR), thyroid hormone receptor alpha (TRα), mineralocorticoid receptor (MR), glucocorticoid receptor (GR), aryl hydrocarbon (AhR), peroxisome proliferator-activated receptor alpha (PPARα), and pregnane × receptor (P × R)) pathways at 120 hpf. Exposure to 0.5 μM TBOEP significantly (p < 0.05, one-way analysis of variance) up-regulated expression of estrogen receptors (ERs, er1, er2a, and er2b) genes and ER-associated genes (vtg4, vtg5, pgr, ncor, and ncoa3), indicating TBOEP modulates the ER pathway. In contrast, expression of most genes (mr, 11βhsd, ube2i,and adrb2b) associated with the mineralocorticoid receptor (MR) pathway were significantly down-regulated. Furthermore, in vitro mammalian cell-based (MDA-kb2 and H4IIE-luc) receptor transactivation assays, were also conducted to investigate possible agonistic or antagonistic effects on AR- and AhR-mediated pathways. In mammalian cells, none of these pathways were

  6. β‑catenin nuclear translocation induced by HIF‑1α overexpression leads to the radioresistance of prostate cancer.

    Science.gov (United States)

    Luo, Yong; Li, Mingchuan; Zuo, Xuemei; Basourakos, Spyridon P; Zhang, Jiao; Zhao, Jiahui; Han, Yili; Lin, Yunhua; Wang, Yongxing; Jiang, Yongguang; Lan, Ling

    2018-04-12

    Hypoxia-inducible factor‑1α (HIF‑1α) is known to play crucial roles in tumor radioresistance; however, the molecular mechanisms responsible for the promotion of tumor radioresistance by HIF‑1α remain unclear. β‑catenin is known to be involved in the metastatic potential of prostate cancer (PCa). In this study, to investigate the role of HIF‑1α and β‑catenin in the radioresistance of PCa, two PCa cell lines, LNCaP and C4‑2B, were grouped as follows: Negative control (no treatment), HIF‑1α overexpression group (transfected with HIF‑1α overexpression plasmid) and β‑catenin silenced group (transfected with HIF‑1α plasmids and β‑catenin-shRNA). Cell proliferation, cell cycle, cell invasion and radiosensitivity were examined under normal or hypoxic conditions. In addition, radiosensitivity was examined in two mouse PCa models (the LNCaP orthotopic BALB/c-nu mice model and the C4‑2B subcutaneous SCID mice model). Our results revealed that in both the LNCaP and C4‑2B cells, transfection with HIF‑1α overexpression plasmid led to an enhanced β‑catenin nuclear translocation, while β‑catenin silencing inhibited β‑catenin nuclear translocation. The enhanced β‑catenin nuclear translocation induced by HIF‑1α overexpression resulted in an enhanced cell proliferation and cell invasion, an altered cell cycle distribution, decreased apoptosis, and improved non‑homologous end joining (NHEJ) repair under normal and irradiation conditions. Similar results were observed in the animal models. HIF‑1α overexpression enhanced β‑catenin nuclear translocation, which led to the activation of the β‑catenin/NHEJ signaling pathway and increased cell proliferation, cell invasion and DNA repair. These results thus suggest that HIF‑1α overexpression promotes the radioresistance of PCa cells.

  7. A comparative study of radioresistance of rare and non-disappearing species of tulipal from the flora of Azerbaijan

    International Nuclear Information System (INIS)

    Ibadov, O.V.; Askerov, I.T.; Medzhidov, M.M.; Alekperov, U.K.

    1983-01-01

    Eichle and Sclimid bulds irradiated by gamma-rays of different doses (1-1 Gy) have been investigated. Morphological and phenological observations are carried out, decorative propertie the tested plants are estimated. It is found that low radiation doses (1-5 Gy) stimulate the sprouting of these species, while high doses suppress it. The Eichler species being somewhat more radioresistant than those of Schmi type

  8. Permanent Distal Occlusion of Middle Cerebral Artery in Rat Causes Local Increased ETB, 5-HT1B and AT1 Receptor-Mediated Contractility Downstream of Occlusion

    DEFF Research Database (Denmark)

    Rasmussen, Marianne N P; Hornbak, Malene; Larsen, Stine S

    2013-01-01

    Background/Aims: In response to experimental stroke, a characteristic functional and expressional upregulation of contractile G-protein-coupled receptors has been uncovered in the affected cerebral vasculature; however, the mechanism initiating this phenomenon remains unknown. Methods: Using...... a model of permanent distal occlusion of rat middle cerebral arteries, we investigated whether there was a regional difference in receptor-mediated contractility of segments located upstream and downstream of the occlusion site. The contractile response to endothelin, angiotensin and 5-hydroxytryptamine...... receptor stimulation was studied by sensitive wire myograph. Results: Only downstream segments exhibited an augmented contractile response to stimulation with each of the three ligands, with the response towards sarafotoxin 6c being especially augmented compared to sham, upstream and contralateral controls...

  9. Down-regulation of survivin by oxaliplatin diminishes radioresistance of head and neck squamous carcinoma cells

    International Nuclear Information System (INIS)

    Khan, Zakir; Khan, Noor; Tiwari, Ram P.; Patro, Ishan K.; Prasad, G.B.K.S.; Bisen, Prakash S.

    2010-01-01

    Background: Oxaliplatin is integrated in treatment strategies against a variety of cancers including radiation protocols. Herein, as a new strategy we tested feasibility and rationale of oxaliplatin in combination with radiation to control proliferation of head and neck squamous cell carcinoma (HNSCC) cells and discussed survivin-related signaling and apoptosis induction. Methods: Cytotoxicity and apoptosis induced by radiation and/or oxaliplatin were examined in relation to survivin status using two HNSCC cell lines viz., Cal27 and NT8e, and one normal 293-cell line. Survivin gene knockdown by siRNA was also tested in relevance to oxaliplatin-mediated radiosensitization effects. Results: Survivin plays a critical role in mediating radiation-resistance in part through suppression of apoptosis via a caspase-dependent mechanism. Oxaliplatin treatment significantly decreased expression of survivin in cancer cells within 24-72 h. Apoptotic cells and caspase-3 activity were increased parallely with decrease in cell viability, if irradiated during this sensitive period. The cytotoxicity of oxaliplatin and radiation combination was greater than additive. Survivin gene knockdown experiments have demonstrated the role of survivin in radiosensitization of cancer cells mediated by oxaliplatin. Conclusions: Higher expression of survivin is a critical factor for radioresistance in HNSCC cell lines. Pre-treatment of cancer cells with oxaliplatin significantly increased the radiosensitivity through induction of apoptosis by potently inhibiting survivin.

  10. Novel pattern of post-γ ray de novo DNA synthesis in a radioresistant human strain

    International Nuclear Information System (INIS)

    Mirzayans, R.; Gentner, N.E.; Paterson, M.C.

    1985-01-01

    Enhanced resistance to radiation cytotoxicity in a fibroblast strain from an afflicted member of a Li-Fraumeni syndrome family may be largely ascribable to a change in the pattern of DNA replicative synthesis following γ ray exposure. That is, the extent of the initial radiogenic inhibition of replicative synthesis and the time interval before its subsequent recovery were both found to be greater in radioresistant (RR) compared to normal cells. In addition, the post-recovery replication rates in the RR cells were both higher and longer lasting than those in the control cells. A similar differential pattern was also seen following treatment with 4NQO, another DNA-damaging agent to which this RR strain displays enhanced resistance. Moreover, several conventional DNA repair assays indicated that the RR cells repair radiogenic damage at normal rates. The authors therefore suggest that the increased inhibition and prolonged lag in resumption of replicative synthesis seen in the RR strain upon exposure to certain genotoxic agents may enhance cellular recovery by ''buying additional time'' for processing of potentially lethal lesions

  11. Effect of individual and group housing of mice on the level of radioresistance

    Directory of Open Access Journals (Sweden)

    Dorozhkina O.V.

    2015-12-01

    Full Text Available Aim: to examine the effect of individual and group housing of mice on radioresistance. Material and methods. Effects of individual and group housing of mice on immunity and blood systems were studied on ICR (CD-1 and C57BI6 male mice before and after proton irradiation. Results. Group housing of intact animals resulted in a decline in the number of nucleated cells in the femur bone marrow and thymus mass. The irradiation with proton with energy of 171 MeV at a dose of 1 Gy causes a statistically significant greater reduction of the number of nucleated cells in the femur bone marrow in group-housed mice. A trend toward greater safety of the number of leukocytes in the peripheral blood and higher proliferative activity of bone marrow cells, as well as lower level of aberrant mitoses have been noted in individually-housed mice. Reduction processes in the recovery period of radiation sickness take place at a greater rate in group-housed mice. Conclusion. Group housing of male mice causes increased sensitivity of the blood and immunity systems to the effects of radiation and at the same time accelerates processes of radiation recovery.

  12. Radioresistance of intermediate TCR cells and their localization in the body of mice revealed by irradiation

    International Nuclear Information System (INIS)

    Kimura, Motohiko; Watanabe, Hisami; Ohtsuka, Kazuo; Iiai, Tsuneo; Tsuchida, Masanori; Sato, Shotaro; Abo, Toru

    1993-01-01

    Extrathymic generation of T cells in the liver and in the intestine was recently demonstrated. We investigated herein whether such T cells, especially those in the liver, are present in other organs of mice. This investigation is possible employing our recently introduced method with which even a minor proportion of extrathymic, intermediate T-cell receptor (TCR) cells in organs other than the liver can be identified. Intermediate TCR cells expressed higher levels of IL-2Rβ and lymphocyte function-associated antigen-1 (LFA-1) than bright TCR cells (i.e., T cells of thymic origin) as revealed by two-color staining. Although intermediate TCR cells were present at a small proportion in the spleen and thymus, they predominated in these organs after irradiation (9 Gy) and bone marrow reconstitution, or after low dose irradiation (6 Gy). This was due to that intermediate TCR cells were relatively radioresistant, whereas bright TCR cells were radiosensitive. Microscopic observation and immunochemical staining showed that intermediate TCR cells in the spleen localized in the red pulp and those in the thymus localized in the medulla. These intermediate TCR cells displayed a large light scatter, similar to such cells in the liver. The present results suggest that intermediate TCR cells may proliferate at multiple sites in the body. (author)

  13. Glioblastoma Stem Cells Microenvironment: The Paracrine Roles of the Niche in Drug and Radioresistance

    Directory of Open Access Journals (Sweden)

    Alessia Fidoamore

    2016-01-01

    Full Text Available Among all solid tumors, the high-grade glioma appears to be the most vascularized one. In fact, “microvascular hyperplasia” is a hallmark of GBM. An altered vascular network determines irregular blood flow, so that tumor cells spread rapidly beyond the diffusion distance of oxygen in the tissue, with the consequent formation of hypoxic or anoxic areas, where the bulk of glioblastoma stem cells (GSCs reside. The response to this event is the induction of angiogenesis, a process mediated by hypoxia inducible factors. However, this new capillary network is not efficient in maintaining a proper oxygen supply to the tumor mass, thereby causing an oxygen gradient within the neoplastic zone. This microenvironment helps GSCs to remain in a “quiescent” state preserving their potential to proliferate and differentiate, thus protecting them by the effects of chemo- and radiotherapy. Recent evidences suggest that responses of glioblastoma to standard therapies are determined by the microenvironment of the niche, where the GSCs reside, allowing a variety of mechanisms that contribute to the chemo- and radioresistance, by preserving GSCs. It is, therefore, crucial to investigate the components/factors of the niche in order to formulate new adjuvant therapies rendering more efficiently the gold standard therapies for this neoplasm.

  14. Alterations in transcription factor binding in radioresistant human melanoma cells after ionizing radiation

    International Nuclear Information System (INIS)

    Sahijdak, W.M.; Yang, Chin-Rang; Zuckerman, J.S.; Meyers, M.; Boothman, D.A.

    1994-01-01

    We analyzed alterations in transcription factor binding to specific, known promoter DNA consensus sequences between irradiated and unirradiated radioresistant human melanoma (U1-Mel) cells. The goal of this study was to begin to investigate which transcription factors and DNA-binding sites are responsible for the induction of specific transcripts and proteins after ionizing radiation. Transcription factor binding was observed using DNA band-shift assays and oligonucleotide competition analyses. Confluence-arrested U1-Mel cells were irradiated (4.5 Gy) and harvested at 4 h. Double-stranded oligonucleotides containing known DNA-binding consensus sites for specific transcription factors were used. Increased DNA binding activity after ionizing radiation was noted with oligonucleotides containing the CREB, NF-kB and Sp1 consensus sites. No changes in protein binding to AP-1, AP-2, AP-3, or CTF/NF1, GRE or Oct-1 consensus sequences were noted. X-ray activation of select transcription factors, which bind certain consensus sites in promoters, may cause specific induction or repression of gene transcription. 22 refs., 2 figs

  15. Bimodal cell death induced by high radiation doses in the radioresistant sf9 insect cell line

    International Nuclear Information System (INIS)

    Chandna, S.

    2003-01-01

    Full text: This study was conducted to investigate the mode(s) of cell death induced by high radiation doses in the highly radioresistant Sf9 insect ovarian cell line. Methods: Cells were exposed to γ-radiation doses 200Gy and 500Gy, harvested at various time intervals (6h-72h) following irradiation, and subjected to cell morphology assay, DNA agarose gel electrophoresis, single cell gel electrophoresis (SCGE; comet assay) and Annexin-V labeling for the detection of membrane phosphatidylserine externalization. Cell morphology was assessed in cells entrapped and fixed in agarose gel directly from the cell suspension, thus preventing the possible loss of fragments/ apoptotic bodies. Surviving fraction of Sf9 cells was 0.01 at 200Gy and 98%) undergoing extensive DNA fragmentation at 500Gy, whereas the frequency of cells with DNA fragmentation was considerably less (∼12%) at 200Gy. Conclusions: While the mode of cell death at 200Gy seems to be different from typical apoptosis, a dose of 500Gy induced bimodal cell death, with typical apoptotic as well as the atypical cell death observed at 200Gy

  16. Evidence for induced radioresistance from survival and other end points: An introduction

    International Nuclear Information System (INIS)

    Joiner, M.C.

    1994-01-01

    A substantial body of data published during the past 30 years makes a strong case for the existence of cellular radioprotective mechanisms that can be up-regulated in response to exposure to small doses of ionizing radiation. Either these open-quotes inducedclose quotes mechanisms can protect against a subsequent exposure to radiation that may be substantially larger than the initial open-quotes primingclose quotes or open-quotes conditioningclose quotes dose, or they may influence the shape of the survival response to single doses so that small radiation exposures are more effective per unit dose than larger exposures above a threshold where the induced radioprotection is triggered. Evidence for these effects comes from studies in vitro with protozoa, algae, higher plant cells, insect cells, mammalian and human cells, and studies on animal models in vivo. Work at the molecular level is now confirming that changes in levels of some cytoplasmic and nuclear proteins, and the increased expression of some genes, may occur within a few hours or even minutes of irradiation. This would be sufficiently quick to explain the phenomenon of induced radioresistance although the precise mechanism, whether by repair, cell cycle control or some other process, remains yet undefined. 35 refs

  17. Radioresistance of populations of bank voles Clethrionomys glareolus in radionuclide-contaminated areas

    International Nuclear Information System (INIS)

    Il'enko, A.I.; Krapivko, T.P.

    1994-01-01

    Contamination of extended territories with radionuclides renders the monitoring of natural populations in their habitats an important work to be done in order to determine the directions of evolution caused by long-term exposure to ionizing radiation. In view of this, many years of field and experimental radioecological studies were devoted to animal populations that inhabit the territories contaminated with 137 Cs after the Chernobyl Power Plant disaster. Special emphasis was placed on the investigation of the time course of radiosensitivity of mammalian populations over several generations as a general index of adaptive processes developing in an area with an elevated radiation background. The authors monitored the population of the European bank vole, a species known for its high spontaneous resistance to radiation. In optimal environmental conditions, it has LD 50/30 = 9.7 Gy. The reaction of bank vole populations to radioactive contamination of their environment primarily increases the rate of variation of sensitivity to ionizing radiation. This results in a continuous increase in the radioresistance of the populations to a certains table level. The findings suggest that adaptive processes occur in natural mammalian populations subjected to chronic ionizing irradiation

  18. Microbial radio-resistance of Salmonella Typhimurium in egg increases due to repetitive irradiation with electron beam

    Energy Technology Data Exchange (ETDEWEB)

    Tesfai, Adiam T.; Beamer, Sarah K.; Matak, Kristen E. [West Virginia University, Division of Animal and Nutritional Sciences, PO Box 6108, Morgantown, WV 26508 (United States); Jaczynski, Jacek, E-mail: Jacek.Jaczynski@mail.wvu.ed [West Virginia University, Division of Animal and Nutritional Sciences, PO Box 6108, Morgantown, WV 26508 (United States)

    2011-04-15

    Ionizing radiation improves food safety. However, foodborne pathogens develop increased resistance in response to sub-lethal stresses such as heat, pH, antibiotics, etc. Therefore, it is hypothesized that foodborne pathogens may develop increased radio-resistance to electron beam (e-beam) radiation. The objective was to determine if D{sub 10}-value for Salmonella Typhimurium in de-shelled raw egg (egg white and yolk mixed together) increases due to repetitive processing with e-beam at sub-lethal doses. Survivors were enumerated on non-selective (TSA) and selective (XLD) media. Survivors from the highest dose were isolated and used in subsequent e-beam cycle. This process was repeated four times for a total of five e-beam cycles. D{sub 10}-values for S. Typhimurium enumerated on TSA and XLD following each e-beam cycle were calculated as inverse reciprocal of the slope of survivor curves. D{sub 10}-values for the ATCC strain were 0.59{+-}0.031 and 0.46{+-}0.022 kGy on TSA and XLD, respectively. However, following the fifth e-beam cycle, the respective D{sub 10}-values increased (P<0.05) to 0.69{+-}0.026 and 0.61{+-}0.029 kGy, respectively. S. Typhimurium showed a trend (P>0.05) to develop radio-resistance faster on selective media, likely due to facilitated selection of radio-resistant cells within microbial population following each e-beam cycle. For all five e-beam cycles, S. Typhimurium had higher (P<0.05) D{sub 10}-values on non-selective media, indicating that sub-lethal injury followed by cellular repair and recovery are important for radio-resistance and inactivation of this microorganism. This study demonstrated that e-beam efficiently inactivates S. Typhimurium in raw egg; however, similar to other inactivation techniques and factors affecting microbial growth, S. Typhimurium develops increased radio-resistance if repetitively processed with e-beam at sub-lethal doses.

  19. Caveolin-1 mediated radioresistance of 3D grown pancreatic cancer cells

    International Nuclear Information System (INIS)

    Hehlgans, Stephanie; Eke, Iris; Storch, Katja; Haase, Michael; Baretton, Gustavo B.; Cordes, Nils

    2009-01-01

    Background and purpose: Resistance of pancreatic ductal adenocarcinoma (PDAC) to chemo- and radiotherapy is a major obstacle. The integral membrane protein Caveolin-1 (Cav-1) has been suggested as a potent target in human pancreatic carcinoma cells. Materials and methods: Human pancreatic tumor cells were examined in a three-dimensional (3D) cell culture model with regard to clonogenic survival, apoptosis, radiogenic DNA-double strand breaks and protein expression and phosphorylation under siRNA-mediated knockdown of Cav-1 without and in combination with irradiation (X-rays, 0-6 Gy). Immunohistochemistry was used to assess Cav-1 expression in biopsies from patients with PDAC. Results: Tumor cells in PDAC showed significantly higher Cav-1 expression relative to tumor stroma. Cav-1 knockdown significantly reduced β1 integrin expression and Akt phosphorylation, induced Caspase 3- and Caspase 8-dependent apoptosis and enhanced the radiosensitivity of 3D cell cultures. While cell cycling and Cav-1 promoter activity remained stable, Cav-1 knockdown-induced radiosensitization correlated with elevated numbers of residual DNA-double strand breaks. Conclusions: Our data strongly support the concept of Cav-1 as a potent target in pancreatic carcinoma cells due to radiosensitization and Cav-1 overexpression in tumor cells of PDAC. 3D cell cultures are powerful and useful tools for the testing of novel targeting strategies to optimize conventional radio- and chemotherapy regimes for PDAC.

  20. Possible association between stem-like hallmark and radioresistance in human cervical carcinoma cells.

    Science.gov (United States)

    Kumazawa, Shoko; Kajiyama, Hiroaki; Umezu, Tomokazu; Mizuno, Mika; Suzuki, Shiro; Yamamoto, Eiko; Mitsui, Hiroko; Sekiya, Ryuichiro; Shibata, Kiyosumi; Kikkawa, Fumitaka

    2014-05-01

    We aimed to investigate the possibility of an association between a stem-like hallmark and radiotherapeutic sensitivity in human cervical carcinoma cells. Side-population (SP) cells and non-SP (NSP) cells in HeLa cells were isolated using flow cytometry and Hoechst 33342 efflux. We performed Western blot analysis to evaluate the expression of stem cell markers (CXCR4, Oct3/4, CD133, and SOX2) and apoptosis markers after irradiation. In addition, SP and NSP cells were injected into nude mice and we assessed subcutaneous tumor formation. To examine tolerance of irradiation, colony formation and apoptosis change were confirmed in the SP and NSP cells. SP cells showed a higher expression of CXCR4, Oct3/4, CD133, and SOX2 than NSP cells. The colony size of SP cells cultured on non-coated dishes was larger than that of NSP cells, and NSP cells were easily induced to undergo apoptosis. SP cells tended to form spheroids and showed a higher level of tumorigenicity compared with NSP cells. In addition, nude mice inoculated with SP cells showed greater tumor growth compared with NSP cells. SP cells showed a higher tumorigenicity and lower apoptotic potential, leading to enhanced radiotolerance. Tumor SP cells showed higher-level stem-cell-like characters and radioresistance than NSP cells. SP cells may be useful for new therapeutic approaches for radiation-resistant cervical cancer. © 2014 The Authors. Journal of Obstetrics and Gynaecology Research © 2014 Japan Society of Obstetrics and Gynecology.

  1. Bcl-2 inhibitors potentiate the cytotoxic effects of radiation in Bcl-2 overexpressing radioresistant tumor cells

    International Nuclear Information System (INIS)

    Hara, Takamitsu; Omura-Minamisawa, Motoko; Chao Cheng; Nakagami, Yoshihiro; Ito, Megumi; Inoue, Tomio

    2005-01-01

    Purpose: Bcl-2, an inhibitor of apoptosis frequently shows elevated expression in human tumors, thus resulting in resistance to radiation therapy. Therefore, inhibiting Bcl-2 function may enhance the radiosensitivity of tumor cells. Tetrocarcin A (TC-A) and bcl-2 antisense oligonucleotides exhibit antitumor activity by inhibiting Bcl-2 function and transcription, respectively. We investigated whether these antitumor agents would enhance the cytotoxic effects of radiation in tumor cells overexpressing Bcl-2. Methods and materials: We used HeLa/bcl-2 cells, a stable Bcl-2-expressing cell line derived from wild-type HeLa (HeLa/wt) cells. Cells were incubated with TC-A and bcl-2 antisense oligonucleotides for 24 h after irradiation, and cell viability was then determined. Apoptotic cells were quantified by flow cytometric assay. Results: The HeLa/bcl-2 cells were more resistant to radiation than HeLa/wt cells. At concentrations that are not inherently cytotoxic, both TC-A and bcl-2 antisense oligonucleotides increased the cytotoxic effects of radiation in HeLa/bcl-2 cells, but not in HeLa/wt cells. However, in HeLa/bcl-2 cells, additional treatment with TC-A in combination with radiation did not significantly increase apoptosis. Conclusions: The present results suggest that TC-A and bcl-2 antisense oligonucleotides reduce radioresistance of tumor cells overexpressing Bcl-2. Therefore, a combination of radiotherapy and Bcl-2 inhibitors may prove to be a useful therapeutic approach for treating tumors that overexpress Bcl-2

  2. Critical analysis of salvage radical prostatectomy in the management of radioresistant prostate cancer

    Energy Technology Data Exchange (ETDEWEB)

    Seabra, Daniel; Faria, Eliney; Dauster, Breno; Rodrigues, Gunther; Fava, Gilberto [Pio XII Foundation, Barretos, SP (Brazil). Section of Urology], e-mail: daniel.seabra@terra.com.br

    2009-01-15

    Purpose: To critically evaluate salvage radical prostatectomy (SRP) in the treatment of patients with recurrent prostate cancer (PCa). Materials and Methods: From January 2005 to June 2007, we assessed patients with recurrent localized PCa. Recurrence was suspected when there were three or more successive increases in prostate specific antigen (PSA) after nadir. After the routine imagery examinations, and once localized PCa was confirmed, patients were offered SRP. Following surgery, we evaluated bleeding, rectal injury, urinary incontinence or obstruction and impotence. PSA values were measured at 1, 3, 6, months and thereafter twice a year. Results: Forty-two patients underwent SRP. The average age was 61 years. Following radiotherapy , the mean PSA nadir was 1.5 ng/mL (0.57-5.5). The mean prostate specific antigen doubling time (PSA-DT) was 14 months (6-20). Prior to SRP, the mean PSA was 5.7 ng/mL (2.9-18). The pathologic staging was pT2a: 13%; pT2b: 34%; pT2c: 27%; pT3a: 13%; and pT3b: 13%. Bleeding > 600 mL occurred in 14% of the cases; urethral stenosis in 50%; and urinary incontinence (two or more pads/day) in 72%. The mean follow-up post-SRP ranged from 6 to 30 months. The PSA level rose in 9, of which 6 had PSA-DT < 10 months. Conclusions: SRP is a feasible method in the management of localized radioresistant PCa. PSA-DT has shown to be important for the selection and SRP should not be performed if PSA-DT > 10 months. Due to its increased morbidity, SRP should be only offered to the patients who are more concerned about survival rather than quality of life. (author)

  3. Radioresistance of aboriginal earthworms of Absheron peninsula in radioactive model systems

    International Nuclear Information System (INIS)

    Suleymanova, A.S.; Garibov, A.A.; Abdullayev, A.A.; Naghiyev, J.A.; Farajov, M.F.; Samedov, P.A.

    2011-01-01

    Full tex:Soil animals are the most suitable biological indicators of radioactive pollution because they are parts of nutritional chains and webs, occur in relatively high numbers and can be collected during most parts of the year.The use of earthworms in the soils from Ramana iodine plant area of Absheron peninsula, which is rich with radionuclide and in the soil mixed with RaCl2 and UO2SO4 salt solutions in different concentrations, for resistance to ionizing radiation in soil is reviewed.Effect of radionuclides on vital functions of earthworms and determination of radionuclides (before and after experiments) in contaminated soils by ?-spectrometer were carried out in laboratory condition during a month. Regarding to ?-spectrometric results there were determined that earthworms had absorbed most of radioactive elements and allocated them as coprogenous substances on the upper layer of soil. In Ramana soils mostly the 238U radionuclides were highly accumulated in gut cells of the earthworms. By the influence of radioactive elements it was shown that the earthworms from Ramana iodine plant territory variants had proved particularly sensitive to an increased Ra-radiation background and to iodine factor.It was interestingly established the proportional dependence between rising level of accumulation in earthworms' body and in their coprolites and increasing of radioactive salts containing in the soils treated by UO2SO4 and RaCl2 solutions.Thus there is different level of radioresistance for earthworms and they are among the best bioindicators of polluted soils. There was an obvious perspective of using of earthworms as bioremediators in polluted soil with radionuclides in future as well.

  4. Vive la radiorésistance!: converging research in radiobiology and biogerontology to enhance human radioresistance for deep space exploration and colonization

    Science.gov (United States)

    Cortese, Franco; Klokov, Dmitry; Osipov, Andreyan; Stefaniak, Jakub; Moskalev, Alexey; Schastnaya, Jane; Cantor, Charles; Aliper, Alexander; Mamoshina, Polina; Ushakov, Igor; Sapetsky, Alex; Vanhaelen, Quentin; Alchinova, Irina; Karganov, Mikhail; Kovalchuk, Olga; Wilkins, Ruth; Shtemberg, Andrey; Moreels, Marjan; Baatout, Sarah; Izumchenko, Evgeny; de Magalhães, João Pedro; Artemov, Artem V.; Costes, Sylvain V.; Beheshti, Afshin; Mao, Xiao Wen; Pecaut, Michael J.; Kaminskiy, Dmitry; Ozerov, Ivan V.; Scheibye-Knudsen, Morten; Zhavoronkov, Alex

    2018-01-01

    While many efforts have been made to pave the way toward human space colonization, little consideration has been given to the methods of protecting spacefarers against harsh cosmic and local radioactive environments and the high costs associated with protection from the deleterious physiological effects of exposure to high-Linear energy transfer (high-LET) radiation. Herein, we lay the foundations of a roadmap toward enhancing human radioresistance for the purposes of deep space colonization and exploration. We outline future research directions toward the goal of enhancing human radioresistance, including upregulation of endogenous repair and radioprotective mechanisms, possible leeways into gene therapy in order to enhance radioresistance via the translation of exogenous and engineered DNA repair and radioprotective mechanisms, the substitution of organic molecules with fortified isoforms, and methods of slowing metabolic activity while preserving cognitive function. We conclude by presenting the known associations between radioresistance and longevity, and articulating the position that enhancing human radioresistance is likely to extend the healthspan of human spacefarers as well. PMID:29581875

  5. Depression of NMDA receptor-mediated synaptic transmission by four α2 adrenoceptor agonists on the in vitro rat spinal cord preparation

    Science.gov (United States)

    Faber, E S L; Chambers, J P; Evans, R H

    1998-01-01

    α2-Adrenoceptor agonists have a spinal site of analgesic action. In the current study the synaptic depressant actions of xylazine, detomidine, romifidine and dexmedetomidine have been compared on segmental reflexes containing NMDA receptor-mediated components in the neonatal rat hemisected spinal cord preparation in vitro.Reflexes were evoked in the ventral root following either supramaximal electrical stimulation of the corresponding ipsilateral lumbar dorsal root to evoke the high intensity excitatory postsynaptic potential (e.p.s.p.) involving all primary afferent fibres, or low intensity stimulation to evoke the solely A fibre-mediated low intensity e.p.s.p. The high intensity e.p.s.p. contains a greater NMDA receptor-mediated component.Xylazine, romifidine, detomidine and dexmedetomidine all depressed both the high intensity e.p.s.p. and the low intensity e.p.s.p. giving respective EC50 values of 0.91±0.2 μM (n=12), 23.4±3 nM (n=12), 37.7±7 nM (n=8) and 0.84±0.1 nM (n=4) for depression of the high intensity e.p.s.p. and 0.76±0.1 μM (n=12), 22.0±3 nM (n=12), 24.9±6 nM (n=4) and 2.7±0.6 nM (n=4) for depression of the low intensity e.p.s.p., respectively. Unlike the other three drugs, the two values for dexmedetomidine, showing a greater selectivity for the high intensity e.p.s.p., are significantly different.Each of these depressant actions was reversed by the selective α2-adrenoceptor antagonist atipamezole (1 μM).In contrast to previous reports of the actions of α2-adrenoceptor agonists on the in vitro spinal cord preparation, at concentrations ten fold higher than the above EC50 values xylazine, romifidine, detomidine and dexmedetomidine depressed the initial population spike of motoneurons (MSR). This depression was not reversed by atipamezole.Comparison of the rank order of the present EC50 values for depression of the high intensity e.p.s.p. with potency ratios from in vivo analgesic tests in previous studies show a close

  6. Proteomic profiling identifies PTK2/FAK as a driver of radioresistance in HPV-negative head and neck cancer

    Science.gov (United States)

    Skinner, Heath D.; Giri, Uma; Yang, Liang P.; Woo, Sang Hyeok; Story, Michael; Pickering, Curtis; Byers, Lauren; Williams, Michelle; El Naggar, Adel; Wang, Jing; Diao, Lixia; Shen, Li; Fan, You Hong; Molkentine, David; Beadle, Beth; Meyn, Raymond; Myers, Jeffrey; Heymach, John

    2016-01-01

    Purpose Head and neck squamous cell carcinoma (HNSCC) is commonly treated with radiotherapy, and local failure after treatment remains the major cause of disease-related mortality. To date human papillomavirus (HPV) is the only known clinically validated, targetable biomarkers of response to radiation in HNSCC. Experimental Design We performed proteomic and transcriptomic analysis of targetable biomarkers of radioresistance in HPV-negative HNSCC cell lines in vitro, and tested whether pharmacologic blockade of candidate biomarkers sensitized cells to radiotherapy. Candidate biomarkers were then investigated in several independent cohorts of patients with HNSCC. Results Increased expression of several targets was associated with radioresistance, including FGFR, ERK1, EGFR, and focal adhesion kinase (FAK), also known as PTK2. Chemical inhibition of PTK2/FAK, but not FGFR, led to significant radiosensitization with increased G2/M arrest and potentiated DNA damage. PTK2/FAK overexpression was associated with gene amplification in HPV-negative HNSCC cell lines and clinical tumors. In two independent cohorts of patients with locally advanced HPV-negative HNSCC, PTK2/FAK amplification was highly associated with poorer disease-free survival (DFS) (P=0.012 and P=0.034). PTK2/FAK mRNA expression was also associated with worse DFS (P=0.03). Moreover, both PTK2/FAK mRNA (P=0.021) and copy number (P=0.063) were associated with DFS in the Head and Neck Cancer subgroup of The Cancer Genome Atlas. Conclusion Proteomic analysis identified PTK2/FAK overexpression is a biomarker of radioresistance in locally advanced HNSCC, and PTK2/FAK inhibition radiosensitized HNSCC cells. Combinations of PTK2/FAK inhibition with radiotherapy merit further evaluation as a therapeutic strategy for improving local control in HPV-negative HNSCC. PMID:27036135

  7. GABA(A receptor-mediated acceleration of aging-associated memory decline in APP/PS1 mice and its pharmacological treatment by picrotoxin.

    Directory of Open Access Journals (Sweden)

    Yuji Yoshiike

    Full Text Available Advanced age and mutations in the genes encoding amyloid precursor protein (APP and presenilin (PS1 are two serious risk factors for Alzheimer's disease (AD. Finding common pathogenic changes originating from these risks may lead to a new therapeutic strategy. We observed a decline in memory performance and reduction in hippocampal long-term potentiation (LTP in both mature adult (9-15 months transgenic APP/PS1 mice and old (19-25 months non-transgenic (nonTg mice. By contrast, in the presence of bicuculline, a GABA(A receptor antagonist, LTP in adult APP/PS1 mice and old nonTg mice was larger than that in adult nonTg mice. The increased LTP levels in bicuculline-treated slices suggested that GABA(A receptor-mediated inhibition in adult APP/PS1 and old nonTg mice was upregulated. Assuming that enhanced inhibition of LTP mediates memory decline in APP/PS1 mice, we rescued memory deficits in adult APP/PS1 mice by treating them with another GABA(A receptor antagonist, picrotoxin (PTX, at a non-epileptic dose for 10 days. Among the saline vehicle-treated groups, substantially higher levels of synaptic proteins such as GABA(A receptor alpha1 subunit, PSD95, and NR2B were observed in APP/PS1 mice than in nonTg control mice. This difference was insignificant among PTX-treated groups, suggesting that memory decline in APP/PS1 mice may result from changes in synaptic protein levels through homeostatic mechanisms. Several independent studies reported previously in aged rodents both an increased level of GABA(A receptor alpha1 subunit and improvement of cognitive functions by long term GABA(A receptor antagonist treatment. Therefore, reduced LTP linked to enhanced GABA(A receptor-mediated inhibition may be triggered by aging and may be accelerated by familial AD-linked gene products like Abeta and mutant PS1, leading to cognitive decline that is pharmacologically treatable at least at this stage of disease progression in mice.

  8. The acquired radioresistance in HeLa cells under conditions mimicking hypoxia was attenuated by a decreased expression of HIF subunit genes induced by RNA interference

    International Nuclear Information System (INIS)

    Doi, Nobutaka; Ogawa, Ryohei; Cui, Zheng-Guo; Morii, Akihiro; Watanabe, Akihiko; Kanayama, Shinji; Yoneda, Yuko; Kondo, Takashi

    2015-01-01

    The cancer cells residing in the hypoxic layer are resistant to radiation and these are ones responsible for cancer recurrence after radiation therapy. One of the reasons why hypoxic cancer cells acquire radioresistance may be attributable to changes in the gene expression profile by the activation of hypoxia inducible factors (HIFs). However, the details underlying this process remain unknown. In this study, we investigated the effects of knockdown of HIF subunit genes to elucidate how HIF subunit genes may be involved in the radioresistance acquired by HeLa cells following exposure to a hypoxia mimic. Interestingly, HIF-1α and HIF-2α seemed mutually complementary for each other when either of them was suppressed. We thus suppressed the expression of both genes simultaneously. To do this, we developed a short hairpin RNA (shRNA) targeting a high homology region between HIF-1α and HIF-2α. It was shown that the expression of the shRNA effectively suppressed the acquisition of radioresistance following the hypoxia mimic. Moreover, it was confirmed that suppression of both subunits resulted in the downregulation of stem cell markers and the suppression of spheroid formation during the hypoxia mimicking-conditions. This shRNA-mediated knockdown method targeting a common region shared by a family of genes may offer a new candidate cancer treatment. - Highlights: • Incubation with CoCl 2 confers radioresistance to HeLa cells. • Both HIF-1α and HIF-2α are involved in the acquisition of radioresistance. • An shRNA to a homology region of HIF-1α and HIF-2α suppressed the radioresistance. • The shRNA decreased cells with stem cell markers and a stem cell phenotype

  9. The acquired radioresistance in HeLa cells under conditions mimicking hypoxia was attenuated by a decreased expression of HIF subunit genes induced by RNA interference

    Energy Technology Data Exchange (ETDEWEB)

    Doi, Nobutaka [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); New Products Research & Development, Gene Engineering Division, NIPPON GENE Co., Ltd. (Japan); Ogawa, Ryohei, E-mail: ogawa@med.u-toyama.ac.jp [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Cui, Zheng-Guo [Department of Public Health, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama (Japan); Morii, Akihiro; Watanabe, Akihiko [Department of Urology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama (Japan); Kanayama, Shinji; Yoneda, Yuko [New Products Research & Development, Gene Engineering Division, NIPPON GENE Co., Ltd. (Japan); Kondo, Takashi [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan)

    2015-05-01

    The cancer cells residing in the hypoxic layer are resistant to radiation and these are ones responsible for cancer recurrence after radiation therapy. One of the reasons why hypoxic cancer cells acquire radioresistance may be attributable to changes in the gene expression profile by the activation of hypoxia inducible factors (HIFs). However, the details underlying this process remain unknown. In this study, we investigated the effects of knockdown of HIF subunit genes to elucidate how HIF subunit genes may be involved in the radioresistance acquired by HeLa cells following exposure to a hypoxia mimic. Interestingly, HIF-1α and HIF-2α seemed mutually complementary for each other when either of them was suppressed. We thus suppressed the expression of both genes simultaneously. To do this, we developed a short hairpin RNA (shRNA) targeting a high homology region between HIF-1α and HIF-2α. It was shown that the expression of the shRNA effectively suppressed the acquisition of radioresistance following the hypoxia mimic. Moreover, it was confirmed that suppression of both subunits resulted in the downregulation of stem cell markers and the suppression of spheroid formation during the hypoxia mimicking-conditions. This shRNA-mediated knockdown method targeting a common region shared by a family of genes may offer a new candidate cancer treatment. - Highlights: • Incubation with CoCl{sub 2} confers radioresistance to HeLa cells. • Both HIF-1α and HIF-2α are involved in the acquisition of radioresistance. • An shRNA to a homology region of HIF-1α and HIF-2α suppressed the radioresistance. • The shRNA decreased cells with stem cell markers and a stem cell phenotype.

  10. Escitalopram attenuates β-amyloid-induced tau hyperphosphorylation in primary hippocampal neurons through the 5-HT1A receptor mediated Akt/GSK-3β pathway.

    Science.gov (United States)

    Wang, Yan-Juan; Ren, Qing-Guo; Gong, Wei-Gang; Wu, Di; Tang, Xiang; Li, Xiao-Li; Wu, Fang-Fang; Bai, Feng; Xu, Lin; Zhang, Zhi-Jun

    2016-03-22

    Tau hyperphosphorylation is an important pathological feature of Alzheimer's disease (AD). To investigate whether escitalopram could inhibit amyloid-β (Aβ)-induced tau hyperphosphorylation and the underlying mechanisms, we treated the rat primary hippocampal neurons with Aβ1-42 and examined the effect of escitalopram on tau hyperphosphorylation. Results showed that escitalopram decreased Aβ1-42-induced tau hyperphosphorylation. In addition, escitalopram activated the Akt/GSK-3β pathway, and the PI3K inhibitor LY294002 blocked the attenuation of tau hyperphosphorylation induced by escitalopram. Moreover, the 5-HT1A receptor agonist 8-OH-DPAT also activated the Akt/GSK-3β pathway and decreased Aβ1-42-induced tau hyperphosphorylation. Furthermore, the 5-HT1A receptor antagonist WAY-100635 blocked the activation of Akt/GSK-3β pathway and the attenuation of tau hyperphosphorylation induced by escitalopram. Finally, escitalopram improved Aβ1-42 induced impairment of neurite outgrowth and spine density, and reversed Aβ1-42 induced reduction of synaptic proteins. Our results demonstrated that escitalopram attenuated Aβ1-42-induced tau hyperphosphorylation in primary hippocampal neurons through the 5-HT1A receptor mediated Akt/GSK-3β pathway.

  11. Chronic restraint stress causes a delayed increase in responding for palatable food cues during forced abstinence via a dopamine D1-like receptor-mediated mechanism.

    Science.gov (United States)

    Ball, Kevin T; Best, Olivia; Luo, Jonathan; Miller, Leah R

    2017-02-15

    Relapse to unhealthy eating habits in dieters is often triggered by stress. Animal models, moreover, have confirmed a causal role for acute stress in relapse. The role of chronic stress in relapse vulnerability, however, has received relatively little attention. Therefore, in the present study, we used an abstinence-based relapse model in rats to test the hypothesis that exposure to chronic stress increases subsequent relapse vulnerability. Rats were trained to press a lever for highly palatable food reinforcers in daily 3-h sessions and then tested for food seeking (i.e., responding for food associated cues) both before and after an acute or chronic restraint stress procedure (3h/day×1day or 10days, respectively) or control procedure (unstressed). The second food seeking test was conducted either 1day or 7days after the last restraint. Because chronic stress causes dopamine D1-like receptor-mediated alterations in prefrontal cortex (a relapse node), we also assessed dopaminergic involvement by administering either SCH-23390 (10.0μg/kg; i.p.), a dopamine D1-like receptor antagonist, or vehicle prior to daily treatments. Results showed that chronically, but not acutely, stressed rats displayed increased food seeking 7days, but not 1day, after the last restraint. Importantly, SCH-23390 combined with chronic stress reversed this effect. These results suggest that drugs targeting D 1 -like receptors during chronic stress may help to prevent future relapse in dieters. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Role of a guanine nucleotide-binding protein in α1-adrenergic receptor-mediated Ca2+ mobilization in DDT1 MF-2 cells

    International Nuclear Information System (INIS)

    Cornett, L.E.; Norris, J.S.

    1987-01-01

    In this study the mechanisms involved in α 1 -adrenergic receptor-mediated Ca 2+ mobilization at the level of the plasma membrane were investigated. Stimulation of 45 Ca 2+ efflux from saponin-permeabilized DDT 1 MF-2 cells was observed with the addition of either the α 1 -adrenergic agonist phenylephrine and guanosine-5'-triphosphate or the nonhydrolyzable guanine nucleotide guanylyl-imidodiphosphate. In the presence of [ 32 P] NAD, pertussis toxin was found to catalyze ADP-ribosylation of a M/sub r/ = 40,500 (n = 8) peptide in membranes prepared from DDT 1 , MF-2 cells, possibly the α-subunit of N/sub i/. However, stimulation of unidirectional 45 Ca 2+ efflux by phenylephrine was not affected by previous treatment of cells with 100 ng/ml pertussis toxin. These data suggest that the putative guanine nucleotide-binding protein which couples the α 1 -adrenergic receptor to Ca 2+ mobilization in DDT 1 MF-2 cells is not a pertussis toxin substrate and may possibly be an additional member of guanine nucleotide binding protein family

  13. Heavy metals-bioremediation by highly radioresistant Deinococcus radiodurans biofilm prospective use in nuclear reactor decontamination

    International Nuclear Information System (INIS)

    Shukla, Sudhir K.; Subba Rao, T.

    2015-01-01

    Over the past few decades, rapid growth of chemical industries have enhanced the heavy metal contamination in water, thereby raising environmental concerns. In the nuclear power industry, decontamination procedure also generates radioactive heavy metal containing wastes. Radio-resistant Deinococcus radiodurans R1 is reported to be a potential candidate for the treatment of low active waste material. To use any bacterium for bioremediation purpose, knowledge about its biofilm production characteristics is a prerequisite. This is because biofilm-mediated bioremediation processes are more efficient as compared to processes mediated by their planktonic counterparts. However, so far there are no reports on the biofilm producing capability of D. radiodurans. We observed that tagging of D. radiodurans by a plasmid harbouring gfp and kan R conferred significant biofilm producing property to the bacterium. Chemical analysis of biofilm matrix components produced by D. radiodurans showed that the matrix consists primarily of proteins and carbohydrates with small amount of extracellular DNA (eDNA). Further, we studied the effect of Ca 2+ on D. radiodurans biofilm formation and it was observed that D. radiodurans biofilm formation was enhanced at higher concentrations of Ca 2+ . We investigated the capability of D. radiodurans biofilm to remove the heavy metals Co and Ni from synthetic waste streams. Results showed that Ca 2+ enhanced the bioremediation of both heavy metals (Co, Ni) by D. radiodurans biofilms in a highly significant manner. In the presence of 50 mM Ca 2+ 35% Co removal and 25% Ni removal was observed, when compared to biofilm grown in the absence of Ca 2+ , which showed mere 7% Co and 3% Ni removal, respectively. The results showed that the presence of Ca 2+ significantly enhanced exopolysaccharide and eDNA (both negatively charged) production in the biofilm matrix. This indicated adsorption could be the major mechanism behind enhanced biofilm mediated removal

  14. Identification of CD133-positive radioresistant cells in atypical teratoid/rhabdoid tumor.

    Directory of Open Access Journals (Sweden)

    Shih-Hwa Chiou

    2008-05-01

    Full Text Available Atypical teratoid/rhabdoid tumor (AT/RT is an extremely malignant neoplasm in the central nervous system (CNS which occurs in infancy and childhood. Recent studies suggested that CD133 could be considered a marker for brain cancer stem-like cells (CSCs. However, the role of CD133 in AT/RT has never been investigated. Herein we report the isolation of CD133-positive cells (CD133(+, found to have the potential to differentiate into three germ layer tissues, from tissues of nine AT/RT patients. The migration/invasion/malignancy and radioresistant capabilities of CD133(+ were significantly augmented when compared to CD133(-. The clinical data showed that the amount of CD133(+ in AT/RTs correlated positively with the degree of resistance to radiation therapy. Using cDNA microarray analysis, the genotoxic-response profiles of CD133(+ and CD133(- irradiated with 10 Gy ionizing radiation (IR were analyzed 0.5, 2, 6, 12 and 24 h post-IR. We then validated these microarray data and showed increased phosphorylation after IR of p-ATM, p-RAD17, and p-CHX2 as well as increased expression of BCL-2 protein in CD133(+ compared to CD133(-. Furthermore, we found that CD133(+ can effectively resist IR with cisplatin- and/or TRAIL-induced apoptosis. Immunohistochemical analysis confirmed the up-regulated expression of p-ATM and BCL-2 proteins in IR-treated CD133(+ xenotransgrafts in SCID mice but not in IR-treated CD133(-. Importantly, the effect of IR in CD133(+ transplanted mice can be significantly improved by a combination of BCL-2 siRNA with debromohymenialdisine, an inhibitor of checkpoint kinases. In sum, this is the first report indicating that CD133(+ AT/RT cells demonstrate the characteristics of CSCs. The IR-resistant and anti-apoptotic properties in CD133(+ may reflect the clinical refractory malignancy of AT/RTs and thus the activated p-ATM pathway and BCL-2 expression in CD133(+ could be possible targets to improve future treatment of deadly diseases

  15. Melatonin suppresses thyroid cancer growth and overcomes radioresistance via inhibition of p65 phosphorylation and induction of ROS

    Directory of Open Access Journals (Sweden)

    Zhen-Wei Zou

    2018-06-01

    Full Text Available Thyroid cancer is the most common endocrine carcinoma with increasing incidence worldwide and anaplastic subtypes are frequently associated with cancer related death. Radioresistance of thyroid cancer often leads to therapy failure and cancer-related death. In this study, we found that melatonin showed potent suppressive roles on NF-κB signaling via inhibition of p65 phosphorylation and generated redox stress in thyroid cancer including the anaplastic subtypes. Our data showed that melatonin significantly decreased cell viability, suppressed cell migration and induced apoptosis in thyroid cancer cell lines in vitro and impaired tumor growth in the subcutaneous mouse model in vivo. By contrast, irradiation of thyroid cancer cells resulted in elevated level of phosphorylated p65, which could be reversed by cotreatment with melatonin. Consequently, melatonin synergized with irradiation to induce cytotoxicity to thyroid cancer, especially in the undifferentiated subgroups. Taken together, our results suggest that melatonin may exert anti-tumor activities against thyroid carcinoma by inhibition of p65 phosphorylation and induction of reactive oxygen species. Radio-sensitization by melatonin may have clinical benefits in thyroid cancer. Keywords: Melatonin, Thyroid cancer, Radioresistance, p65, Reactive oxygen species

  16. Cytogenetic Effects of Low Dose Radiation in Mammalian Cells Analysis of the Phenomenon Hypersensitivity and Induced Radioresistence

    CERN Document Server

    Shmakova, N L; Nasonova, E A; Krasavin, E A; Rsjanina, A V

    2001-01-01

    The induction of cytogenetic damage after irradiation of chinese hamster cells and human melanoma cells within dose range 1-200 cGy was studied. The anaphase and metaphase analysis of chromosome damage and micronuclei test were applied. The hypersensitivity (HRS) at doses below 20 cGy and the increased radioresistence at higher doses (IR) were shown with all cytogenetic criteria for both cell lines. The phenomenon of HRS/IR was reproduced in synchronic as well as in asynchronic population of chinese hamster cells. This fact shows that HRS was caused by high radiosensitivity of all cells and can not be explained by any differential sensitivity of cells in different phases of the cell cycle. So it was supposed that the increasing radioresistence is determined by the inclusion of the inducible repair processes in all cells. This conclusion agress with the fact that there was no evidence of HRS on dose-effect curves and that some part of pre-existent damage was repaired after preliminary irradiation with low dose...

  17. The role of oxidative stress in EBV lytic reactivation, radioresistance and the potential preventive and therapeutic implications.

    Science.gov (United States)

    Hu, Jianmin; Li, Hongde; Luo, Xiangjian; Li, Yueshuo; Bode, Ann; Cao, Ya

    2017-11-01

    Epstein-Barr virus (EBV) is an important cancer causing virus. Cancer associated with EBV account for approximately 1.5% of all cancers, and represent 1.8% of all cancer deaths worldwide. EBV reactivation plays an important role in the development of EBV-related diseases and is closely related with patients' survival and clinical stages of EBV-related cancers. The therapy regarding to EBV-related cancers is very urgent, especially in endemic areas. Generating oxidative stress is a critical mechanism by which host cells defend against infection by virus. In addition, ROS-mediated oxidative stress plays a significant but paradoxical role acting as a "double-edged sword" to regulate cellular response to radiation, which is the main therapy strategy for EBV-related cancers, especially nasopharyngeal carcinoma. Therefore, in this review we primarily discuss the possible interplay among the oxidative stress, EBV lytic reactivation and radioresistance. Understanding the role of oxidative stress in EBV lytic reactivation and radioresistance will assist in the development of effective strategies for prevention and treatment of EBV-related cancers. © 2017 UICC.

  18. Intrathymic radioresistant stem cells follow an IL-2/IL-2R pathway during thymic regeneration after sublethal irradiation

    International Nuclear Information System (INIS)

    Zuniga-Pfluecker, J.C.K.; Kruisbeek, A.M.

    1990-01-01

    Sublethally irradiated mice undergo thymic regeneration which follows a phenotypic pattern of events similar to that observed during normal fetal development. Thymic regeneration after irradiation is the product of a limited pool of intrathymic radioresistant stem cells undergoing simultaneous differentiation. We show that in this model of T cell development, thymic regeneration follows a pathway in which the IL-2R is transiently expressed on CD4-/CD8- cells. IL-2R expression occurred during the exponential growth period of thymic regeneration, and IL-2R blocking prevented this explosive growth. Flow cytometry analysis revealed that the IL-2R blockade affected primarily the development of the immature CD3-/CD4-/CD8- (triple negative) cells and their ability to generate CD3+/CD4+/CD8+ or CD3+/CD4+/CD8- and CD3+/CD4-/CD8+ thymocytes. Thus, our findings demonstrate that blocking of the IL-2R resulted in an arrest in proliferation and differentiation by intrathymic radioresistant stem cells, indicating that the IL-2/IL-2R pathway is necessary for the expansion of immature triple negative T cells

  19. Metabolic profiling and correlation analysis for the determination of killer compounds of proliferating and clonogenic HRT-18 colon cancer cells from Lafoensia pacari.

    Science.gov (United States)

    Reichert, Cristiane Loiva; da Silva, Denise Brentan; Carollo, Carlos Alexandre; Weffort-Santos, Almeriane Maria; de Moraes Santos, Cid Aimbiré

    2018-06-18

    Lafoensia pacari A. St.-Hil., belonging to the family Lythraceae and popularly known as 'dedaleira' and 'mangava-brava,' is a native tree of the Brazilian Cerrado, and its barks have been traditionally used as a tonic to treat inflammatory conditions, particularly related to gastric ulcers, wounds or fevers and various types of cancer. We have previously demonstrated the apoptogenic effects of the methanolic extract of L. pacari using various cancer cell lines. In the present study, this extract has been partitioned into fractions to identify the components that might be responsible for the apoptogenic effects using HRT-18 cells, which have been previously demonstrated to be sensitive to this extract. A standard methanolic extract was prepared and fractionated by centrifugal partition chromatography. The fractions were submitted to cytotoxicity and clonogenic assays to monitor the effects in parallel with LC-DAD-MS and statistical analyses to suggest the potential bioactive compounds. Besides ellagic acid, the primary constituent of the plant and also the biomarker of the species, one punicalin isomer, three pedunculagin I isomers, two castalagin isomers, three punicalagin HHDP-gallagyl-hexoside isomers, one ellagic acid deoxyhexose conjugate and one methyl ellagic acid deoxyhexose conjugate were putatively identified. The barks of L. pacari are rich in ellagic acid and various hydrolysable tannins, some of which were reported for the first time in this species, such as punicalagin and ellagitannins. This mixture of substances had the ability to kill proliferating cells and abrogate the growth of clonogenic cells in a similar manner shown by the methanolic extract of our previous study. The collective data reported herein suggest that the biological activities of the L. pacari barks used by the Cerrado's population to treat cancer conditions are due to the apoptogenic effects promoted by a mixed content of ellagitannins. Copyright © 2018. Published by Elsevier B.V.

  20. A preliminary investigation into the extent of increased radioresistance or hyper-radiosensitivity in cells of hamster cell lines known to be deficient in DNA repair

    International Nuclear Information System (INIS)

    Skov, K.; Marples, B.; Matthews, J.B.; Zhou, H.; Joiner, M.C.

    1994-01-01

    The response to low doses of X rays was assessed in cells of three hamster cell lines which are defective in DNA repair and was compared with their parental lines. Cells of the V79-derived double-strand break repair-deficient line XR-V15B showed no radioresistance in the 0.5-Gy range compared with the V79B wild type, but instead showed an exponential response. Cells of the single-strand break repair-deficient line EM9 showed hyper-radiosensitivity and exhibited increased radioresistance. Most interestingly, cells of the UV-20 cell line appeared to respond exponentially, as a continuation of the hyper-radiosensitive portion of the curve, with no evidence of increased radioresistance. This line is defective in an incision step of excision repair and is sensitive to crosslinking agents. Further studies are warranted to address the possible role of single- and double-strand break repair and excision repair in hyper-radiosensitivity and increased radioresistance. 24 refs., 4 figs

  1. Lambda bacteriophage gene products and x-ray sensitivity of Escherichia coli: comparison of red-dependent and gam-dependent radioresistance

    International Nuclear Information System (INIS)

    Trgovcevic, Z.; Rupp, W.D.

    1975-01-01

    When gene products of lambda bacteriophage are introduced into a cell by transient induction of a lysogen, increased resistance of the cells to x rays results. This phenomenon has been called phage-induced radioresistance. Genetic studies show at least two classes of induced radioresistance. The first type depends on the products of the lambda red genes and is observed in bacteria that are mutated in the recB gene. It is thought that the lambda red products compensate for the missing RecBC nuclease in the repair of x-ray damage. An optimal effect is obtained even when the lambda red products are supplied 1 h after irradiation. The lesions that are affected by the red-dependent process are probably not deoxyribonucleic acid strand breaks because the extent of deoxyribonucleic acid strand rejoining is not altered by the red products. The second type of phage-induced radioresistance requires the gam product of lambda and is observed in wild-type and polA strains. The lambda gam + gene product must be present immediately after irradiation to exert its full effect. In its presence, DNA breakdown is decreased, and a greater fraction of DNA is converted back to high molecular weight. Strains carrying lex, recA, or certain other combinations of mutations do not show any detectable phage-induced radioresistance. (U.S.)

  2. The influence of nalidixic acid and nicotinamide of the radiation-induced cytogenetic injury to hexaploid wheat varities contrast by radioresistance

    International Nuclear Information System (INIS)

    Selezneva, E.M.; Sarapul'tsev, B.I.

    1990-01-01

    Nalidixic acid modifies the cytogenetic injury when only applied to seeds of a radiosensitive variety, Moskovskaya 35. The radioprotective effect of nicotinamide on both radiosensitive and radioresistant hexaploid wheat varities is observed being dependent on the extent which the genetic apparatus is impaired

  3. Role of insulin-like growth factor-I receptor (IGF-IR) in survival kinetics and radioresistance of mouse embryo fibroblasts in a hypoxic environment

    International Nuclear Information System (INIS)

    Okochi, Kiyoshi

    2002-01-01

    The role of insulin-like growth factor-I receptor (IGF-IR) in survival kinetics and radioresistance of fibroblasts in a severely hypoxic environment (partial oxygen pressure of less than 3 mmHg) was analyzed, in both low and high cell-density conditions. Mouse embryonic fibroblasts R(-), with a targeted disruption of the IGF-IR gene, and R(+) cells, derived from R(-) cells stably transfected with a plasmid containing a human IGF-IR cDNA, were used for this purpose. Survival time in hypoxia was longer in R(+) cells than R(-) cells, which correlated with highly elevated expression of caspase 3-like activity in R(-) cells, but not with HIF-Iα expression. Under euoxia, R(+) cells were more radioresistant, by a factor of 1.9, than R(-) cells. Under hypoxia, R(+) cells became more radioresistant, with an oxygen-enhancement ratio (OER) of 2.7, than R(-) cells, with an OER of 1.5, in a low cell density. However, unexpected hyper-radiosensitivity in hypoxia was observed for both R(+) and R(-) cells in a high cell density, which further increased with incubation time in hypoxia following X-irradiation. The hyper-radiosensitivity was more pronounced for R(-) cells. The result thus implies that IGF-IR may be an important target molecule for radioresistant tumors in radiotherapy. (author)

  4. CB1 receptor-mediated signaling underlies the hippocampal synaptic, learning, and memory deficits following treatment with JWH-081, a new component of spice/K2 preparations.

    Science.gov (United States)

    Basavarajappa, Balapal S; Subbanna, Shivakumar

    2014-02-01

    Recently, synthetic cannabinoids have been sprayed onto plant material, which is subsequently packaged and sold as "Spice" or "K2" to mimic the effects of marijuana. A recent report identified several synthetic additives in samples of "Spice/K2", including JWH-081, a synthetic ligand for the cannabinoid receptor 1 (CB1). The deleterious effects of JWH-081 on brain function are not known, particularly on CB1 signaling, synaptic plasticity, learning and memory. Here, we evaluated the effects of JWH-081 on pCaMKIV, pCREB, and pERK1/2 signaling events followed by long-term potentiation (LTP), hippocampal-dependent learning and memory tasks using CB1 receptor wild-type (WT) and knockout (KO) mice. Acute administration of JWH-081 impaired CaMKIV phosphorylation in a dose-dependent manner, whereas inhibition of CREB phosphorylation in CB1 receptor WT mice was observed only at higher dose of JWH-081 (1.25 mg/kg). JWH-081 at higher dose impaired CaMKIV and CREB phosphorylation in a time-dependent manner in CB1 receptor WT mice but not in KO mice and failed to alter ERK1/2 phosphorylation. In addition, SR treated or CB1 receptor KO mice have a lower pCaMKIV/CaMKIV ratio and higher pCREB/CREB ratio compared with vehicle or WT littermates. In hippocampal slices, JWH-081 impaired LTP in CB1 receptor WT but not in KO littermates. Furthermore, JWH-081 at higher dose impaired object recognition, spontaneous alternation and spatial memory on the Y-maze in CB1 receptor WT mice but not in KO mice. Collectively our findings suggest that deleterious effects of JWH-081 on hippocampal function involves CB1 receptor mediated impairments in CaMKIV and CREB phosphorylation, LTP, learning and memory in mice. © 2013 Wiley Periodicals, Inc.

  5. Ultrastructural evidence for the accumulation of insulin in nuclei of intact 3T3-L1 adipocytes by an insulin-receptor mediated process

    International Nuclear Information System (INIS)

    Smith, R.M.; Jarett, L.

    1987-01-01

    Monomeric ferritin-labeled insulin (F/sub m/-Ins), a biologically active, electron-dense marker of occupied insulin receptors, was used to characterize the internalization of insulin in 3T3-L1 adipocytes. F/sub m/-Ins bound specifically to insulin receptors and was internalized in a time- and temperature-dependent manner. In the nucleus, several F/sub m/-Ins particles usually were found in the same general location-near nuclear pores, associated with the periphery of the condensed chromatin. Addition of a 250-fold excess of unlabeled insulin or incubation at 15 0 C reduced the number of F/sub m/-Ins particles found in nuclei after 90 min by 99% or 92%, respectively. Nuclear accumulation of unlabeled ferritin was only 2% of that found with F/sub m/-Ins after 90 min at 37 0 C. Biochemical experiments utilizing 125 I-labeled insulin and subcellular fractionation indicated that intact 3T3-L1 adipocytes internalized insulin rapidly and that ≅ 3% of the internalized ligand accumulated in nuclei after 1 hr. These data provide biochemical and high-resolution ultrastructural evidence that 3T3-L1 adipocytes accumulate potentially significant amounts of insulin in nuclei by an insulin receptor-mediated process. The transport of insulin or the insulin-receptor complex to nuclei in this cell or in others may be directly involved in the long-term biological effects of insulin - in particular, in the control of DNA and RNA synthesis

  6. Receptor-mediated inhibition of adenylate cyclase and stimulation of arachidonic acid release in 3T3 fibroblasts. Selective susceptibility to islet-activating protein, pertussis toxin

    International Nuclear Information System (INIS)

    Murayama, T.; Ui, M.

    1985-01-01

    Thrombin exhibited diverse effects on mouse 3T3 fibroblasts. It (a) decreased cAMP in the cell suspension, (b) inhibited adenylate cyclase in the Lubrol-permeabilized cell suspension in a GTP-dependent manner, increased releases of (c) arachidonic acid and (d) inositol from the cell monolayer prelabeled with these labeled compounds, (e) increased 45 Ca 2+ uptake into the cell monolayer, and (f) increased 86 Rb + uptake into the cell monolayer in a ouabain-sensitive manner. Most of the effects were reproduced by bradykinin, platelet-activating factor, and angiotensin II. The receptors for these agonists are thus likely to be linked to three separate effector systems: the adenylate cyclase inhibition, the phosphoinositide breakdown leading to Ca 2+ mobilization and phospholipase A2 activation, and the Na,K-ATPase activation. Among the effects of these agonists, (a), (b), (c), and (e) were abolished, but (d) and (f) were not, by prior treatment of the cells with islet-activating protein (IAP), pertussis toxin, which ADP-ribosylates the Mr = 41,000 protein, the alpha-subunit of the inhibitory guanine nucleotide regulatory protein (Ni), thereby abolishing receptor-mediated inhibition of adenylate cyclase. The effects (a), (c), (d), and (e) of thrombin, but not (b), were mimicked by A23187, a calcium ionophore. The effects of A23187, in contrast to those of receptor agonists, were not affected by the treatment of cells with IAP. Thus, the IAP substrate, the alpha-subunit of Ni, or the protein alike, may play an additional role in signal transduction arising from the Ca 2+ -mobilizing receptors, probably mediating process(es) distal to phosphoinositide breakdown and proximal to Ca 2+ gating

  7. Striatal adenosine A2A receptor-mediated positron emission tomographic imaging in 6-hydroxydopamine-lesioned rats using [18F]-MRS5425

    International Nuclear Information System (INIS)

    Bhattacharjee, Abesh Kumar; Lang Lixin; Jacobson, Orit; Shinkre, Bidhan; Ma Ying; Niu Gang; Trenkle, William C.; Jacobson, Kenneth A.; Chen Xiaoyuan; Kiesewetter, Dale O.

    2011-01-01

    Introduction: A 2A receptors are expressed in the basal ganglia, specifically in striatopallidal GABAergic neurons in the striatum (caudate-putamen). This brain region undergoes degeneration of presynaptic dopamine projections and depletion of dopamine in Parkinson's disease. We developed an 18 F-labeled A 2A analog radiotracer ([ 18 F]-MRS5425) for A 2A receptor imaging using positron emission tomography (PET). We hypothesized that this tracer could image A 2A receptor changes in the rat model for Parkinson's disease, which is created following unilateral injection of the monoaminergic toxin 6-hydroxydopamine (6-OHDA) into the substantia nigra. Methods: [ 18 F]-MRS5425 was injected intravenously in anesthetized rats, and PET imaging data were collected. Image-derived percentage injected doses per gram (%ID/g) in regions of interest was measured in the striatum of normal rats and in rats unilaterally lesioned with 6-OHDA after intravenous administration of saline (baseline), D 2 agonist quinpirole (1.0 mg/kg) or D 2 antagonist raclopride (6.0 mg/kg). Results: Baseline %ID/g reached a maximum at 90 s and maintained plateau for 3.5 min, and then declined slowly thereafter. In 6-OHDA-lesioned rats, %ID/g was significantly higher in the lesioned side compared to the intact side, and the baseline total %ID/g (data from both hemispheres were combined) was significantly higher compared to quinpirole stimulation starting from 4.5 min until the end of acquisition at 30 min. Raclopride did not produce any change in uptake compared to baseline or between the hemispheres. Conclusion: Thus, increase of A 2A receptor-mediated uptake of radioactive MRS5425 could be a superior molecular target for Parkinson's imaging.

  8. Striatal adenosine A{sub 2A} receptor-mediated positron emission tomographic imaging in 6-hydroxydopamine-lesioned rats using [{sup 18}F]-MRS5425

    Energy Technology Data Exchange (ETDEWEB)

    Bhattacharjee, Abesh Kumar; Lang Lixin; Jacobson, Orit [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Shinkre, Bidhan [Chemical Biology Unit, Laboratory of Cell Biochemistry and Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 (United States); Ma Ying [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Niu Gang [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Department of Radiology and Imaging Sciences, Warren Grant Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Trenkle, William C. [Chemical Biology Unit, Laboratory of Cell Biochemistry and Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 (United States); Jacobson, Kenneth A. [Molecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 (United States); Chen Xiaoyuan [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States); Kiesewetter, Dale O., E-mail: dk7k@nih.gov [Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, MD 20892 (United States)

    2011-08-15

    Introduction: A{sub 2A} receptors are expressed in the basal ganglia, specifically in striatopallidal GABAergic neurons in the striatum (caudate-putamen). This brain region undergoes degeneration of presynaptic dopamine projections and depletion of dopamine in Parkinson's disease. We developed an {sup 18}F-labeled A{sub 2A} analog radiotracer ([{sup 18}F]-MRS5425) for A{sub 2A} receptor imaging using positron emission tomography (PET). We hypothesized that this tracer could image A{sub 2A} receptor changes in the rat model for Parkinson's disease, which is created following unilateral injection of the monoaminergic toxin 6-hydroxydopamine (6-OHDA) into the substantia nigra. Methods: [{sup 18}F]-MRS5425 was injected intravenously in anesthetized rats, and PET imaging data were collected. Image-derived percentage injected doses per gram (%ID/g) in regions of interest was measured in the striatum of normal rats and in rats unilaterally lesioned with 6-OHDA after intravenous administration of saline (baseline), D{sub 2} agonist quinpirole (1.0 mg/kg) or D{sub 2} antagonist raclopride (6.0 mg/kg). Results: Baseline %ID/g reached a maximum at 90 s and maintained plateau for 3.5 min, and then declined slowly thereafter. In 6-OHDA-lesioned rats, %ID/g was significantly higher in the lesioned side compared to the intact side, and the baseline total %ID/g (data from both hemispheres were combined) was significantly higher compared to quinpirole stimulation starting from 4.5 min until the end of acquisition at 30 min. Raclopride did not produce any change in uptake compared to baseline or between the hemispheres. Conclusion: Thus, increase of A{sub 2A} receptor-mediated uptake of radioactive MRS5425 could be a superior molecular target for Parkinson's imaging.

  9. The role of natural radioresistance and ecological specialization of a specie in radio adaptation (as exemplified by natural rodent populations)

    Energy Technology Data Exchange (ETDEWEB)

    Grigorkina, E.; Olenev, G. [Institute of Plant and Animal Ecology, Ural Branch RAS, Ekaterinburg, (Russian Federation)

    2004-07-01

    The problem of mammal radio-adaptation is closely connected with problems of micro-evolution and prediction of the fate of irradiated populations. This report gives new materials on radio-adaptation of small mammals inhabiting the East Ural Radioactive Trace (EURT) which has been formed after the Kyshtym accident in 1957 year. The EURT zone is a unique area for studying long-term consequences of chronic low-dose irradiation of small mammal populations many generations being born after the accident. The role of natural radioresistance, ecological specialization and biological characteristics of a specie in the development of radio-adaptation are discussed. The objects of investigation were rodents: 1) Ellobius talpinus is a peculiar specialized specie with low ability to migrate, burrowing underground way of life and lifespan up to 6 years; 2) Sylvaemus uralensis, Apodemus agrarius, Clethrionomys rutilus widespread aboveground species, very active migrators with a 1.5 year lifespan. Significant differences were found among species in natural radioresistance to acute gamma-irradiation. LD{sub 50/30} is 5.0{+-}0.7 Gy for the Ellobius talpinus, 7.0{+-}0.4 Gy for the Sylvaemus uralensis, 10.0{+-}0.2 Gy for the Apodemus agrarius, 12.8{+-}0.2 Gy for the Clethrionomys rutilus. Despite the high radiosensitivity the Ellobius talpinus was more tolerant to chronic irradiation (over 45 years inhabiting the EURT, soil pollution by {sup 90}Sr was 950-1050 Ci/km{sup 2} - 35-39 MBq/m{sup 2}) in a complex of morpho-physiological, haematological and immunological parameters, than other species with active migration activity (the initial pollution of soil by {sup 90}Sr was 400-540 Ci/km{sup 2} - 15-20 MBq/m{sup 2}). This phenomenon is explained by radio-adaptation which developed in the Ellobius talpinus due to isolation of their settlement in the periphery of the area in conditions of radio-contamination. Various radioresistance to acute and chronic irradiation, disproportion of

  10. Cancer-initiating cells derived from established cervical cell lines exhibit stem-cell markers and increased radioresistance

    International Nuclear Information System (INIS)

    López, Jacqueline; Poitevin, Adela; Mendoza-Martínez, Veverly; Pérez-Plasencia, Carlos; García-Carrancá, Alejandro

    2012-01-01

    Cancer-initiating cells (CICs) are proposed to be responsible for the generation of metastasis and resistance to therapy. Accumulating evidences indicates CICs are found among different human cancers and cell lines derived from them. Few studies address the characteristics of CICs in cervical cancer. We identify biological features of CICs from four of the best-know human cell lines from uterine cervix tumors. (HeLa, SiHa, Ca Ski, C-4 I). Cells were cultured as spheres under stem-cell conditions. Flow cytometry was used to detect expression of CD34, CD49f and CD133 antigens and Hoechst 33342 staining to identify side population (SP). Magnetic and fluorescence-activated cell sorting was applied to enrich and purify populations used to evaluate tumorigenicity in nude mice. cDNA microarray analysis and in vitro radioresistance assay were carried out under standard conditions. CICs, enriched as spheroids, were capable to generate reproducible tumor phenotypes in nu-nu mice and serial propagation. Injection of 1 × 10 3 dissociated spheroid cells induced tumors in the majority of animals, whereas injection of 1 × 10 5 monolayer cells remained nontumorigenic. Sphere-derived CICs expressed CD49f surface marker. Gene profiling analysis of HeLa and SiHa spheroid cells showed up-regulation of CICs markers characteristic of the female reproductive system. Importantly, epithelial to mesenchymal (EMT) transition-associated markers were found highly expressed in spheroid cells. More importantly, gene expression analysis indicated that genes required for radioresistance were also up-regulated, including components of the double-strand break (DSB) DNA repair machinery and the metabolism of reactive oxygen species (ROS). Dose-dependent radiation assay indicated indeed that CICs-enriched populations exhibit an increased resistance to ionizing radiation (IR). We characterized a self-renewing subpopulation of CICs found among four well known human cancer-derived cell lines (HeLa, Si

  11. MiR-20a Induces Cell Radioresistance by Activating the PTEN/PI3K/Akt Signaling Pathway in Hepatocellular Carcinoma

    International Nuclear Information System (INIS)

    Zhang, Yuqin; Zheng, Lin; Ding, Yi; Li, Qi; Wang, Rong; Liu, Tongxin; Sun, Quanquan; Yang, Hua; Peng, Shunli; Wang, Wei; Chen, Longhua

    2015-01-01

    Purpose: To investigate the role of miR-20a in hepatocellular carcinoma (HCC) cell radioresistance, which may reveal potential strategies to improve treatment. Methods and Materials: The expression of miR-20a and PTEN were detected in HCC cell lines and paired primary tissues by quantitative real-time polymerase chain reaction. Cell radiation combined with colony formation assays was administrated to discover the effect of miR-20a on radiosensitivity. Bioinformatics prediction and luciferase assay were used to identify the target of miR-20a. The phosphatidylinositol 3-kinase inhibitor LY294002 was used to inhibit phosphorylation of Akt, to verify whether miR-20a affects HCC cell radioresistance through activating the PTEN/PI3K/Akt pathway. Results: MiR-20a levels were increased in HCC cell lines and tissues, whereas PTEN was inversely correlated with it. Overexpression of miR-20a in Bel-7402 and SMMC-7721 cells enhances their resistance to the effect of ionizing radiation, and the inhibition of miR-20a in HCCLM3 and QGY-7701 cells sensitizes them to it. PTEN was identified as a direct functional target of miR-20a for the induction of radioresistance. Overexpression of miR-20a activated the PTEN/PI3K/Akt signaling pathway. Additionally, the kinase inhibitor LY294002 could reverse the effect of miR-20a–induced radioresistance. Conclusion: MiR-20a induces HCC cell radioresistance by activating the PTEN/PI3K/Akt pathway, which suggests that miR-20a/PTEN/PI3K/Akt might represent a target of investigation for developing effective therapeutic strategies against HCC

  12. Searching for a strategy to gamma-sterilize Portuguese cork stoppers - preliminary studies on bioburden, radioresistance and sterility assurance level

    International Nuclear Information System (INIS)

    Botelho, M.L.; Almeida-Vara, E.; Andrade, M.E.; Tenreiro, R.

    1988-01-01

    A gamma radiation plant will start running next year in Portugal, to sterilize medical devices and wine cork stoppers. As Portugal is the first world producer, manufacturer and exporter of wine cork stoppers, an efficient sterilizing procedure is required to overcome moulding from long term shipping. Preliminary research on cork stoppers bioburden and microflora radioresistance allowed to establish reliable D 10 and Sterility Assurance Level (SAL) values. Studied samples showed an initial average contamination of about 10 4 c.f.u. per cork stopper. The determined D 10 values for fungi were not higher than 2 kGy. In these conditions, a SAL of 10 -4 can be expected when the product is treated at a minimum absorbed dose of 15 kGy. (author)

  13. Upregulation of Long Noncoding RNA Small Nucleolar RNA Host Gene 18 Promotes Radioresistance of Glioma by Repressing Semaphorin 5A

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Rong [Department of Radiation Oncology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong (China); Department of Radiation Oncology, Fujian Medical University Union Hospital, Fuzhou, Fujian (China); Yao, Qiwei [Department of Radiation Oncology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong (China); Department of Radiation Oncology, Teaching Hospital of Fujian Medical University, Fujian Provincial Cancer Hospital, Fuzhou, Fujian (China); Ren, Chen; Liu, Ying; Yang, Hongli; Xie, Guozhu; Du, Shasha [Department of Radiation Oncology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong (China); Yang, Kaijun [Department of Neurosurgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong (China); Yuan, Yawei, E-mail: yuanyawei2015@outlook.com [Department of Radiation Oncology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong (China); Department of Radiation Oncology, Cancer Hospital Center of Guangzhou Medical University, Guangzhou, Guangdong (China)

    2016-11-15

    Purpose: Although increasing evidence has shown that long noncoding RNAs play an important regulatory role in carcinogenesis and tumor progression, little is known about the role of small nucleolar RNA host gene 18 (SNHG18) in cancer. The goal of this study was to investigate the expression of SNHG18 and its clinical significance in glioma. Methods and Materials: Differences in the lncRNA expression profile between M059K and M059J cells were assessed by lncRNA expression microarray analysis. The expression and localization of SNHG18 in glioma cells or tissues was evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH), respectively. the clinical associations of SNHG18 in glioma was evaluated by qRT-PCR, ISH and immunohistochemistry. The role of SNHG18 in glioma radiosensitivity was evaluated by colony formation assays, immunofluorescence, Western blot and tumor growth inhibition study. Results: The present study investigated the clinical associations of SNHG18 and its role in glioma. Our results showed that the expression of SNHG18 was remarkably upregulated in clinical glioma tissues compared with normal brain tissues. SNHG18 expression was associated with the clinical tumor grade and correlated negatively with isocitrate dehydrogenase 1 mutation. In addition, knockdown of SNHG18 with short hairpin RNA suppressed the radioresistance of glioma cells, and transgenic expression of SNHG18 had the opposite effect. Furthermore, xenograft tumors grown from cells with SNHG18 deletion were more radiosensitive than tumors grown from control cells. Further studies revealed that SNHG18 promotes radioresistance by inhibiting semaphorin 5A and that inhibition of semaphorin 5A expression abrogated the radiosensitizing effect caused by SNHG18 deletion. Conclusions: Our findings provide new insights into the role of SNHG18 in glioma and suggest its potential as a target for glioma therapy.

  14. Pim-3 contributes to radioresistance through regulation of the cell cycle and DNA damage repair in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xiang-Yuan; Wang, Zhen [Cancer Research Institute, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Li, Bei [Department of Nuclear Medicine, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Zhang, Ying-Jian, E-mail: yjzhang111@aliyun.com [Department of Nuclear Medicine, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Li, Ying-Yi, E-mail: liyingyi@fudan.edu.cn [Cancer Research Institute, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China)

    2016-04-22

    Resistance of cancer cells to chemoradiotherapy is a major clinical problem in pancreatic cancer treatment. Therefore, understanding the molecular basis of cellular resistance and identifying novel targets are essential for improving treatment efficacy for pancreatic cancer patients. Previous studies have demonstrated a significant role for Pim-3 in pancreatic cancer survival against gemcitabine-induced genotoxic stress. Here, we observed that radiation treatment enhanced Pim-3 expression in human pancreatic cancer cells in vitro. Stable overexpression of Pim-3 in pancreatic cancer cells significantly protected cells against radiation treatment by attenuating G2/M phase cell cycle arrest and DNA damage response. Silencing of Pim-3 expression significantly elevated the phosphorylation of histone variant H2AX, a marker of DNA double strand breaks, and decreased the activation of ataxia-telangiectasia-mutated (ATM) kinase, along with its downstream targets, eventually enhancing the radiosensitivity of human pancreatic cancer cells in vitro and in vivo. Hence, we demonstrated a novel function for Pim-3 in human pancreatic cancer cell survival against radiation. Targeting Pim-3 may be a promising way to improve treatment efficacy in combination with radiotherapy in human pancreatic cancer. - Highlights: • This is first study to demonstrate that Pim-3 is endogenously induced by ionizing radiation in pancreatic cancer cells, and Pim-3 overexpression enhanced radioresistance of pancreatic cancer cells both in vitro and in vivo. • This is first study to provide evidence that radioresistance induced by Pim-3 is mainly attributed to Pim-3 induces activation of ATM, which subsequently activates checkpoint 1, leading to amplification of DNA repair through cell cycle arrest and DNA repair pathways. • This is first study to indicate that targeting Pim-3 may be a promising strategy to provide better treatment efficacy in combination with radiotherapy in human pancreatic

  15. KNK437, abrogates hypoxia-induced radioresistance by dual targeting of the AKT and HIF-1α survival pathways

    International Nuclear Information System (INIS)

    Oommen, Deepu; Prise, Kevin M.

    2012-01-01

    Highlights: ► KNK437, a benzylidene lactam compound, is a novel radiosensitizer. ► KNK437 inhibits AKT signaling and abrogates the accumulation of HIF-1α under hypoxia. ► KNK437 abrogates hypoxia induced resistance to radiation. -- Abstract: KNK437 is a benzylidene lactam compound known to inhibit stress-induced synthesis of heat shock proteins (HSPs). HSPs promote radioresistance and play a major role in stabilizing hypoxia inducible factor-1α (HIF-1α). HIF-1α is widely responsible for tumor resistance to radiation under hypoxic conditions. We hypothesized that KNK437 sensitizes cancer cells to radiation and overrides hypoxia-induced radioresistance via destabilizing HIF-1α. Treatment of human cancer cells MDA-MB-231 and T98G with KNK437 sensitized them to ionizing radiation (IR). Surprisingly, IR did not induce HSPs in these cell lines. As hypothesized, KNK437 abrogated the accumulation of HIF-1α in hypoxic cells. However, there was no induction of HSPs under hypoxic conditions. Moreover, the proteosome inhibitor MG132 did not restore HIF-1α levels in KNK437-treated cells. This suggested that the absence of HIF-1α in hypoxic cells was not due to the enhanced protein degradation. HIF-1α is mainly regulated at the level of post-transcription and AKT is known to modulate the translation of HIF-1α mRNA. Interestingly, pre-treatment of cells with KNK437 inhibited AKT signaling. Furthermore, down regulation of AKT by siRNA abrogated HIF-1α levels under hypoxia. Interestingly, KNK437 reduced cell survival in hypoxic conditions and inhibited hypoxia-induced resistance to radiation. Taken together, these data suggest that KNK437 is an effective radiosensitizer that targets multiple pro-survival stress response pathways.

  16. Radioresistant head and neck squamous cell carcinoma cells: Intracellular signaling, putative biomarkers for tumor recurrences and possible therapeutic targets

    International Nuclear Information System (INIS)

    Skvortsov, Sergej; Jimenez, Connie R.; Knol, Jaco C.; Eichberger, Paul; Schiestl, Bernhard; Debbage, Paul; Skvortsova, Ira; Lukas, Peter

    2011-01-01

    Purpose: Treatment of local and distant head and neck cancer recurrences after radiotherapy remains an unsolved problem. In order to identify potential targets for use in effective therapy of recurrent tumors, we have investigated protein patterns in radioresistant (FaDu-IRR and SCC25-IRR, “IRR cells”) as compared to parental (FaDu and SCC25) head and neck carcinoma cells. Methods and materials: Radiation resistant IRR cells were derived from parental cells after repeated exposure to ionizing radiation 10 times every two weeks at a single dose of 10 Gy, resulting in a total dose of 100 Gy. Protein profiling in parental and IRR cells was carried out using two-dimensional differential gel electrophoresis (2D-DIGE) followed by MALDI-TOF/TOF mass spectrometry. Cell viability, cell migration assays and Western blot analysis were used to confirm results obtained using the proteome approach. Results: Forty-five proteins that were similarly modulated in FaDu-IRR and SCC25-IRR cells compared to parental cells were selected to analyze their common targets. It was found that these either up- or down-regulated proteins are closely related to the enhancement of cell migration which is regulated by Rac1 protein. Further investigations confirmed that Rac1 is up-regulated in IRR cells, and inhibiting its action reduces the migratory abilities of these cells. Additionally, the Rac1 inhibitor exerts cytostatic effects in HNSCC cells, mostly in migratory cells. Conclusions: Based on these results, we conclude that radioresistant HNSCC cells possess enhanced metastatic abilities that are regulated by a network of migration-related proteins. Rac1 protein may be considered as a putative biomarker of HNSCC radiation resistance, and as a potential therapeutic target for treating local and distant HNSCC recurrences.

  17. Pim-3 contributes to radioresistance through regulation of the cell cycle and DNA damage repair in pancreatic cancer cells

    International Nuclear Information System (INIS)

    Chen, Xiang-Yuan; Wang, Zhen; Li, Bei; Zhang, Ying-Jian; Li, Ying-Yi

    2016-01-01

    Resistance of cancer cells to chemoradiotherapy is a major clinical problem in pancreatic cancer treatment. Therefore, understanding the molecular basis of cellular resistance and identifying novel targets are essential for improving treatment efficacy for pancreatic cancer patients. Previous studies have demonstrated a significant role for Pim-3 in pancreatic cancer survival against gemcitabine-induced genotoxic stress. Here, we observed that radiation treatment enhanced Pim-3 expression in human pancreatic cancer cells in vitro. Stable overexpression of Pim-3 in pancreatic cancer cells significantly protected cells against radiation treatment by attenuating G2/M phase cell cycle arrest and DNA damage response. Silencing of Pim-3 expression significantly elevated the phosphorylation of histone variant H2AX, a marker of DNA double strand breaks, and decreased the activation of ataxia-telangiectasia-mutated (ATM) kinase, along with its downstream targets, eventually enhancing the radiosensitivity of human pancreatic cancer cells in vitro and in vivo. Hence, we demonstrated a novel function for Pim-3 in human pancreatic cancer cell survival against radiation. Targeting Pim-3 may be a promising way to improve treatment efficacy in combination with radiotherapy in human pancreatic cancer. - Highlights: • This is first study to demonstrate that Pim-3 is endogenously induced by ionizing radiation in pancreatic cancer cells, and Pim-3 overexpression enhanced radioresistance of pancreatic cancer cells both in vitro and in vivo. • This is first study to provide evidence that radioresistance induced by Pim-3 is mainly attributed to Pim-3 induces activation of ATM, which subsequently activates checkpoint 1, leading to amplification of DNA repair through cell cycle arrest and DNA repair pathways. • This is first study to indicate that targeting Pim-3 may be a promising strategy to provide better treatment efficacy in combination with radiotherapy in human pancreatic

  18. Suppression of in vitro cell-mediated lympholysis generation by alloactivated lymphocytes. Examination of radioresistant suppressive activity

    International Nuclear Information System (INIS)

    Orosz, C.G.; Ferguson, R.M.

    1986-01-01

    We investigated the radioresistant (1000 rads) suppression of CML generation mediated by alloactivated murine splenocytes. Suppressive cells were generated in MLCs by stimulation of (A X 6R)F1 splenocytes with irradiated C57BL/10 splenocytes. Suppressive cells could lyse targets bearing H-2b alloantigens, but would not lyse parental B10.T(6R) or B10.A targets. Suppressive activity was detected by including the alloactivated (A X 6R)F1 cells in B10.T(6R) anti-B10.A(1R) MLCs. Relative to the suppressive (A X 6R)F1 cells, the B10.A(1R) lymphocytes display both parental and suppressor-inducing alloantigens. In the absence of a suppressive population, B10.A(1R) stimulators cause B10.T(6R) splenocytes to generate cytolytic activity specific for both H-2Db (suppressor-inducing) and H-2Kk (suppressor-borne) target determinants. The irradiated, alloactivated (A X 6R)F1 cells decrease the H-2Db-specific CML generated in this system, thus mediating apparent antigen-specific suppression. However, cytolytic activity concomitantly generated in the same culture against the unrelated H-2Kk target determinants is similarly reduced by the (A X 6R)F1 cells. Thus, radioresistant suppression by alloactivated splenocytes is not necessarily antigen-specific. The irradiated (A X 6R)F1 cells would not suppress the generation of H-2Kk-specific CTL in B10.T(6R) anti-B10.A MLCs. Hence, the irradiated (A X 6R)F1 cells can impede CML generation against third-party alloantigens if, and only if, those alloantigens are coexpressed with suppressor-inducing alloantigens on the stimulator cells in suppressed MLCs. Similar results were also obtained using a different histoincompatible lymphocyte combination

  19. Upregulation of Long Noncoding RNA Small Nucleolar RNA Host Gene 18 Promotes Radioresistance of Glioma by Repressing Semaphorin 5A

    International Nuclear Information System (INIS)

    Zheng, Rong; Yao, Qiwei; Ren, Chen; Liu, Ying; Yang, Hongli; Xie, Guozhu; Du, Shasha; Yang, Kaijun; Yuan, Yawei

    2016-01-01

    Purpose: Although increasing evidence has shown that long noncoding RNAs play an important regulatory role in carcinogenesis and tumor progression, little is known about the role of small nucleolar RNA host gene 18 (SNHG18) in cancer. The goal of this study was to investigate the expression of SNHG18 and its clinical significance in glioma. Methods and Materials: Differences in the lncRNA expression profile between M059K and M059J cells were assessed by lncRNA expression microarray analysis. The expression and localization of SNHG18 in glioma cells or tissues was evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH), respectively. the clinical associations of SNHG18 in glioma was evaluated by qRT-PCR, ISH and immunohistochemistry. The role of SNHG18 in glioma radiosensitivity was evaluated by colony formation assays, immunofluorescence, Western blot and tumor growth inhibition study. Results: The present study investigated the clinical associations of SNHG18 and its role in glioma. Our results showed that the expression of SNHG18 was remarkably upregulated in clinical glioma tissues compared with normal brain tissues. SNHG18 expression was associated with the clinical tumor grade and correlated negatively with isocitrate dehydrogenase 1 mutation. In addition, knockdown of SNHG18 with short hairpin RNA suppressed the radioresistance of glioma cells, and transgenic expression of SNHG18 had the opposite effect. Furthermore, xenograft tumors grown from cells with SNHG18 deletion were more radiosensitive than tumors grown from control cells. Further studies revealed that SNHG18 promotes radioresistance by inhibiting semaphorin 5A and that inhibition of semaphorin 5A expression abrogated the radiosensitizing effect caused by SNHG18 deletion. Conclusions: Our findings provide new insights into the role of SNHG18 in glioma and suggest its potential as a target for glioma therapy.

  20. Monte Carlo model to simulate the effects of DNA damage resulting from accumulation of 125I decays during development of colonies and clonogenic survival assays

    International Nuclear Information System (INIS)

    Lobachevsky, P.; Karagiannis, T.; Martin, R.F.

    1998-01-01

    Full text: Exposure of cultured cells to an internal source of ionising radiation, such as a radioactive isotope, differs substantially from external irradiation in the determination of delivered dose. In some cases, the radioactive isotope cannot be quickly and completely removed from cells before plating for clonogenic survival assay. This provides an additional dose of irradiation which is not easy to calculate. The contribution of this phenomenon to the cell survival is especially important if a radioactive isotope is incorporated into DNA, or a DNA-binding ligand is labelled with the isotope. The correction of the cell survival due to additional dose cannot be calculated using a simple analytical expression, since the isotope is present in the cells during colony growth. We have developed a Monte Carlo model which simulates the process of the colony growth, and takes into account the extent of damage from isotope decays accumulated between consequent cell divisions. The model considers such factors as cell cycle time, radiosensitivity, colony growth inhibition, isotope specific (per cell) activity, partition of isotope in daughter cells, isotope half-life time, isotope efflux. The model allows estimation of the impact of the irradiation during colony formation on the distribution of colony size, and on the calculation of the survival correction factor, which depends mainly on the isotope cell-specific activity. We applied the model to interpret the difference in survival of K652 cells exposed to 125 I decays with various cell-specific activities: 0.45, 3.21 and 7.42 decays/cell/hour. The cells were treated with 125 I - labelled Hoechst 33258 which binds to DNA in cell nucleus. After accumulation of 125 I decays under non-growth conditions, cells were plated for clonogenic survival assay. The survival correction factors calculated from the model for the given values of 125 I cell-specific activity are in good correlation with differences between experimental

  1. P2Y receptor-mediated transient relaxation of rat longitudinal ileum preparations involves phospholipase C activation, intracellular Ca(2+) release and SK channel activation.

    Science.gov (United States)

    Mader, Felix; Krause, Ludwig; Tokay, Tursonjan; Hakenberg, Oliver W; Köhling, Rüdiger; Kirschstein, Timo

    2016-05-01

    Purinergic signaling plays a major role in the enteric nervous system, where it governs gut motility through a number of P2X and P2Y receptors. The aim of this study was to investigate the P2Y receptor-mediated motility in rat longitudinal ileum preparations. Ileum smooth muscle strips were prepared from rats, and fixed in an organ bath. Isometric contraction and relaxation responses of the muscle strips were measured with force transducers. Drugs were applied by adding of stock solutions to the organ bath to yield the individual final concentrations. Application of the non-hydrolyzable P2 receptor agonists α,β-Me-ATP or 2-Me-S-ADP (10, 100 μmol/L) dose-dependently elicited a transient relaxation response followed by a sustained contraction. The relaxation response was largely blocked by SK channel blockers apamin (500 nmol/L) and UCL1684 (10 μmol/L), PLC inhibitor U73122 (100 μmol/L), IP3 receptor blocker 2-APB (100 μmol/L) or sarcoendoplasmic Ca(2+) ATPase inhibitor thapsigargin (1 μmol/L), but not affected by atropine, NO synthase blocker L-NAME or tetrodotoxin. Furthermore, α,β-Me-ATP-induced relaxation was suppressed by P2Y1 receptor antagonist MRS2179 (50 μmol/L) or P2Y13 receptor antagonist MRS2211 (100 μmol/L), and was abolished by co-application of the two antagonists, whereas 2-Me-S-ADP-induced relaxation was abolished by P2Y6 receptor antagonist MRS2578 (50 μmol/L). In addition, P2Y1 receptor antagonist MRS2500 (1 μmol/L) not only abolished α,β-Me-ATP-induced relaxation, but also suppressed 2-Me-S-ADP-induced relaxation. P2Y receptor agonist-induced transient relaxation of rat ileum smooth muscle strips is mediated predominantly by P2Y1 receptor, but also by P2Y6 and P2Y13 receptors, and involves PLC, IP3, Ca(2+) release and SK channel activation, but is independent of acetylcholine and NO release.

  2. Dopamine D2 receptor-mediated G-protein activation in rat striatum: functional autoradiography and influence of unilateral 6-hydroxydopamine lesions of the substantia nigra.

    Science.gov (United States)

    Newman-Tancredi, A; Cussac, D; Brocco, M; Rivet, J M; Chaput, C; Touzard, M; Pasteau, V; Millan, M J

    2001-11-30

    , the present data indicate that, in rat striatum, the actions of quinelorane are mediated primarily by D2 receptors, and suggest that behavioural hypersensitivity to this agonist, induced by unilateral SNPC lesions, is associated with an increase in D2, but not D3 or D4, receptor-mediated G-protein activation.

  3. Does the cell radioresistance acquired by low dose-rate gamma irradiation depend on genetic factors or physiological changes. Study carried out on inactive cells of the unicellular green alga Chlorella pyrenoidosa CHICK

    International Nuclear Information System (INIS)

    Dettwiller, Pascale.

    1982-09-01

    Inactive cells of the unicellular green alga Chlorella pyrenoidosa CHICK were used to test the following hypothesis: the radioresistance acquired by these cells after irradiation at low dose rate (0.06 Gy/mn) is due to the selection or induction of radioresistant clones. Clone cultures were grown mainly from colonies exhibiting defects (high cell loss, slowed growth, pigment deficiency). Of thirty clones studied, three only of second and third separations possessed the radioresistance of their original population. On the basis of these results, backed up by a first experiment which shows the loss of cell radioresistance when continuous irradiation is stopped, the initial hypothesis may be dismissed and research directed towards changes relative to cell restoration processes by irradiation at low dose rates [fr

  4. Circadian rhythms in the incidence of apoptotic cells and number of clonogenic cells in intestinal crypts after radiation using normal and reversed light conditions

    International Nuclear Information System (INIS)

    Ijiri, K.; Potten, C.S.

    1988-01-01

    Variations in the number of radiation-induced morphologically dead or dying cells (apoptotic cells) in the crypts in the small intestine of the mouse have been studied throughout a 24-h period under a normal light regimen. A clear circadian rhythm was displayed in the apoptotic incidence 3 or 6 h after irradiation for each gamma-ray dose studied (range 0.14-9.0 Gy). The most prominent circadian rhythm was obtained after 0.5 Gy. Peak time of day for inducing apoptosis was 06.00-09.00 h, and the trough occurred at 18.00-21.00 h. Some mice were also transferred to a reversed light cycle, and irradiated on different days after transfer. Apoptosis induced by 0.5 Gy or 9.0 Gy, or number of surviving crypts (microcolonies) after 11.0 Gy or 13.0 Gy was examined. The transition point for reversal of circadian rhythm in apoptosis (after 0.5 Gy) occurred 7 days after transfer and the rhythm was reversed by 14 days. The rhythm for crypt survival (i.e. for clonogenic cell radiosensitivity) was disturbed on 1 day and transition point for reversal occurred 3 days after transfer. The rhythm became reversed by 7 days. (author)

  5. Natural Killer Cells Improve Hematopoietic Stem Cell Engraftment by Increasing Stem Cell Clonogenicity In Vitro and in a Humanized Mouse Model.

    Science.gov (United States)

    Escobedo-Cousin, Michelle; Jackson, Nicola; Laza-Briviesca, Raquel; Ariza-McNaughton, Linda; Luevano, Martha; Derniame, Sophie; Querol, Sergio; Blundell, Michael; Thrasher, Adrian; Soria, Bernat; Cooper, Nichola; Bonnet, Dominique; Madrigal, Alejandro; Saudemont, Aurore

    2015-01-01

    Cord blood (CB) is increasingly used as a source of hematopoietic stem cells (HSC) for transplantation. Low incidence and severity of graft-versus-host disease (GvHD) and a robust graft-versus-leukemia (GvL) effect are observed following CB transplantation (CBT). However, its main disadvantages are a limited number of HSC per unit, delayed immune reconstitution and a higher incidence of infection. Unmanipulated grafts contain accessory cells that may facilitate HSC engraftment. Therefore, the effects of accessory cells, particularly natural killer (NK) cells, on human CB HSC (CBSC) functions were assessed in vitro and in vivo. CBSC cultured with autologous CB NK cells showed higher levels of CXCR4 expression, a higher migration index and a higher number of colony forming units (CFU) after short-term and long-term cultures. We found that CBSC secreted CXCL9 following interaction with CB NK cells. In addition, recombinant CXCL9 increased CBSC clonogenicity, recapitulating the effect observed of CB NK cells on CBSC. Moreover, the co-infusion of CBSC with CB NK cells led to a higher level of CBSC engraftment in NSG mouse model. The results presented in this work offer the basis for an alternative approach to enhance HSC engraftment that could improve the outcome of CBT.

  6. Molecular and functional analysis of DIR1; a novel gene with a potential role in induced radioresistance

    International Nuclear Information System (INIS)

    Young, S.M.; McKeen, H.; Valentine, A.; Burke, G.; Hirst, D.; Robson, T.

    2003-01-01

    Full text: There is now little doubt about the existence of radioprotective mechanisms that are upregulated following exposure to small doses of ionizing radiation and other DNA-damaging agents. The identification of genes whose expression is altered following exposure to a low dose of ionizing radiation will be an important step in understanding these phenomena. We have identified a novel gene, DIR1, that is transiently repressed by low radiation doses (Robson et al.,1997 and 1999) and is otherwise expressed in a wide range of cell lines and tissues. The repression of this gene is in the dose range where induced radioresistance is observed in a number of cell survival studies (Joiner et al., 2001) implicating this gene in induced radioresistance. Using antisense strategies, we have demonstrated that the DIR1 gene product appears to be involved in cell survival and DNA repair in a range of cell lines following exposure to X-rays (Robson et al., 1999 and 2000). Using microchip array analysis we have been able to identify a number of genes activated as a consequence of DIR1 repression. Preliminary data implicate genes involved in repair, cell cycle and stress response and include ATM and BRCA2. We are now confirming these responses using northern and western blot analysis. Yeast two hybrid analysis has also been useful in demonstrating interacting proteins. One protein, which interacts with DIR1 is similar to murine UIP28, a RING finger protein which interacts with the ubiquitin conjugating enzyme, UbcM4. Interestingly, the ubiquitin (Ub)/proteosome pathway regulates many cellular processes including apoptosis, cell cycle progression, stress responses, development and transcriptional regulation. Further characterisation of these downstream genes and interacting proteins will allow us to:- i) dissect the cellular pathways involved in adaptation to oxidative and genotoxic stress ii) elucidate the mechanisms involved in many disease pathologies iii) identify new

  7. Decursin reduce radio-resistance of hypoxic regions under the proton beam therapy by induced HIF-1α degradation

    International Nuclear Information System (INIS)

    Jung, Myung Hwan; Kim, Kye Ryung

    2013-01-01

    Protons induce cancer-cell apoptosis in vitro and block blood vessel formation in vivo through the generation of reactive oxygen species (ROS). The fact that proton severely inhibits blood vessel development in zebrafish embryos suggests a higher sensitivity of vascular endothelial cells to proton beam. Decursin, a coumarin compound, was originally isolated from Angelica gigas Nakai (Dang Gui). A. gigas root has been traditionally used in Korean folk medicine for the treatment of anemia and other common diseases. In previous reports, decursin was reported to exhibit anti-tumor activity against various cancer cells and to inhibit the activities of the androgen and androgen-receptor (AR) signaling pathway in prostate cancer, induction of cell cycle arrest and apoptosis in various cancer cells, such as prostate, breast, bladder, and colon cancer cells. Decursin also inhibits VEGF-induced angiogenesis through the suppression of the VEGFR-2-signaling pathway. However, the mechanism of decursin mediates change of HIF-1α activities is not clear. In this research, we identified regulations of the HIF-1α and the anti-angiogenesis effects of decursin in proton-beam-irradiated human lung cancer, prostate cancer and Hepatic cancer cells. We investigated the underlying mechanisms of positive effects of protonbeam-induced anti-angiogenesis. Our data indicate that the groups co-treated with decursin and a proton-beam had significant reduced HIF-1α activity compared with the groups treated with only a proton beam under the hypoxic condition caused by DFX(desferrioxamine). Decursin was found to induced HIF-1α degradation. Therefore, we suggest that decursin may be a potential candidate for use as a sensitizer for proton-beaminduced cell apoptosis. Here we have shown that decursin successfully reduced HIF-1α stability under hypoxic condition by induced desferrioxamine. We showed novel candidates for anti-angiogenic compound, decursin, leading to complete inhibition of radio-resistance

  8. Decursin reduce radio-resistance of hypoxic regions under the proton beam therapy by induced HIF-1α degradation

    Energy Technology Data Exchange (ETDEWEB)

    Jung, Myung Hwan; Kim, Kye Ryung [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2013-10-15

    Protons induce cancer-cell apoptosis in vitro and block blood vessel formation in vivo through the generation of reactive oxygen species (ROS). The fact that proton severely inhibits blood vessel development in zebrafish embryos suggests a higher sensitivity of vascular endothelial cells to proton beam. Decursin, a coumarin compound, was originally isolated from Angelica gigas Nakai (Dang Gui). A. gigas root has been traditionally used in Korean folk medicine for the treatment of anemia and other common diseases. In previous reports, decursin was reported to exhibit anti-tumor activity against various cancer cells and to inhibit the activities of the androgen and androgen-receptor (AR) signaling pathway in prostate cancer, induction of cell cycle arrest and apoptosis in various cancer cells, such as prostate, breast, bladder, and colon cancer cells. Decursin also inhibits VEGF-induced angiogenesis through the suppression of the VEGFR-2-signaling pathway. However, the mechanism of decursin mediates change of HIF-1α activities is not clear. In this research, we identified regulations of the HIF-1α and the anti-angiogenesis effects of decursin in proton-beam-irradiated human lung cancer, prostate cancer and Hepatic cancer cells. We investigated the underlying mechanisms of positive effects of protonbeam-induced anti-angiogenesis. Our data indicate that the groups co-treated with decursin and a proton-beam had significant reduced HIF-1α activity compared with the groups treated with only a proton beam under the hypoxic condition caused by DFX(desferrioxamine). Decursin was found to induced HIF-1α degradation. Therefore, we suggest that decursin may be a potential candidate for use as a sensitizer for proton-beaminduced cell apoptosis. Here we have shown that decursin successfully reduced HIF-1α stability under hypoxic condition by induced desferrioxamine. We showed novel candidates for anti-angiogenic compound, decursin, leading to complete inhibition of radio-resistance

  9. High linear-energy-transfer radiation can overcome radioresistance of glioma stem-like cells to low linear-energy-transfer radiation.

    Science.gov (United States)

    Hirota, Yuki; Masunaga, Shin-Ichiro; Kondo, Natsuko; Kawabata, Shinji; Hirakawa, Hirokazu; Yajima, Hirohiko; Fujimori, Akira; Ono, Koji; Kuroiwa, Toshihiko; Miyatake, Shin-Ichi

    2014-01-01

    Ionizing radiation is applied as the standard treatment for glioblastoma multiforme (GBM). However, radiotherapy remains merely palliative, not curative, because of the existence of glioma stem cells (GSCs), which are regarded as highly radioresistant to low linear-energy-transfer (LET) photons. Here we analyzed whether or not high-LET particles can overcome the radioresistance of GSCs. Glioma stem-like cells (GSLCs) were induced from the GBM cell line A172 in stem cell culture medium. The phenotypes of GSLCs and wild-type cells were confirmed using stem cell markers. These cells were irradiated with (60)Co gamma rays or reactor neutron beams. Under neutron-beam irradiation, high-LET proton particles can be produced through elastic scattering or nitrogen capture reaction. Radiosensitivity was assessed by a colony-forming assay, and the DNA double-strand breaks (DSBs) were assessed by a histone gamma-H2AX focus detection assay. In stem cell culture medium, GSLCs could form neurosphere-like cells and express neural stem cell markers (Sox2 and Musashi) abundantly in comparison with their parental cells. GSLCs were significantly more radioresistant to gamma rays than their parental cells, but neutron beams overcame this resistance. There were significantly fewer gamma-H2AX foci in the A172 GSLCs 24 h after irradiation with gamma rays than in their parental cultured cells, while there was no apparent difference following neutron-beam irradiation. High-LET radiation can overcome the radioresistance of GSLCs by producing unrepairable DNA DSBs. High-LET radiation therapy might have the potential to overcome GBM's resistance to X-rays in a clinical setting.

  10. High linear-energy-transfer radiation can overcome radioresistance of glioma stem-like cells to low linear-energy-transfer radiation

    International Nuclear Information System (INIS)

    Hirota, Yuki; Kawabata, Shinji; Kuroiwa, Toshihiko; Miyatake, Shin-ichi; Masunaga, Shin-ichiro; Kondo, Natsuko; Ono, Koji; Hirakawa, Hirokazu; Yajima, Hirohiko; Fujimori, Akira

    2014-01-01

    Ionizing radiation is applied as the standard treatment for glioblastoma multiforme (GBM). However, radiotherapy remains merely palliative, not curative, because of the existence of glioma stem cells (GSCs), which are regarded as highly radioresistant to low linear-energy-transfer (LET) photons. Here we analyzed whether or not high-LET particles can overcome the radioresistance of GSCs. Glioma stem-like cells (GSLCs) were induced from the GBM cell line A172 in stem cell culture medium. The phenotypes of GSLCs and wild-type cells were confirmed using stem cell markers. These cells were irradiated with 60 Co gamma rays or reactor neutron beams. Under neutron-beam irradiation, high-LET proton particles can be produced through elastic scattering or nitrogen capture reaction. Radiosensitivity was assessed by a colony-forming assay, and the DNA double-strand breaks (DSBs) were assessed by a histone gamma-H2AX focus detection assay. In stem cell culture medium, GSLCs could form neurosphere-like cells and express neural stem cell markers (Sox2 and Musashi) abundantly in comparison with their parental cells. GSLCs were significantly more radioresistant to gamma rays than their parental cells, but neutron beams overcame this resistance. There were significantly fewer gamma-H2AX foci in the A172 GSLCs 24 h after irradiation with gamma rays than in their parental cultured cells, while there was no apparent difference following neutron-beam irradiation. High-LET radiation can overcome the radioresistance of GSLCs by producing unrepairable DNA DSBs. High-LET radiation therapy might have the potential to overcome GBM's resistance to X-rays in a clinical setting. (author)

  11. Regulation of radiation-induced protein kinase Cδ activation in radiation-induced apoptosis differs between radiosensitive and radioresistant mouse thymic lymphoma cell lines

    International Nuclear Information System (INIS)

    Nakajima, Tetsuo; Yukawa, Osami; Tsuji, Hideo; Ohyama, Harumi; Wang, Bing; Tatsumi, Kouichi; Hayata, Isamu; Hama-Inaba, Hiroko

    2006-01-01

    Protein kinase Cδ (PKCδ) has an important role in radiation-induced apoptosis. The expression and function of PKCδ in radiation-induced apoptosis were assessed in a radiation-sensitive mouse thymic lymphoma cell line, 3SBH5, and its radioresistant variant, XR223. Rottlerin, a PKCδ-specific inhibitor, completely abolished radiation-induced apoptosis in 3SBH5. Radiation-induced PKCδ activation correlated with the degradation of PKCδ, indicating that PKCδ activation through degradation is involved in radiation-induced apoptosis in radiosensitive 3SBH5. In radioresistant XR223, radiation-induced PKCδ activation was lower than that in radiosensitive 3SBH5. Cytosol PKCδ levels in 3SBH5 decreased markedly after irradiation, while those in XR223 did not. There was no apparent change after irradiation in the membrane fractions of either cell type. In addition, basal cytosol PKCδ levels in XR223 were higher than those in 3SBH5. These results suggest that the radioresistance in XR223 to radiation-induced apoptosis is due to a difference in the regulation of radiation-induced PKCδ activation compared to that of 3SBH5. On the other hand, Atm -/- mouse thymic lymphoma cells were more radioresistant to radiation-induced apoptosis than wild-type mouse thymic lymphoma cells. Irradiated wild-type cells, but not Atm -/- cells, had decreased PKCδ levels, indicating that the Atm protein is involved in radiation-induced apoptosis through the induction of PKCδ degradation. The decreased Atm protein levels induced by treatment with Atm small interfering RNA had no effect on radiation-induced apoptosis in 3SBH5 cells. These results suggest that the regulation of radiation-induced PKCδ activation, which is distinct from the Atm-mediated cascade, determines radiation sensitivity in radiosensitive 3SBH5 cells

  12. HDAC4 and HDAC6 sustain DNA double strand break repair and stem-like phenotype by promoting radioresistance in glioblastoma cells.

    Science.gov (United States)

    Marampon, Francesco; Megiorni, Francesca; Camero, Simona; Crescioli, Clara; McDowell, Heather P; Sferra, Roberta; Vetuschi, Antonella; Pompili, Simona; Ventura, Luca; De Felice, Francesca; Tombolini, Vincenzo; Dominici, Carlo; Maggio, Roberto; Festuccia, Claudio; Gravina, Giovanni Luca

    2017-07-01

    The role of histone deacetylase (HDAC) 4 and 6 in glioblastoma (GBM) radioresistance was investigated. We found that tumor samples from 31 GBM patients, who underwent temozolomide and radiotherapy combined treatment, showed HDAC4 and HDAC6 expression in 93.5% and 96.7% of cases, respectively. Retrospective clinical data analysis demonstrated that high-intensity HDAC4 and/or HDAC6 immunostaining was predictive of poor clinical outcome. In vitro experiments revealed that short hairpin RNA-mediated silencing of HDAC4 or HDAC6 radiosensitized U87MG and U251MG GBM cell lines by promoting DNA double-strand break (DSBs) accumulation and by affecting DSBs repair molecular machinery. We found that HDAC6 knock-down predisposes to radiation therapy-induced U251MG apoptosis- and U87MG autophagy-mediated cell death. HDAC4 silencing promoted radiation therapy-induced senescence, independently by the cellular context. Finally, we showed that p53 WT expression contributed to the radiotherapy lethal effects and that HDAC4 or HDAC6 sustained GBM stem-like radioresistant phenotype. Altogether, these observations suggest that HDAC4 and HDAC6 are guardians of irradiation-induced DNA damages and stemness, thus promoting radioresistance, and may represent potential prognostic markers and therapeutic targets in GBM. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Prognostic factors for outcomes after whole-brain irradiation of brain metastases from relatively radioresistant tumors: a retrospective analysis

    Directory of Open Access Journals (Sweden)

    Schild Steven E

    2010-10-01

    Full Text Available Abstract Background This study investigated potential prognostic factors in patients treated with whole-brain irradiation (WBI alone for brain metastases from relatively radioresistant tumors such as malignant melanoma, renal cell carcinoma, and colorectal cancer. Additionally, a potential benefit from escalating the radiation dose was investigated. Methods Data from 220 patients were retrospectively analyzed for overall survival and local control. Nine potential prognostic factors were evaluated: tumor type, WBI schedule, age, gender, Karnofsky performance score, number of brain metastases, extracerebral metastases, interval from diagnosis of cancer to WBI, and recursive partitioning analysis (RPA class. Results Survival rates at 6 and 12 months were 32% and 19%, respectively. In the multivariate analysis, WBI doses >30 Gy (p = 0.038, KPS ≥70 (p Conclusions Improved outcomes were associated with WBI doses >30 Gy, better performance status, fewer brain metastases, lack of extracerebral metastases, and lower RPA class. Patients receiving WBI alone appear to benefit from WBI doses >30 Gy. However, such a benefit is limited to RPA class 1 or 2 patients.

  14. Natural cytotoxicity in immunodeficiency diseases: preservation of natural killer activity and the in vivo appearance of radioresistant killing

    International Nuclear Information System (INIS)

    Pierce, G.F.; Polmar, S.H.; Schacter, B.Z.; Brovall, C.; Hornick, D.L.; Sorensen, R.U.

    1986-01-01

    We studied spontaneous natural killer (NK) cell activity and radiation-resistant NK mediated cytotoxicity in four patients with clinically documented severe combined immune deficiency disease (SCID), and in one subject each with intestinal lymphangiectasia and cartilage-hair hypoplasia. We observed the preservation of spontaneous NK activity in all patients despite the presence of profound B- and T-lymphocytopenia and clinical immunodeficiency. NK activity was associated with relatively normal circulating numbers of OKM1+ lymphocytes, a population known to contain NK effectors. Spontaneous NK activity resistant to 3000 rad was increased in all patients, indicating the presence of activated natural killer cells in vivo. The concept of a chronically activated immune system in these patients was further supported by the presence of increased Ia positive T cells in all subjects tested, suggesting that radioresistant NK activity may be a useful parameter to measure when assessing in vivo immune activation. Our data, as well as that of others, supports the hypothesis that at least one population of NK cells is a distinct lineage arising at the differentiation level of myeloid and lymphoid stem cells in the bone marrow

  15. Interactions of checkpoint-genes RAD9, RAD17, RAD24 and RAD53 determining radioresistance of Yeast Saccharomyces Cerevisiae

    International Nuclear Information System (INIS)

    Koltovaya, N.A.; Nikulushkina, Yu.V.; Roshchina, M.P.; Devin, A.B.

    2007-01-01

    The mechanisms of genetic control of progress through the division cell cycle (checkpoint-control) in yeast Saccharomyces cerevisiae have been studied intensively. To investigate the role of checkpoint-genes RAD9, RAD17, RAD24, RAD53 in cell radioresistance we have investigated cell sensitivity of double mutants to γ-ray. Double mutants involving various combinations with rad9Δ show epistatic interactions, i.e. the sensitivity of the double mutants to γ-ray was no greater than that of more sensitive of the two single mutants. This suggests that all these genes govern the same pathway. This group of genes was named RAD9-epistasis group. It is interesting to note that the genes RAD9 and RAD53 have positive effect but RAD17 and RAD24 have negative effect on radiosensitivity of yeast cells. Interactions between mutations may differ depending on the agent γ-ray or UV-light, for example mutations rad9Δ and rad24Δ show additive effect for γ-ray and epistatic effect for UV-light

  16. Prenatal diagnosis of ataxia-telangiectasia and Nijmegen Breakage Syndrome by the assay of radioresistant DNA synthesis

    International Nuclear Information System (INIS)

    Kleijer, W.J.; Kraan, M. van der; Los, F.J.; Jaspers, N.G.J.

    1994-01-01

    Prenatal diagnosis was performed in 16 pregnancies at risk of ataxia-telangiectasia (A-T) or Nijmegen Breakage Syndrome (NBS). Radioresistant DNA synthesis (RDS) was investigated in cultured chorionic villus (CV) cells and/or amniotic fluid (AF) cells. In four pregnancies, an affected foetus was diagnosed with increased RDS in cultured CV cells. In three of the four cases confirmation of the diagnosis was obtained by analysis of AF cells and/or skin fibroblasts from the foetus cultured after termination of the pregnancy; in the fourth case a fibroblast culture from the aborted foetus failed. In one case, only AF cells could be analysed in a late stage of pregnancy; pregnancy was terminated due to intermediate/equivocal results but the foetus fibroblasts showed normal RDS. Normal RDS was demonstrated in the other 11 pregnancies at 25% risk either by analysis of CB cells (nine cases) or of AF cells (two cases). In some cases the (normal) results on the CV cells were corroborated by subsequent analysis of Af cells. The results suggest that RDS analysis of CV cells allows reliable prenatal diagnosis of A-T/NBS. However, amniocentesis may be necessary to confirm normal results on CV cells if the foetus is female (because of the risk of maternal cell contamination) or in the rare case of equivocal results. (author)

  17. ATM Is Required for the Prolactin-Induced HSP90-Mediated Increase in Cellular Viability and Clonogenic Growth After DNA Damage.

    Science.gov (United States)

    Karayazi Atici, Ödül; Urbanska, Anna; Gopinathan, Sesha Gopal; Boutillon, Florence; Goffin, Vincent; Shemanko, Carrie S

    2018-02-01

    Prolactin (PRL) acts as a survival factor for breast cancer cells, but the PRL signaling pathway and the mechanism are unknown. Previously, we identified the master chaperone, heat shock protein 90 (HSP90) α, as a prolactin-Janus kinase 2 (JAK2)-signal transducer and activator of transcription 5 (STAT5) target gene involved in survival, and here we investigated the role of HSP90 in the mechanism of PRL-induced viability in response to DNA damage. The ataxia-telangiectasia mutated kinase (ATM) protein plays a critical role in the cellular response to double-strand DNA damage. We observed that PRL increased viability of breast cancer cells treated with doxorubicin or etoposide. The increase in cellular resistance is specific to the PRL receptor, because the PRL receptor antagonist, Δ1-9-G129R-hPRL, prevented the increase in viability. Two different HSP90 inhibitors, 17-allylamino-17-demethoxygeldanamycin and BIIB021, reduced the PRL-mediated increase in cell viability of doxorubicin-treated cells and led to a decrease in JAK2, ATM, and phosphorylated ATM protein levels. Inhibitors of JAK2 (G6) and ATM (KU55933) abolished the PRL-mediated increase in cell viability of DNA-damaged cells, supporting the involvement of each, as well as the crosstalk of ATM with the PRL pathway in the context of DNA damage. Drug synergism was detected between the ATM inhibitor (KU55933) and doxorubicin and between the HSP90 inhibitor (BIIB021) and doxorubicin. Short interfering RNA directed against ATM prevented the PRL-mediated increase in cell survival in two-dimensional cell culture, three-dimensional collagen gel cultures, and clonogenic cell survival, after doxorubicin treatment. Our results indicate that ATM contributes to the PRL-JAK2-STAT5-HSP90 pathway in mediating cellular resistance to DNA-damaging agents. Copyright © 2018 Endocrine Society.

  18. Altered Cross-Linking of HSP27 by Zerumbone as a Novel Strategy for Overcoming HSP27-Mediated Radioresistance

    International Nuclear Information System (INIS)

    Choi, Seo-Hyun; Lee, Yoon-Jin; Seo, Woo Duck; Lee, Hae-June; Nam, Joo-Won; Lee, Yoo Jin; Kim, Joon; Seo, Eun-Kyoung; Lee, Yun-Sil

    2011-01-01

    Purpose: HSP27 or HSP25 negatively regulates apoptosis pathways after radiation or chemotherapeutic agents. Abrogation of HSP27 function may be a candidate target for overcoming radio- and chemoresistance. Methods and Materials: Zerumbone (ZER), a cytotoxic component isolated from Zingiber zerumbet smith. Clonogenic survival assay and flow cytometry after Annexin V staining were performed to determine in vitro sensitization effects of ZER with ionizing radiation. A nude mouse xenografting system was also applied to detect in vivo radiosensitizing effects of ZER. Results: ZER produced cross-linking of HSP27, which was dependent on inhibition of the monomeric form of HSP27. ZER was directly inserted between the disulfide bond in the HSP27 dimer and modified normal HSP27 dimerization. Pretreatment with ZER before radiation inhibited the binding affinity between HSP27 and apoptotic molecules, such as cytochrome c and PKCδ, and induced sensitization in vitro and in an in vivo xenografted nude mouse system. Structural analogs lacking only the carbonyl group in ZER, such as α-humulene (HUM) and 8-hydroxy-humulen (8-OH-HUM), did not affect normal cross-linking of HSP27 and did not induce radiosensitization. Conclusions: We suggest that altered cross-linking of HSP27 by ZER is a good strategy for abolishing HSP27-mediated resistance.

  19. Down-regulation of 5-HT1B and 5-HT1D receptors inhibits proliferation, clonogenicity and invasion of human pancreatic cancer cells.

    Directory of Open Access Journals (Sweden)

    Nilgun Gurbuz

    Full Text Available Pancreatic ductal adenocarcinoma is characterized by extensive local tumor invasion, metastasis and early systemic dissemination. The vast majority of pancreatic cancer (PaCa patients already have metastatic complications at the time of diagnosis, and the death rate of this lethal type of cancer has increased over the past decades. Thus, efforts at identifying novel molecularly targeted therapies are priorities. Recent studies have suggested that serotonin (5-HT contributes to the tumor growth in a variety of cancers including prostate, colon, bladder and liver cancer. However, there is lack of evidence about the impact of 5-HT receptors on promoting pancreatic cancer. Having considered the role of 5-HT-1 receptors, especially 5-HT1B and 5-HT1D subtypes in different types of malignancies, the aim of this study was to investigate the role of 5-HT1B and 5-HT1D receptors in PaCa growth and progression and analyze their potential as cytotoxic targets. We found that knockdown of 5-HT1B and 5-HT1D receptors expression, using specific small interfering RNA (siRNA, induced significant inhibition of proliferation and clonogenicity of PaCa cells. Also, it significantly suppressed PaCa cells invasion and reduced the activity of uPAR/MMP-2 signaling and Integrin/Src/Fak-mediated signaling, as integral tumor cell pathways associated with invasion, migration, adhesion, and proliferation. Moreover, targeting 5-HT1B and 5-HT1D receptors down-regulates zinc finger ZEB1 and Snail proteins, the hallmarks transcription factors regulating epithelial-mesenchymal transition (EMT, concomitantly with up-regulating of claudin-1 and E-Cadherin. In conclusion, our data suggests that 5-HT1B- and 5-HT1D-mediated signaling play an important role in the regulation of the proliferative and invasive phenotype of PaCa. It also highlights the therapeutic potential of targeting of 5-HT1B/1D receptors in the treatment of PaCa, and opens a new avenue for biomarkers identification

  20. Down-regulation of 5-HT1B and 5-HT1D receptors inhibits proliferation, clonogenicity and invasion of human pancreatic cancer cells.

    Directory of Open Access Journals (Sweden)

    Nilgun Gurbuz

    Full Text Available Pancreatic ductal adenocarcinoma is characterized by extensive local tumor invasion, metastasis and early systemic dissemination. The vast majority of pancreatic cancer (PaCa patients already have metastatic complications at the time of diagnosis, and the death rate of this lethal type of cancer has increased over the past decades. Thus, efforts at identifying novel molecularly targeted therapies are priorities. Recent studies have suggested that serotonin (5-HT contributes to the tumor growth in a variety of cancers including prostate, colon, bladder and liver cancer. However, there is lack of evidence about the impact of 5-HT receptors on promoting pancreatic cancer. Having considered the role of 5-HT-1 receptors, especially 5-HT1B and 5-HT1D subtypes in different types of malignancies, the aim of this study was to investigate the role of 5-HT1B and 5-HT1D receptors in PaCa growth and progression and analyze their potential as cytotoxic targets. We found that knockdown of 5-HT1B and 5-HT1D receptors expression, using specific small interfering RNA (siRNA, induced significant inhibition of proliferation and clonogenicity of PaCa cells. Also, it significantly suppressed PaCa cells invasion and reduced the activity of uPAR/MMP-2 signaling and Integrin/Src/Fak-mediated signaling, as integral tumor cell pathways associated with invasion, migration, adhesion, and proliferation. Moreover, targeting 5-HT1B and 5-HT1D receptors down-regulates zinc finger ZEB1 and Snail proteins, the hallmarks transcription factors regulating epithelial-mesenchymal transition (EMT, concomitantly with up-regulating of claudin-1 and E-Cadherin. In conclusion, our data suggests that 5-HT1B- and 5-HT1D- mediated signaling play an important role in the regulation of the proliferative and invasive phenotype of PaCa. It also highlights the therapeutic potential of targeting of 5-HT1B/1D receptors in the treatment of PaCa, and opens a new avenue for biomarkers identification

  1. Elimination of clonogenic tumor cells from HL-60, Daudi, and U-937 cell lines by laser photoradiation therapy: implications for autologous bone marrow purging

    International Nuclear Information System (INIS)

    Gulliya, K.S.; Pervaiz, S.

    1989-01-01

    Laser photoradiation therapy was tested in an in vitro model for its efficacy in the elimination of non-Hodgkin's lymphoma cells. Results show that at 31.2 J/cm2 of laser light in the presence of 20 micrograms/mL of merocyanine 540 (MC540) there was greater than 5 log reduction in Burkitt's lymphoma (Daudi) cells. Similar tumor cell kill was obtained for leukemia (HL-60) cells at a laser light dose of 93.6 J/cm2. However, to obtain the same efficiency of killing for histiocytic lymphoma (U-937) cells, a higher dose of MC540 (25 micrograms/mL) was required. Clonogenic tumor stem cell colony formation was reduced by greater than 5 logs after laser photoradiation therapy. Under identical conditions for each cell line the percent survival for granulocyte-macrophage colony-forming units (CFU-GM, 45.9%, 40%, 17.5%), granulocyte/erythroid/macrophage/megakaryocyte (GEMM, 40.1%, 20.1%, 11.5%), colony-forming units (CFU-C, 16.2%, 9.1%, 1.8%), and erythroid burst-forming units (BFU-E, 33.4%, 17.8%, 3.9%) was significantly higher than the tumor cells. Mixing of gamma ray-irradiated normal marrow cells with tumor cells (1:1 and 10:1 ratio) did not interfere with the elimination of tumor cells. The effect of highly purified recombinant interferon alpha (rIFN) on laser photoradiation therapy of tumor cells was also investigated. In the presence of rIFN (30 to 3,000 U/mL), the viability of leukemic cells was observed to increase from 0% to 1.5% with a concurrent decrease in membrane polarization, suggesting an increase in fluidity of cell membrane in response to rIFN. However, at higher doses of rIFN (6,000 to 12,000 U/mL) this phenomenon was not observed. The viability of lymphoma cells remained unaffected at all doses of rIFN tested

  2. Mitochondrial-Targeted Decyl-Triphenylphosphonium Enhances 2-Deoxy-D-Glucose Mediated Oxidative Stress and Clonogenic Killing of Multiple Myeloma Cells.

    Directory of Open Access Journals (Sweden)

    Jeanine Schibler

    Full Text Available Therapeutic advances have markedly prolonged overall survival in multiple myeloma (MM but the disease currently remains incurable. In a panel of MM cell lines (MM.1S, OPM-2, H929, and U266, using CD138 immunophenotyping, side population staining, and stem cell-related gene expression, we demonstrate the presence of stem-like tumor cells. Hypoxic culture conditions further increased CD138low stem-like cells with upregulated expression of OCT4 and NANOG. Compared to MM cells, these stem-like cells maintained lower steady-state pro-oxidant levels with increased uptake of the fluorescent deoxyglucose analog. In primary human MM samples, increased glycolytic gene expression correlated with poorer overall and event-free survival outcomes. Notably, stem-like cells showed increased mitochondrial mass, rhodamine 123 accumulation, and orthodox mitochondrial configuration while more condensed mitochondria were noted in the CD138high cells. Glycolytic inhibitor 2-deoxyglucose (2-DG induced ER stress as detected by qPCR (BiP, ATF4 and immunoblotting (BiP, CHOP and increased dihydroethidium probe oxidation both CD138low and CD138high cells. Treatment with a mitochondrial-targeting agent decyl-triphenylphosphonium (10-TPP increased intracellular steady-state pro-oxidant levels in stem-like and mature MM cells. Furthermore, 10-TPP mediated increases in mitochondrial oxidant production were suppressed by ectopic expression of manganese superoxide dismutase. Relative to 2-DG or 10-TPP alone, 2-DG plus 10-TPP combination showed increased caspase 3 activation in MM cells with minimal toxicity to the normal hematopoietic progenitor cells. Notably, treatment with polyethylene glycol conjugated catalase significantly reduced 2-DG and/or 10-TPP-induced apoptosis of MM cells. Also, the combination of 2-DG with 10-TPP decreased clonogenic survival of MM cells. Taken together, this study provides a novel strategy of metabolic oxidative stress-induced cytotoxicity of MM

  3. Differential Bystander Signaling Between Radioresistant Chondrosarcoma Cells and Fibroblasts After X-Ray, Proton, Iron Ion and Carbon Ion Exposures

    Energy Technology Data Exchange (ETDEWEB)

    Wakatsuki, Masaru, E-mail: wa@mbe.nifty.com [Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, Chiba (Japan); Department of Radiation Oncology, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts (United States); Magpayo, Nicole; Kawamura, Hidemasa; Held, Kathryn D. [Department of Radiation Oncology, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts (United States)

    2012-09-01

    Purpose: Chondrosarcoma is well known as a radioresistant tumor, but the mechanisms underlying that resistance are still unclear. The bystander effect is well documented in the field of radiation biology. We investigated the bystander response induced by X-rays, protons, carbon ions, and iron ions in chondrosarcoma cells using a transwell insert co-culture system that precludes physical contact between targeted and bystander cells. Methods and Materials: Human chondrosarcoma cells were irradiated with 0.1-, 0.5-, 1-, and 2-Gy X-rays, protons, carbon ions or iron ions using a transwell insert co-culture system. Formation of micronuclei and p53 binding protein 1 staining in bystander and irradiated cells were analyzed and bystander signaling between mixed cultures of chondrosarcoma cells, and normal human skin fibroblasts was investigated. Results: In this study, we show that the fraction of cells with DNA damages in irradiated chondrosarcoma cells showed dose-dependent increases with all beams. However, the fraction of cells with DNA damages in all bystander chondrosarcoma cells did not show any change from the levels in control cells. In the bystander signaling between mixed cultures of chondrosarcoma cells and fibroblasts, the amount of micronucleus formation in all bystander chondrosarcoma cells co-cultured with irradiated fibroblasts were the same as the levels for control cells. However, all bystander fibroblasts co-cultured with irradiated chondrosarcoma cells showed significant increases in the fraction of micronucleated cells compared to the rate of control cells. Conclusions: We conclude that chondrosarcoma cells in the transwell insert co-culture system could release bystander stimulations but could not develop bystander responses.

  4. Differential Bystander Signaling Between Radioresistant Chondrosarcoma Cells and Fibroblasts After X-Ray, Proton, Iron Ion and Carbon Ion Exposures

    International Nuclear Information System (INIS)

    Wakatsuki, Masaru; Magpayo, Nicole; Kawamura, Hidemasa; Held, Kathryn D.

    2012-01-01

    Purpose: Chondrosarcoma is well known as a radioresistant tumor, but the mechanisms underlying that resistance are still unclear. The bystander effect is well documented in the field of radiation biology. We investigated the bystander response induced by X-rays, protons, carbon ions, and iron ions in chondrosarcoma cells using a transwell insert co-culture system that precludes physical contact between targeted and bystander cells. Methods and Materials: Human chondrosarcoma cells were irradiated with 0.1-, 0.5-, 1-, and 2-Gy X-rays, protons, carbon ions or iron ions using a transwell insert co-culture system. Formation of micronuclei and p53 binding protein 1 staining in bystander and irradiated cells were analyzed and bystander signaling between mixed cultures of chondrosarcoma cells, and normal human skin fibroblasts was investigated. Results: In this study, we show that the fraction of cells with DNA damages in irradiated chondrosarcoma cells showed dose-dependent increases with all beams. However, the fraction of cells with DNA damages in all bystander chondrosarcoma cells did not show any change from the levels in control cells. In the bystander signaling between mixed cultures of chondrosarcoma cells and fibroblasts, the amount of micronucleus formation in all bystander chondrosarcoma cells co-cultured with irradiated fibroblasts were the same as the levels for control cells. However, all bystander fibroblasts co-cultured with irradiated chondrosarcoma cells showed significant increases in the fraction of micronucleated cells compared to the rate of control cells. Conclusions: We conclude that chondrosarcoma cells in the transwell insert co-culture system could release bystander stimulations but could not develop bystander responses.

  5. Role of endogeneous thiols in the determination of radio-resistance of E. coli B culture in different phases of its growth

    Energy Technology Data Exchange (ETDEWEB)

    Konstantinova, M M [AN SSSR, Moscow. Inst. Biologii Razvitiya

    1975-05-01

    An investigation was made of the link between the radioresistance of various stages of development of a culture of E. coli B with the level of sulfhydryl groups in the cells at each stage. The analysis was carried out by comparing the content of sulfhydryl groups in the cells, with their viability after irradiation and by determining the effectiveness of radio-protective and radio-sensitizing agents on a culture of various ages. It was shown that the thiols are the factor that determines the variation in radiosensitivity of a culture of E. coli B in the logarithmic and stationary phases of growth.

  6. Radiation-induced phosphorylation of P53 protects radioresistant Spodoptera frugiperda 9 cells by suppressing microRNA-31-Bim-Bax mediated apoptosis

    International Nuclear Information System (INIS)

    Kumar, Ashish; Chandna, Sudhir

    2016-01-01

    In this study, we demonstrate the role of microRNA-31 (miR-31) in the regulation of radiation-induced apoptosis in model radioresistant insect cell line Sf9 (derived from the ovaries of insect Spodoptera frugiperda) which carries well-conserved apoptotic response. We also investigated the miR-31 expression regulation by p53 homologue in these cells. Our initial in silico analysis confirmed perfect conservation of mature miR-31 across various insect orders, hence we designed biotinylated probes from Bombyx mori sequence for successful detection of miR-31 in Sf9 cells

  7. Comparison of SDR presumed by AAMI and the radio-resistance distribution of the microbiological populations exist on the Turkish made single use syringes

    International Nuclear Information System (INIS)

    Konac, T.; Siyakus, G.; Basguel, M.; Uenal, Z.

    2001-01-01

    The concept of radiation sterilization of medical devices was introduced in Turkey 10 years ago through the irradiation facilities in Ankara and Istanbul. It has been preferred by international community due to technological advantages,such as freedom from chemical residues, safer sterility assurance, etc.. This technology is regulated by ISO 11137 (1), EN 552 (2) standards and their references. The scope of this study was to compare the radio-resistance distribution of the microbial population on the Turkish single use syringes and SDR

  8. CREB mediates ICAM-3: inducing radio-resistance, cell growth and migration/invasion of the human nonsmall cell lung cancer cell

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jong Kuk; So, Kwang Sup; Bae, In Hwa; Um, Hong Duck [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2009-05-15

    The ICAM family proteins comprises cell surface molecules that are homologous to NCAM and are members of the single passed type 1 immunoglobulin superfamily (IgSF) that are anchored at the cellular membrane. The ICAM family consists of five subfamilies (ICAM-1 to ICAM-5) of heavily glycosylated cell surface receptors with common functional or structural homology. The extracellular domains of ICAM protein have roles in immune response and inflammation through various cell-cell interactions. The cytoplasmic tail residues of ICAM-3 participate in intracellular signaling such as calcium mobilization and tyrosine phosphorylation. Interestingly, the ICAM proteins appear to have a dual role in cancer. ICAM molecules may target and block tumor progression by stimulation of an immune response such as leukocyte activation. Conversely, other investigations have shown that ICAM molecules are involved in cancer malignancy because their increased expressions are associated with a poor diagnosis, lower survival rates and invasion in several cancers including melanoma, breast cancer and leukemia. We have also reported that an increase of ICAM-3 expression in several cancer cells and specimens of cervical cancer patient induce enhanced radio-resistance by the activation of focal adhesion kinase (FAK) and promote cancer cell proliferation by the activation of Akt and p44/42 MAPK. Therefore, these previous reports imply that ICAM-3 has various undefined roles in cancer. In this study, we investigated whether ICAM-3 increase cell migration and invasion through CREB activation and CREB has a role of increase of radioresistance and cell growth.

  9. Radioresistance of inorganic glasses

    International Nuclear Information System (INIS)

    Vorob'ev, A.A.; Zavadovskaya, E.K.; Fedorov, B.V.; Starodubtsev, V.A.

    1977-01-01

    Regularities are considered in the variation of properties of glass due to irradiations. On the basis of previous theoretical statements and experimental investigations, it is inferred that the irradiation resistance of glasses of the same type, synthesis conditions, content of impurities and amount of imperfections, is a function of the ''element-oxygen'' bond energy. The irradiation resistance depends on the number and the nature of glass structure imperfections. The averaged level of bonding forces is indicative of the glass formation temperature; the imperfections in glasses are formed in structure elements whose amount predominates as compared to the others. Electric charges which accumulate on the crack surface tend to increase its size, thus lessening even further the electric strength of the dielectric. The greater the irradiation time, the greater the number of irradiation imperfections causing a drop in the electric strength of glass. When choosing a glass for service in a radiation field, it is necessary to select those of a highest temperature of glass formation and with a least amount of imperfections

  10. The effect of radiosensitizers on the survival response of hypoxic mammalian cells: The low X-ray dose region, hypersensitivity and induced radioresistance

    International Nuclear Information System (INIS)

    Skov, K.A.; MacPhail, H.S.; Marples, B.

    1994-01-01

    It has been shown previously that the extent of chemical modification of the hypoxic radiation response is dependent on dose. Some types of sensitizer are more effective at low doses (to 4 Gy) than at higher doses. Since such drugs are possible adjuvants to radiotherapy, the mechanisms responsible for the variable response at clinical doses are summarized, and the effects of cisplatin and buthionine sulfoximine on the purported induced response to radiation in hypoxic cells are presented. Cisplatin at a low, nontoxic concentration (1 μM) appears to abolish the increased radioresistant portion of the survival response. A role for high-mobility-group protein binding by platinum drugs is hypothesized to explain their interaction with radiation, and conversely, it is suggested that the heretofore unexplained different behavior of certain hypoxic sensitizers at low doses could be, at least in part, an effect on the induction of resistance. 36 refs., 2 figs

  11. Activation of mitochondrial promoter PH-binding protein in a radio-resistant Chinese hamster cell strain associated with Bcl-2

    International Nuclear Information System (INIS)

    Roychoudhury, Paromita; Ghosh, Utpal; Bhattacharyya, Nitai P.; Chaudhuri, Keya

    2006-01-01

    The cellular response to ionizing radiation is mediated by a complex interaction of number of proteins involving different pathways. Previously, we have shown that up regulation of mitochondrial genes ND1, ND4, and COX1 transcribed from the heavy strand promoter (P H ) has been increased in a radio-resistant cell strain designated as M5 in comparison with the parental Chinese hamster V79 cells. These genes are also up regulated in Chinese hamster V79 cells VB13 that express exogenous human Bcl2. In the present study, the expression of the gene ND6 that is expressed from the light strand promoter (P L ) was found to be similar in both the cell lines, as determined by RT-PCR. To test the possibility that this differential expression of mitochondrial genes under these two promoters was mediated by differences in proteins' affinity to interact with these promoters, we have carried out electrophoretic mobility shift assay (EMSA) using mitochondrial cell extracts from these two cell lines. Our result of these experiments revealed that two different proteins formed complex with the synthetic promoters and higher amount of protein from M5 cell extracts interacted with the P H promoter in comparison to that observed with cell extracts from Chinese hamster V79 cells. The promoter-specific differential binding of proteins was also observed in VB13. These results showed that differential mitochondrial gene expression observed earlier in the radio-resistant M5 cells was due to enhanced interaction proteins with the promoters P H and mediated by the expression of Bcl2

  12. TGF-β and Hypoxia/Reoxygenation Promote Radioresistance of A549 Lung Cancer Cells through Activation of Nrf2 and EGFR

    Directory of Open Access Journals (Sweden)

    Sae-lo-oom Lee

    2016-01-01

    Full Text Available Although many studies have examined the roles of hypoxia and transforming growth factor- (TGF- β separately in the tumor microenvironment, the effects of simultaneous treatment with hypoxia/reoxygenation and TGF-β on tumor malignancy are unclear. Here, we investigated the effects of redox signaling and oncogenes on cell proliferation and radioresistance in A549 human lung cancer cells in the presence of TGF-β under hypoxia/reoxygenation conditions. Combined treatment with TGF-β and hypoxia activated epidermal growth factor receptor (EGFR and nuclear factor (erythroid-derived 2-like 2 (Nrf2, a redox-sensitive transcription factor. Interestingly, Nrf2 knockdown suppressed the effects of combined treatment on EGFR phosphorylation. In addition, blockade of EGFR signaling also suppressed induction of Nrf2 following combined treatment with hypoxia and TGF-β, indicating that the combined treatment induced positive crosstalk between Nrf2 and EGFR. TGF-β and hypoxia/reoxygenation increased the accumulation of reactive oxygen species (ROS, while treatment with N-acetyl-L-cysteine abolished the activation of Nrf2 and EGFR. Treatment with TGF-β under hypoxic conditions increased the proliferation of A549 cells compared with that after vehicle treatment. Moreover, cells treated with the combined treatment exhibited resistance to ionizing radiation (IR, and knockdown of Nrf2 increased IR-induced cell death under these conditions. Thus, taken together, our findings suggested that TGF-β and hypoxia/reoxygenation promoted tumor progression and radioresistance of A549 cells through ROS-mediated activation of Nrf2 and EGFR.

  13. Daoy medulloblastoma cells that express CD133 are radioresistant relative to CD133- cells, and the CD133+ sector is enlarged by hypoxia

    International Nuclear Information System (INIS)

    Blazek, Ed R.; Foutch, Jennifer L.; Maki, Guitta

    2007-01-01

    Purpose: Primary medulloblastoma and glioblastoma multiforme tumor cells that express the surface marker CD133 are believed to be enriched for brain tumor stem cells because of their unique ability to initiate or reconstitute tumors in immunodeficient mice. This study sought to characterize the radiobiological properties and marker expression changes of CD133+ vs. CD133- cells of an established medulloblastoma cell line. Methods and Materials: Daoy and D283 Med cell lines were stained with fluorescently labeled anti-CD133 antibody and sorted into CD133+ and CD133- populations. The effect of oxygen (2% vs. 20%) on CD133 expression was measured. Both populations were analyzed for marker stability, cell cycle distribution, and radiosensitivity. Results: CD133+ Daoy cells restored nearly native CD133+ and CD133- populations within 18 days, whereas CD133- cells remained overwhelmingly CD133-. Culturing Daoy cells in 2% oxygen rather than the standard 20% oxygen increased their CD133 expression 1.6-fold. CD133+ Daoy cells were radioresistant via the β-parameter of the linear-quadratic model relative to CD133- Daoy cells, although their α-parameters and cell cycle distributions were identical. Conclusions: Restoration of the original CD133+ and CD133- populations from CD133+ Daoy cells in serum is further evidence that CD133+ cells are functionally distinct from CD133- cells. The radioresistance of CD133+ compared with CD133- Daoy cells is consistent with better repair of sublethal damage. Enlargement of the CD133+ sector is a new feature of the hypoxic response

  14. Monoclonal antibody to the type I insulin-like growth factor (IGF-I) receptor blocks IGF-I receptor-mediated DNA synthesis: clarification of the mitogenic mechanisms of IGF-I and insulin in human skin fibroblasts

    International Nuclear Information System (INIS)

    Flier, J.S.; Usher, P.; Moses, A.C.

    1986-01-01

    Insulin and insulin-like growth factor type I (IGF-I) stimulate an overlapping spectrum of biological responses in human skin fibroblasts. Although insulin and IGF-I are known to stimulate the incorporation of [ 3 H]thymidine into DNA in these cells, the identify of the receptor(s) that mediates this effect has not been fully clarified. The mouse anti-human IGF-I receptor antibody αIR-3 binds with specificity to IGF-I but not to insulin receptors in human placental membranes; it also specifically inhibits the binding of 125 I-labeled IGF-I but not 125 I-labeled insulin to suspensions of human skin fibroblasts in a dose-dependent manner. αIR-3 competitively inhibits IGF-I-mediated stimulation of [ 3 H]thymidine incorporation into DNA. This inhibition is dependent on the concentration of αIR-3 and in the presence of a fixed antibody concentration can be partially overcome by high concentrations of IGF-I. In contrast, at concentrations of 3 H]thymidine incorporation is not inhibited by αIR-3. However, the incremental effects of higher concentrations (> 1 μg/ml) of insulin on [ 3 H]thymidine incorporation are inhibited by αIR-3. αIR-3 is a highly specific antagonist of IGF-I receptor-mediated mitogenesis in human skin fibroblasts. By using this antibody, it is shown directly that insulin can act through the IGF-I receptor to stimulate DNA synthesis but can also activate this effect through the insulin receptor itself

  15. Opioid receptor mediated anticonvulsant effect of pentazocine.

    Science.gov (United States)

    Khanna, N; Khosla, R; Kohli, J

    1998-01-01

    Intraperitoneal (i.p.) administration of (+/-) pentazocine (10, 30 & 50 mg/kg), a Sigma opioid agonist, resulted in a dose dependent anticonvulsant action against maximal electroshock seizures in mice. This anticonvulsant effect of pentazocine was not antagonized by both the doses of naloxone (1 and 10 mg/kg) suggesting thereby that its anticonvulsant action is probably mediated by Sigma opiate binding sites. Its anticonvulsant effect was potentiated by both the anticonvulsant drugs viz. diazepam and diphenylhydantoin. Morphine, mu opioid agonist, on the other hand, failed to protect the animals against maximal electroshock seizures when it was given in doses of 10-40 mg/kg body wt.

  16. Molecular Mechanisms of Dopamine Receptor Mediated Neuroprotection

    National Research Council Canada - National Science Library

    Sealfon, Stuart

    2000-01-01

    ... of the cellular changes characteristic of this process. Evidence from our laboratory and others suggest that activation of dopamine receptors can oppose the induction of apoptosis in dopamine neurons...

  17. Inositol trisphosphate receptor mediated spatiotemporal calcium signalling.

    Science.gov (United States)

    Miyazaki, S

    1995-04-01

    Spatiotemporal Ca2+ signalling in the cytoplasm is currently understood as an excitation phenomenon by analogy with electrical excitation in the plasma membrane. In many cell types, Ca2+ waves and Ca2+ oscillations are mediated by inositol 1,4,5-trisphosphate (IP3) receptor/Ca2+ channels in the endoplasmic reticulum membrane, with positive feedback between cytosolic Ca2+ and IP3-induced Ca2+ release creating a regenerative process. Remarkable advances have been made in the past year in the analysis of subcellular Ca2+ microdomains using confocal microscopy and of Ca2+ influx pathways that are functionally coupled to IP3-induced Ca2+ release. Ca2+ signals can be conveyed into the nucleus and mitochondria. Ca2+ entry from outside the cell allows repetitive Ca2+ release by providing Ca2+ to refill the endoplasmic reticulum stores, thus giving rise to frequency-encoded Ca2+ signals.

  18. Early LC3 lipidation induced by d-limonene does not rely on mTOR inhibition, ERK activation and ROS production and it is associated with reduced clonogenic capacity of SH-SY5Y neuroblastoma cells.

    Science.gov (United States)

    Berliocchi, Laura; Chiappini, Carlotta; Adornetto, Annagrazia; Gentile, Debora; Cerri, Silvia; Russo, Rossella; Bagetta, Giacinto; Corasaniti, Maria Tiziana

    2018-02-01

    d-Limonene is a natural monoterpene abundant in Citrus essential oils. It is endowed with several biological activities, including inhibition of carcinogenesis and promotion of tumour regression. Recently, d-limonene has been shown to modulate autophagic markers in vitro at concentrations found in vivo, in clinical pharmacokinetic studies. Autophagy is an intracellular catabolic process serving as both an adaptive metabolic response and a quality control mechanism. Because autophagy defects have been linked to a wide range of human pathologies, including neurodegeneration and cancer, there is a need for new pharmacological tools to control deregulated autophagy. To better understand the effects of d-limonene on autophagy, to identify the molecular mechanisms through which this monoterpene rapidly triggers LC3 lipidation and to evaluate the role for autophagy in long-term effects of d-limonene. Human SH-SY5Y neuroblastoma, HepG2 hepatocellular carcinoma and MCF7 breast cancer cells were used. Endogenous LC3-II levels were evaluated by western blotting. Autophagic flux assay was performed using bafilomycin A1 and chloroquine. Intracellular distribution of LC3 protein was studied by confocal microscopy analysis of LC3B-GFP transduced cells. Expression of lysosomal-membrane protein LAMP-1 was assessed by immunofluorescence analysis. Phosphorylated levels of downstream substrates of mTOR kinase (p70S6 kinase, 4E-BP1, and ULK1) and ERK were analyzed by western blotting. Production of reactive oxygen species (ROS) was assessed by live confocal microscopy of cells loaded with CellROX ® Green Reagent. Clonogenic assay was used to evaluate the ability of treated cells to proliferate and form colonies. LC3 lipidation promoted by d-limonene correlates with autophagosome formation and stimulation of basal autophagy. LC3 lipidation does not rely on inhibition of mTOR kinase, which instead appears to be transiently activated. In addition, d-limonene rapidly activates ERK and

  19. Ga-66 labeled somatostatin analogue DOTA-DPhe1-Tyr3-octreotide as a potential agent for positron emission tomography imaging and receptor mediated internal radiotherapy of somatostatin receptor positive tumors

    International Nuclear Information System (INIS)

    Ugur, Oemer; Kothari, Paresh J.; Finn, Ronald D.; Zanzonico, Pat; Ruan, Shutian; Guenther, Ilonka; Maecke, Helmut R.; Larson, Steven M.

    2002-01-01

    Radionuclide labeled somatostatin analogues selectively target somatostatin receptor (SSTR)-expressing tumors as a basis for diagnosis and treatment of these tumors. Recently, a DOTA-functionalized somatostatin analogue, DOTATOC (DOTA-DPhe 1 -Tyr 3 -octreotide) has been developed. This compound has been shown to be superior to the other somatostatin analogues as indicated by its uniquely high tumor-to-non-target tissue ratio. DOTATOC can be labeled with a variety of radiometals including gallium radioisotopes. Gallium-66 is a positron emitting radionuclide (T 1/2 =9.5 hr; β + =56%), that can be produced in carrier free form by a low-beam energy cyclotron. In this study we investigated SSTR targeting characteristics of 66 Ga-DOTATOC in AR42J rat pancreas tumor implanted nude mice as a potential agent for diagnosis and receptor-mediated internal radiotherapy of SSTR-expressing tumors. We compared our results with 67 Ga- and 68 Ga- labeled DOTATOC. The radiolabeling procedure gave labeling yield ranged from 85-95% and radiochemical and chemical purity was >95%. In-vitro competitive binding curves and in-vivo competitive displacement studies with an excess of unlabeled peptide indicates that there is specific binding of the radioligand to SSTR. Animal biodistribution data and serial microPET TM images demonstrated rapid tumor uptake and rapid clearance from the blood and all tissues except kidney. Maximum % ID/g values for tumor were 10.0±0.7, 13.2±2.1 and 9.8±1.5 for 66 Ga-, 67 Ga-, and 68 Ga-DOTATOC, respectively. Calculated tumor, kidney and bone marrow doses for 66 Ga-DOTATOC based on biodistribution data were 178, 109 and 1.2 cGy/MBq, respectively. We conclude that 66 Ga labeled DOTATOC can be used for PET diagnosis and quantitative imaging-based dosimetry of SSTR positive tumors. 66 Ga-DOTATOC may also be used in higher doses for ablation of these tumors. However, kidney is the critical organ for toxicity (tumor/kidney ratio 1.64), and high kidney uptake must

  20. Ga-66 labeled somatostatin analogue DOTA-DPhe{sup 1}-Tyr{sup 3}-octreotide as a potential agent for positron emission tomography imaging and receptor mediated internal radiotherapy of somatostatin receptor positive tumors

    Energy Technology Data Exchange (ETDEWEB)

    Ugur, Oemer E-mail: ougur@hacettepe.edu.tr; Kothari, Paresh J.; Finn, Ronald D.; Zanzonico, Pat; Ruan, Shutian; Guenther, Ilonka; Maecke, Helmut R.; Larson, Steven M

    2002-02-01

    Radionuclide labeled somatostatin analogues selectively target somatostatin receptor (SSTR)-expressing tumors as a basis for diagnosis and treatment of these tumors. Recently, a DOTA-functionalized somatostatin analogue, DOTATOC (DOTA-DPhe{sup 1}-Tyr{sup 3}-octreotide) has been developed. This compound has been shown to be superior to the other somatostatin analogues as indicated by its uniquely high tumor-to-non-target tissue ratio. DOTATOC can be labeled with a variety of radiometals including gallium radioisotopes. Gallium-66 is a positron emitting radionuclide (T{sub 1/2} =9.5 hr; {beta}{sup +}=56%), that can be produced in carrier free form by a low-beam energy cyclotron. In this study we investigated SSTR targeting characteristics of {sup 66}Ga-DOTATOC in AR42J rat pancreas tumor implanted nude mice as a potential agent for diagnosis and receptor-mediated internal radiotherapy of SSTR-expressing tumors. We compared our results with {sup 67}Ga- and {sup 68}Ga- labeled DOTATOC. The radiolabeling procedure gave labeling yield ranged from 85-95% and radiochemical and chemical purity was >95%. In-vitro competitive binding curves and in-vivo competitive displacement studies with an excess of unlabeled peptide indicates that there is specific binding of the radioligand to SSTR. Animal biodistribution data and serial microPET{sup TM} images demonstrated rapid tumor uptake and rapid clearance from the blood and all tissues except kidney. Maximum % ID/g values for tumor were 10.0{+-}0.7, 13.2{+-}2.1 and 9.8{+-}1.5 for {sup 66}Ga-, {sup 67}Ga-, and {sup 68}Ga-DOTATOC, respectively. Calculated tumor, kidney and bone marrow doses for {sup 66}Ga-DOTATOC based on biodistribution data were 178, 109 and 1.2 cGy/MBq, respectively. We conclude that {sup 66}Ga labeled DOTATOC can be used for PET diagnosis and quantitative imaging-based dosimetry of SSTR positive tumors. {sup 66}Ga-DOTATOC may also be used in higher doses for ablation of these tumors. However, kidney is the

  1. h5-HT1B receptor-mediated constitutive Gαi3-protein activation in stably transfected Chinese hamster ovary cells: an antibody capture assay reveals protean efficacy of 5-HT

    Science.gov (United States)

    Newman-Tancredi, Adrian; Cussac, Didier; Marini, Laetitia; Touzard, Manuelle; Millan, Mark J

    2003-01-01

    Serotonin 5-HT1B receptors couple to G-proteins of the Gi/o family. However, their activation of specific G-protein subtypes is poorly characterised. Using an innovative antibody capture/guanosine-5′-0-(3-[35S]thio)-triphosphate ([35S]GTPγS) binding strategy, we characterised Gαi3 subunit activation by h5-HT1B receptors stably expressed in Chinese hamster ovary (CHO) cells. The agonists, 5-HT, alniditan and BMS181,101, stimulated Gαi3, whereas methiothepin and SB224,289 behaved as inverse agonists. The selective 5-HT1B receptor ligand, S18127, modestly stimulated Gαi3 and reversed the actions of both 5-HT and methiothepin. S18127 (1 μM) also produced parallel, dextral shifts of the 5-HT and methiothepin isotherms. Isotopic dilution experiments ([35S]GTPγS versus GTPγS) revealed high-affinity [35S]GTPγS binding to Gαi3 subunits in the absence of receptor ligands indicating constitutive activity. High-affinity [35S]GTPγS binding was increased 2.8-fold by 5-HT with an increase in the affinity of GTPγS for Gαi3 subunits. In contrast, methiothepin halved the number of high-affinity binding sites and decreased their affinity. h5-HT1B receptor-mediated Gαi3 subunit activation was dependent on the concentration of NaCl. At 300 mM, 5-HT stimulated [35S]GTPγS binding, basal Gαi3 activation was low and methiothepin was inactive. In contrast, at 10 mM NaCl, basal activity was enhanced and the inverse agonist activity of methiothepin was accentuated. Under these conditions, 5-HT decreased Gαi3 activation. In conclusion, at h5-HT1B receptors expressed in CHO cells: (i) inverse agonist induced inhibition of Gαi3, and its reversal by S18127, reveals constitutive activation of this Gα subunit; (ii) constitutive Gαi3 activation can be quantified by isotopic dilution [35S]GTPγS binding and (iii) decreasing NaCl concentrations enhances Gαi3 activation and leads to protean agonist properties of 5-HT: that is a switch to inhibition of Gαi3. PMID:12684263

  2. h5-HT(1B) receptor-mediated constitutive Galphai3-protein activation in stably transfected Chinese hamster ovary cells: an antibody capture assay reveals protean efficacy of 5-HT.

    Science.gov (United States)

    Newman-Tancredi, Adrian; Cussac, Didier; Marini, Laetitia; Touzard, Manuelle; Millan, Mark J

    2003-03-01

    1. Serotonin 5-HT(1B) receptors couple to G-proteins of the Gi/o family. However, their activation of specific G-protein subtypes is poorly characterised. Using an innovative antibody capture/guanosine-5'-0-(3-[(35)S]thio)-triphosphate ([(35)S]GTPgammaS) binding strategy, we characterised Galpha(i3) subunit activation by h5-HT(1B) receptors stably expressed in Chinese hamster ovary (CHO) cells. 2. The agonists, 5-HT, alniditan and BMS181,101, stimulated Galpha(i3), whereas methiothepin and SB224,289 behaved as inverse agonists. The selective 5-HT(1B) receptor ligand, S18127, modestly stimulated Galpha(i3) and reversed the actions of both 5-HT and methiothepin. S18127 (1 micro M) also produced parallel, dextral shifts of the 5-HT and methiothepin isotherms. 3. Isotopic dilution experiments ([(35)S]GTPgammaS versus GTPgammaS) revealed high-affinity [(35)S]GTPgammaS binding to Galpha(i3) subunits in the absence of receptor ligands indicating constitutive activity. High-affinity [(35)S]GTPgammaS binding was increased 2.8-fold by 5-HT with an increase in the affinity of GTPgammaS for Galpha(i3) subunits. In contrast, methiothepin halved the number of high-affinity binding sites and decreased their affinity. 4. h5-HT(1B) receptor-mediated Galpha(i3) subunit activation was dependent on the concentration of NaCl. At 300 mM, 5-HT stimulated [(35)S]GTPgammaS binding, basal Galpha(i3) activation was low and methiothepin was inactive. In contrast, at 10 mM NaCl, basal activity was enhanced and the inverse agonist activity of methiothepin was accentuated. Under these conditions, 5-HT decreased Galpha(i3) activation. 5. In conclusion, at h5-HT(1B) receptors expressed in CHO cells: (i) inverse agonist induced inhibition of Galpha(i3), and its reversal by S18127, reveals constitutive activation of this Galpha subunit; (ii) constitutive Galpha(i3) activation can be quantified by isotopic dilution [(35)S]GTPgammaS binding and (iii) decreasing NaCl concentrations enhances Galpha(i3

  3. Extracting the normal lung dose–response curve from clinical DVH data: a possible role for low dose hyper-radiosensitivity, increased radioresistance

    International Nuclear Information System (INIS)

    Gordon, J J; Snyder, K; Zhong, H; Barton, K; Sun, Z; Chetty, I J; Matuszak, M; Ten Haken, R K

    2015-01-01

    In conventionally fractionated radiation therapy for lung cancer, radiation pneumonitis’ (RP) dependence on the normal lung dose-volume histogram (DVH) is not well understood. Complication models alternatively make RP a function of a summary statistic, such as mean lung dose (MLD). This work searches over damage profiles, which quantify sub-volume damage as a function of dose. Profiles that achieve best RP predictive accuracy on a clinical dataset are hypothesized to approximate DVH dependence.Step function damage rate profiles R(D) are generated, having discrete steps at several dose points. A range of profiles is sampled by varying the step heights and dose point locations. Normal lung damage is the integral of R(D) with the cumulative DVH. Each profile is used in conjunction with a damage cutoff to predict grade 2 plus (G2+) RP for DVHs from a University of Michigan clinical trial dataset consisting of 89 CFRT patients, of which 17 were diagnosed with G2+ RP.Optimal profiles achieve a modest increase in predictive accuracy—erroneous RP predictions are reduced from 11 (using MLD) to 8. A novel result is that optimal profiles have a similar distinctive shape: enhanced damage contribution from low doses (<20 Gy), a flat contribution from doses in the range ∼20–40 Gy, then a further enhanced contribution from doses above 40 Gy. These features resemble the hyper-radiosensitivity / increased radioresistance (HRS/IRR) observed in some cell survival curves, which can be modeled using Joiner’s induced repair model.A novel search strategy is employed, which has the potential to estimate RP dependence on the normal lung DVH. When applied to a clinical dataset, identified profiles share a characteristic shape, which resembles HRS/IRR. This suggests that normal lung may have enhanced sensitivity to low doses, and that this sensitivity can affect RP risk. (paper)

  4. γ-Glutamylcysteine synthetase (γ-GCS) as a target for overcoming chemo- and radio-resistance of human hepatocellular carcinoma cells.

    Science.gov (United States)

    Lin, Li-Ching; Chen, Chi-Fen; Ho, Chun-Te; Liu, Jun-Jen; Liu, Tsan-Zon; Chern, Chi-Liang

    2018-04-01

    This study uncovered that the genetically endowed intracellular glutathione contents (iGSH) regulated by the catalytic subunit of γ‑glutamylcysteine synthetase heavy chain (γ‑GCSh) as a prime target for overcoming both the inherited and stimuli-activated chemo- and radio-resistance of hepatocellular carcinoma (HCC) cells. Reactive oxygen species (ROS) production and mitochondrial membrane potential (Δψm) were determined by the probe-based flow cytometry. The TUNEL assay was used as an index of radio-sensitivity and the MTT assay was used as an index of chemo-sensitivity against various anti-cancer agents. iGSH and γ‑GCSh activity were measured by HPLC methods. γ‑GCSh-overexpressing GCS30 cell line was established by tetracycline-controlled Tet-OFF gene expression system in SK-Hep-1 cells. The relative radio-sensitivities of a panel of five HCC cells were found to be correlated negatively with both the contents of iGSH and their corresponding γ‑GCSh activities with an order of abundance being Hep G2 > Hep 3B > J5 > Mahlavu > SK-Hep-1, respectively. Similarly, the cytotoxicity response patterns of these HCC cells against arsenic trioxide (ATO), a ROS-producing anti-cancer drug, were exactly identical to the order of ranking instigated by the radiotherapy (RT) treatment. Next, γ‑GCSh-overexpressing GCS30 cells were found to possess excellent ability to profoundly mitigate both the drop of Δψm and apoptotic TUNEL-positive cell population engendered by ATO, cisplatin, doxorubicin, and RT treatments. Our data unequivocally demonstrate that γ‑GCSh may represent a prime target for overcoming anti-cancer drugs and RT resistance for HCC cells. Copyright © 2018 Elsevier Inc. All rights reserved.

  5. Auto-SCT induces a phenotypic shift from CMP to GMP progenitors, reduces clonogenic potential and enhances in vitro and in vivo cycling activity defined by (18)F-FLT PET scanning.

    Science.gov (United States)

    Woolthuis, C; Agool, A; Olthof, S; Slart, R H J A; Huls, G; Smid, W M; Schuringa, J J; Vellenga, E

    2011-01-01

    Autologous SCT (auto-SCT) introduces a reduced tolerance to chemotherapy even in patients with adequate engraftment, suggesting long-term effects of the transplantation procedure on the BM capacity. To study the hematopoietic cell compartment after auto-SCT, CD34(+) BM cells (n = 16) from patients at 6-9 months after auto-SCT were studied with regard to the progenitor subsets, colony frequency and cell cycle status. The BM compartments were studied in vivo using PET tracer 3-fluoro-3-deoxy-L-thymidine (¹⁸F-FLT PET). BM CD34(+) cells after auto-SCT were compared with normal CD34(+) cells and showed a phenotypic shift from common myeloid progenitor (CMP mean percentage 3.7 vs 19.4%, P=0.001) to granulocyte-macrophage progenitor (GMP mean percentage 51.8 vs 27.6%, P=0.01). In addition, a reduced clonogenic potential and higher cycling activity especially of the GMP fraction (41% ± 4 in G2/S phase vs 19% ± 2, P = 0.03) were observed in BM after auto-SCT compared with normal. The enhanced cycling activity was confirmed in vivo by showing a significantly higher uptake of the ¹⁸F-FLT PET tracer by the BM compartment. This study shows that auto-SCT results in defects of the hematopoietic compartment at least 6 months after auto-SCT, characterized by changes in the composition of progenitor subsets and enhanced in vitro and in vivo cycling activity.

  6. Radioresistant Spodoptera frugiperda 9 insect cells display excessive resistance to 'endoplasmic reticulum' stress and calcium disturbances via pre-emptive activation of unfolded protein response pathway

    International Nuclear Information System (INIS)

    Guleria, Ayushi; Chandna, Sudhir

    2016-01-01

    Endoplasmic Reticulum (ER) performs multiple cellular functions such as proper folding of newly synthesized proteins and calcium homeostasis. ER stress triggers unfolded protein response (UPR) that attempts to restore normal ER function and resists damage-induced cell death. Lepidopteran Sf9 insect cells (derived from Spodoptera frugiperda) display 100-200 times higher radioresistance than mammalian cells. We have earlier reported that gamma-radiation doses <1000 Gy fail to trigger increase in cytosolic calcium in Sf9 cells, indicating resilience to calcium/ ER disturbances

  7. Single Strand Annealing Plays a Major Role in RecA-Independent Recombination between Repeated Sequences in the Radioresistant Deinococcus radiodurans Bacterium.

    Directory of Open Access Journals (Sweden)

    Solenne Ithurbide

    2015-10-01

    Full Text Available The bacterium Deinococcus radiodurans is one of the most radioresistant organisms known. It is able to reconstruct a functional genome from hundreds of radiation-induced chromosomal fragments. Our work aims to highlight the genes involved in recombination between 438 bp direct repeats separated by intervening sequences of various lengths ranging from 1,479 bp to 10,500 bp to restore a functional tetA gene in the presence or absence of radiation-induced DNA double strand breaks. The frequency of spontaneous deletion events between the chromosomal direct repeats were the same in recA+ and in ΔrecA, ΔrecF, and ΔrecO bacteria, whereas recombination between chromosomal and plasmid DNA was shown to be strictly dependent on the RecA and RecF proteins. The presence of mutations in one of the repeated sequence reduced, in a MutS-dependent manner, the frequency of the deletion events. The distance between the repeats did not influence the frequencies of deletion events in recA+ as well in ΔrecA bacteria. The absence of the UvrD protein stimulated the recombination between the direct repeats whereas the absence of the DdrB protein, previously shown to be involved in DNA double strand break repair through a single strand annealing (SSA pathway, strongly reduces the frequency of RecA- (and RecO- independent deletions events. The absence of the DdrB protein also increased the lethal sectoring of cells devoid of RecA or RecO protein. γ-irradiation of recA+ cells increased about 10-fold the frequencies of the deletion events, but at a lesser extend in cells devoid of the DdrB protein. Altogether, our results suggest a major role of single strand annealing in DNA repeat deletion events in bacteria devoid of the RecA protein, and also in recA+ bacteria exposed to ionizing radiation.

  8. Cell death induced by ionizing radiations in human radio-resistant tumours: in-vitro and in-vivo study of mechanisms involved in its induction by different types of radiations and pharmacological modulation

    International Nuclear Information System (INIS)

    Altmeyer, Anais

    2010-01-01

    Whereas chemo-radiotherapy protocols revealed to be very efficient when taking tumours into care, the treatment of some tumours remains very limited due to their critical location or to the weak radio-sensitivity to conventional radiations. One way to work around this problem is to use high linear energy transfer radiations or hadron therapy, in combination with radio-sensitizers. This research thesis reports the assessment of radio-sensitizer effects of different molecules on human radio-resistant cell lines and more particularly the SK-Hep1 line from a hepatocellular carcinoma. In vitro studies have been performed and then in vivo studies by using fast neutron irradiation on a mice liver sample. Observations made by optic fibre confocal microscopy and transmission electronic microscopy confirmed in vitro observations: the prevailing cell death after such an irradiation is the autophagic cell death. It shows the importance of the autophagic phenomenon induced by radiations with high linear transfer energy. This could lead to new therapeutic protocols for radio-resistant cancers [fr

  9. Study of radioresistance of microorganisms

    International Nuclear Information System (INIS)

    Olbrichova, D.

    1983-01-01

    Strain radiosensitivity was evaluated on the basis of the determination of inactivation curve parameters and the determination of the D 10 value. The course of curves was investigated for two 60 Co sources, RCHM-gamma-20 (USSR) with an activity of 2.54x10 14 Bq and AECL (type J-6000, Canada) with an activity of 4.92x10 16 Bq. The curve parameters were determined for microorganisms in buffer medium and for cultures in dehydrated condition. Coccus germs were irradiated with doses of 1, 2, 3, 4, 5 and 6 kGy and aerobic sporulates with doses of 3, 6, 9, 12 and 15 kGy. For comparing the resistance of isolated strains the t-test was used. The most resistant of the isolated strains were Bacillus cereus, Bacillus subtilis, Bacillus pumilus and Bacillus megatherium. (E.S.)

  10. Double-negative feedback loop between long non-coding RNA TUG1 and miR-145 promotes epithelial to mesenchymal transition and radioresistance in human bladder cancer cells.

    Science.gov (United States)

    Tan, Jiemei; Qiu, Kaifeng; Li, Mingyi; Liang, Ying

    2015-10-07

    LncRNAs have a critical role in the regulation of cellular processes such as cancer progression and metastasis. In the present study, we confirmed that TUG1 was overexpressed in bladder cancer tissues and established cell lines. Knockdown of TUG1 inhibited bladder cancer cell metastasis both in vitro and in vivo. Furthermore, we found that TUG1 promoted cancer cell invasion and radioresistance through inducing epithelial-to-mesenchymal transition (EMT). Interestingly, TUG1 decreased the expression of miR-145 and there was a reciprocal repression between TUG1 and miR-145 in an Argonaute2-dependent manner. ZEB2 was identified as a down-stream target of miR-145 and TUG1 exerted its function through the miR-145/ZEB2 axis. In summary, our data indicated that blocking TUG1 function may be an effective anti-cancer therapy. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  11. MiR-146b-5p overexpression attenuates stemness and radioresistance of glioma stem cells by targeting HuR/lincRNA-p21/β-catenin pathway

    Science.gov (United States)

    Yang, Wei; Yu, Hongquan; Shen, Yueming; Liu, Yingying; Yang, Zhanshan; Sun, Ting

    2016-01-01

    A stem-like subpopulation existed in GBM cells, called glioma stem cells (GSCs), might contribute to cancer invasion, angiogenesis, immune evasion, and therapeutic resistance, providing a rationale to eliminate GSCs population and their supporting niche for successful GBM treatment. LincRNA-p21, a novel regulator of cell proliferation, apoptosis and DNA damage response, is found to be downregulated in several types of tumor. However, little is known about the role of lincRNA-p21 in stemness and radioresistance of GSCs and its regulating mechanisms. In this study, we found that lincRNA-p21 negatively regulated the expression and activity of β-catenin in GSCs. Downregulation of lincRNA-p21 in GSCs was resulted from upregulation of Hu antigen R (HuR) expression caused by miR-146b-5p downregulation. MiR-146b-5p overexpression increased apoptosis and radiosensitivity, decreased cell viability, neurosphere formation capacity and stem cell marker expression, and induced differentiation in GSCs. Moreover, knock-down lincRNA-p21 or HuR and β-catenin overexpression could rescue the phenotypic changes resulted from miR-146b-5p overexpression in GSCs. These findings suggest that targeting the miR-146b-5p/HuR/lincRNA-p21/β-catenin signaling pathway may be valuable therapeutic strategies against glioma. PMID:27166258

  12. Suppression of Thyroid Hormone Receptor-Mediated Transcription ...

    African Journals Online (AJOL)

    TH)-induced TR-mediated transcription. We further examined the effects of methamidophos on TR-thyroid hormone response element (TRE) binding using the liquid chemiluminescent DNA pull-down assay (LCDPA), and found no dissociation of ...

  13. Adrenergic receptor-mediated modulation of striatal firing patterns.

    Science.gov (United States)

    Ohta, Hiroyuki; Kohno, Yu; Arake, Masashi; Tamura, Risa; Yukawa, Suguru; Sato, Yoshiaki; Morimoto, Yuji; Nishida, Yasuhiro; Yawo, Hiromu

    2016-11-01

    Although noradrenaline and adrenaline are some of the most important neurotransmitters in the central nervous system, the effects of noradrenergic/adrenergic modulation on the striatum have not been determined. In order to explore the effects of adrenergic receptor (AR) agonists on the striatal firing patterns, we used optogenetic methods which can induce continuous firings. We employed transgenic rats expressing channelrhodopsin-2 (ChR2) in neurons. The medium spiny neuron showed a slow rising depolarization during the 1-s long optogenetic striatal photostimulation and a residual potential with 8.6-s half-life decay after the photostimulation. As a result of the residual potential, five repetitive 1-sec long photostimulations with 20-s onset intervals cumulatively increased the number of spikes. This 'firing increment', possibly relating to the timing control function of the striatum, was used to evaluate the AR modulation. The β-AR agonist isoproterenol decreased the firing increment between the 1st and 5th stimulation cycles, while the α 1 -AR agonist phenylephrine enhanced the firing increment. Isoproterenol and adrenaline increased the early phase (0-0.5s of the photostimulation) firing response. This adrenergic modulation was inhibited by the β-antagonist propranolol. Conversely, phenylephrine and noradrenaline reduced the early phase response. β-ARs and α 1 -ARs work in opposition controlling the striatal firing initiation and the firing increment. Copyright © 2016 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.

  14. Outline of therapeutic interventions with muscarinic receptor-mediated transmission

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; Šantrůčková, Eva; Randáková, Alena; Janíčková, Helena; Zimčík, Pavel; Rudajev, Vladimír; Michal, Pavel; El-Fakahany, E. E.; Doležal, Vladimír

    2014-01-01

    Roč. 63, Suppl.1 (2014), S177-S189 ISSN 0862-8408 R&D Projects: GA ČR(CZ) GA305/09/0681; GA ČR(CZ) GAP304/12/0259; GA MŠk(CZ) 7E10060 Institutional support: RVO:67985823 Keywords : cholinergic transmission * muscarinic receptors * therapy * Alzheimer's disease, * schizophrenia Subject RIV: ED - Physiology Impact factor: 1.293, year: 2014

  15. Fas receptor-mediated apoptosis : a clinical application?

    NARCIS (Netherlands)

    Timmer, T; de Vries, EGE; de Jong, S

    Fas is a membrane protein belonging to the death receptor family. Cross-linking of Fas by its ligand, FasL, or agonistic anti-Fas antibodies, induces apoptosis of cells expressing Fas on the membrane by triggering a cascade of caspases. Since many different tumours express Fas on their membrane,

  16. Dynamics of Receptor-Mediated Nanoparticle Internalization into Endothelial Cells

    Science.gov (United States)

    Gonzalez-Rodriguez, David; Barakat, Abdul I.

    2015-01-01

    Nanoparticles offer a promising medical tool for targeted drug delivery, for example to treat inflamed endothelial cells during the development of atherosclerosis. To inform the design of such therapeutic strategies, we develop a computational model of nanoparticle internalization into endothelial cells, where internalization is driven by receptor-ligand binding and limited by the deformation of the cell membrane and cytoplasm. We specifically consider the case of nanoparticles targeted against ICAM-1 receptors, of relevance for treating atherosclerosis. The model computes the kinetics of the internalization process, the dynamics of binding, and the distribution of stresses exerted between the nanoparticle and the cell membrane. The model predicts the existence of an optimal nanoparticle size for fastest internalization, consistent with experimental observations, as well as the role of bond characteristics, local cell mechanical properties, and external forces in the nanoparticle internalization process. PMID:25901833

  17. Of pheromones and kairomones: what receptors mediate innate emotional responses?

    Science.gov (United States)

    Fortes-Marco, Lluis; Lanuza, Enrique; Martinez-Garcia, Fernando

    2013-09-01

    Some chemicals elicit innate emotionally laden behavioral responses. Pheromones mediate sexual attraction, parental care or agonistic confrontation, whereas predators' kairomones elicit defensive behaviors in their preys. This essay explores the hypothesis that the detection of these semiochemicals relies on highly specific olfactory and/or vomeronasal receptors. The V1R, V2R, and formyl-peptide vomeronasal receptors bind their ligands in highly specific and sensitive way, thus being good candidates for pheromone- or kairomone-detectors (e.g., secreted and excreted proteins, peptides and lipophilic volatiles). The olfactory epithelium also expresses specific receptors, for example trace amine-associated receptors (TAAR) and guanylyl cyclase receptors (GC-D and other types), some of which bind kairomones and putative pheromones. However, most of the olfactory neurons express canonical olfactory receptors (ORs) that bind many ligands with different affinity, being not suitable for mediating responses to pheromones and kairomones. In this respect, trimethylthiazoline (TMT) is considered a fox-derived kairomone for mice and rats, but it seems to be detected by canonical ORs. Therefore, we have reassessed the kairomonal nature of TMT by analyzing the behavioral responses of outbred (CD1) and inbred mice (C57BL/J6) to TMT. Our results confirm that both mouse strains avoid TMT, which increases immobility in C57BL/J6, but not CD1 mice. However, mice of both strains sniff at TMT throughout the test and show no trace of TMT-induced contextual conditioning (immobility or avoidance). This suggests that TMT is not a kairomone but, similar to a loud noise, in high concentrations it induces aversion and stress as unspecific responses to a strong olfactory stimulation. Copyright © 2013 Wiley Periodicals, Inc.

  18. Effect of ARA9 on dioxin receptor mediated transcription

    International Nuclear Information System (INIS)

    Lees, M.J.; Whitelaw, M.L.

    2002-01-01

    The dioxin (Aryl hydrocarbon) receptor (DR) is a unique bHLH transcription factor which is activated by binding of planar aromatic hydrocarbons typified by dioxin (TCDD). The active receptor is key to metabolism of aryl hydrocarbon xenobiotics by being a potent inducer of CYP1A1 gene activity. Chlorinated dioxins are inert to metabolism and initiate multifarious toxicities, including potent tumour promotion. These ill-effects are mediated by the activated DR and we are studying the mechanisms by which the ligand binding domain of the DR controls activity of the protein. The DR ligand binding domain resides within a PAS (Per/Arnt/Sim homology) region which is contiguous with the bHLH. The latent bHLH/PAS dioxin receptor (DR) is found in the cytoplasm of most mammalian cell types in a complex with heat shock protein 90, a novel immunophilin like protein termed ARA9/XAP2/AIP, and the co-chaperone p23. Here we use antisense ARA9 constructs to reveal that in the absence of ARA9, the DR is unable to form a transcriptionally active complex. Co-expression of antisense ARA9 with a form of the DR which is constitutively targeted to the nucleus leads to dramatically decreased levels of the nuclear DR protein, implying that ARA9 may function beyond its currently proposed role in cytoplasmic retention of the latent DR

  19. Ligand Receptor-Mediated Regulation of Growth in Plants.

    Science.gov (United States)

    Haruta, Miyoshi; Sussman, Michael R

    2017-01-01

    Growth and development of multicellular organisms are coordinately regulated by various signaling pathways involving the communication of inter- and intracellular components. To form the appropriate body patterns, cellular growth and development are modulated by either stimulating or inhibiting these pathways. Hormones and second messengers help to mediate the initiation and/or interaction of the various signaling pathways in all complex multicellular eukaryotes. In plants, hormones include small organic molecules, as well as larger peptides and small proteins, which, as in animals, act as ligands and interact with receptor proteins to trigger rapid biochemical changes and induce the intracellular transcriptional and long-term physiological responses. During the past two decades, the availability of genetic and genomic resources in the model plant species, Arabidopsis thaliana, has greatly helped in the discovery of plant hormone receptors and the components of signal transduction pathways and mechanisms used by these immobile but highly complex organisms. Recently, it has been shown that two of the most important plant hormones, auxin and abscisic acid (ABA), act through signaling pathways that have not yet been recognized in animals. For example, auxins stimulate cell elongation by bringing negatively acting transcriptional repressor proteins to the proteasome to be degraded, thus unleashing the gene expression program required for increasing cell size. The "dormancy" inducing hormone, ABA, binds to soluble receptor proteins and inhibits a specific class of protein phosphatases (PP2C), which activates phosphorylation signaling leading to transcriptional changes needed for the desiccation of the seeds prior to entering dormancy. While these two hormone receptors have no known animal counterparts, there are also many similarities between animal and plant signaling pathways. For example, in plants, the largest single gene family in the genome is the protein kinase family (approximately 5% of the protein coding genes), although the specific function for only a few dozen of these kinases is clearly established. Recent comparative genomics studies have revealed that parasitic nematodes and pathogenic microbes produce plant peptide hormone mimics that target specific plant plasma membrane receptor-like protein kinases, thus usurping endogenous signaling pathways for their own pathogenic purposes. With biochemical, genetic, and physiological analyses of the regulation of hormone receptor signal pathways, we are thus just now beginning to understand how plants optimize the development of their body shape and cope with constantly changing environmental conditions. © 2017 Elsevier Inc. All rights reserved.

  20. Environmental phthalate monoesters activate pregnane X receptor-mediated transcription

    International Nuclear Information System (INIS)

    Hurst, Christopher H.; Waxman, David J.

    2004-01-01

    Phthalate esters, widely used as plasticizers in the manufacture of products made of polyvinyl chloride, induce reproductive and developmental toxicities in rodents. The mechanism that underlies these effects of phthalate exposure, including the potential role of members of the nuclear receptor superfamily, is not known. The present study investigates the effects of phthalates on the pregnane X receptor (PXR), which mediates the induction of enzymes involved in steroid metabolism and xenobiotic detoxification. The ability of phthalate monoesters to activate PXR-mediated transcription was assayed in a HepG2 cell reporter assay following transfection with mouse PXR (mPXR), human PXR (hPXR), or the hPXR allelic variants V140M, D163G, and A370T. Mono-2-ethylhexyl phthalate (MEHP) increased the transcriptional activity of both mPXR and hPXR (5- and 15-fold, respectively) with EC 50 values of 7-8 μM. mPXR and hPXR were also activated by monobenzyl phthalate (MBzP, up to 5- to 6-fold) but were unresponsive to monomethyl phthalate and mono-n-butyl phthalate (M(n)BP) at the highest concentrations tested (300 μM). hPXR-V140M and hPXR-A370T exhibited patterns of phthalate responses similar to the wild-type receptor. By contrast, hPXR-D163G was unresponsive to all phthalate monoesters tested. Further studies revealed that hPXR-D163G did respond to rifampicin, but required approximately 40-fold higher concentrations than wild-type receptor, suggesting that the ligand-binding domain D163G variant has impaired ligand-binding activity. The responsiveness of PXR to activation by phthalate monoesters demonstrated here suggests that these ubiquitous environmental chemicals may, in part, exhibit their endocrine disruptor activities by altering PXR-regulated steroid hormone metabolism with potential adverse health effects in exposed individuals

  1. Investigating the Regulation of Estrogen Receptor-Mediated Transcription

    National Research Council Canada - National Science Library

    Thackray, Varykina

    2002-01-01

    ...-mediated regulation of specific target genes are still lacking. We have developed an estrogen responsive system in the fruit fly, Drosophila melanogaster in order to explore the functional interactions between ER and other cellular proteins...

  2. Investigating the Regulation of Estrogen Receptor-Mediated Transcription

    National Research Council Canada - National Science Library

    Thackray, Varykina

    2001-01-01

    ...-mediated regulation of specific target genes are still lacking. We have developed an estrogen responsive system in the fruit fly, Drosophila melanogaster in order to explore the functional interactions between ER and other cellular proteins...

  3. Androgen Receptor-Mediated Escape Mechanisms from Androgen Ablation Therapy

    Science.gov (United States)

    2005-10-01

    pathway independent of phosphatidylinositol 3’-kinase and Akt/protein kinase B. Cancer Res 59:1449-53. 158. Manin M, Baron S, Goossens K, et al. 2002...thyroid hormone-stimulated cell proliferation. Mol Cell Biol 21:6626-39 189. Richard -Parpaillon L, Heligon C, Chesnel F, Boujard D, Philpott A 2002 The IGF

  4. Erythropoietin's Beta Common Receptor Mediates Neuroprotection in Spinal Cord Neurons.

    Science.gov (United States)

    Foley, Lisa S; Fullerton, David A; Mares, Joshua; Sungelo, Mitchell; Weyant, Michael J; Cleveland, Joseph C; Reece, T Brett

    2017-12-01

    Paraplegia from spinal cord ischemia-reperfusion (SCIR) remains an elusive and devastating complication of complex aortic operations. Erythropoietin (EPO) attenuates this injury in models of SCIR. Upregulation of the EPO beta common receptor (βcR) is associated with reduced damage in models of neural injury. The purpose of this study was to examine whether EPO-mediated neuroprotection was dependent on βcR expression. We hypothesized that spinal cord neurons subjected to oxygen-glucose deprivation would mimic SCIR injury in aortic surgery and EPO treatment attenuates this injury in a βcR-dependent fashion. Lentiviral vectors with βcR knockdown sequences were tested on neuron cell cultures. The virus with greatest βcR knockdown was selected. Spinal cord neurons from perinatal wild-type mice were harvested and cultured to maturity. They were treated with knockdown or nonsense virus and transduced cells were selected. Three groups (βcR knockdown virus, nonsense control virus, no virus control; n = 8 each) were subjected to 1 hour of oxygen-glucose deprivation. Viability was assessed. βcR expression was quantified by immunoblot. EPO preserved neuronal viability after oxygen-glucose deprivation (0.82 ± 0.04 versus 0.61 ± 0.01; p neuron preservation was similar in the nonsense virus and control mice (0.82 ± 0.04 versus 0.80 ± 0.05; p = 0.77). EPO neuron preservation was lost in βcR knockdown mice compared with nonsense control mice (0.46 ± 0.03 versus 0.80 ± 0.05; p neuronal loss after oxygen-glucose deprivation in a βcR-dependent fashion. This receptor holds immense clinical promise as a target for pharmacotherapies treating spinal cord ischemic injury. Copyright © 2017 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.

  5. Study of multidrug resistance and radioresistance

    International Nuclear Information System (INIS)

    Kang, Yoon Koo; Yoo, Young Do

    1999-04-01

    We investigated the mechanism of 5-FU, adriamycin, radiation resistance in Korean gastric cancer cells. First we investigated the relation between Rb and multidrug resistance. Rb stable transfectants exhibited 5- to 10- fold more resistance to adriamycin than the control cells. These Rb transfectants showed increased MDR1 expression. We also investigated up-regulation in radiation-resistant tumor tissues. HSP27, MRP-8, GST, and NKEF-B were up-regulated in radiation resistant tumor. Expression of NKEF-B was also increased by radiation exposure in Head and Neck cells. These results demonstrated that NKEF-B is a stress response protein and it may have an important role in radiation resistance

  6. Induction of transient radioresistance in human erythrocytes

    International Nuclear Information System (INIS)

    Krokosz, Anita; Szweda-Lewandowska, Zofia

    2006-01-01

    Human erythrocytes suspended in an isotonic Na-phosphate buffer, pH 7.4 (hematocrit of 2%), were irradiated with γ-rays with single and split doses under air or N 2 O in order to determine the physicochemical changes caused by the dose inducing an increase in resistance to radiation-induced hemolysis. The obtained results showed that under the applied irradiation conditions, the dose of 0.4 kGy induced changes in erythrocytes, which were responsible for temporary resistance of erythrocytes to hemolysis. We concluded that the observed resistance is caused mainly by the structural changes in proteins

  7. Autophagy contributes to resistance of tumor cells to ionizing radiation.

    Science.gov (United States)

    Chaachouay, Hassan; Ohneseit, Petra; Toulany, Mahmoud; Kehlbach, Rainer; Multhoff, Gabriele; Rodemann, H Peter

    2011-06-01

    Autophagy signaling is a novel important target to improve anticancer therapy. To study the role of autophagy on resistance of tumor cells to ionizing radiation (IR), breast cancer cell lines differing in their intrinsic radiosensitivity were used. Breast cancer cell lines MDA-MB-231 and HBL-100 were examined with respect to clonogenic cell survival and induction of autophagy after radiation exposure and pharmacological interference of the autophagic process. As marker for autophagy the appearance of LC3-I and LC3-II proteins was analyzed by SDS-PAGE and Western blotting. Formation of autophagic vacuoles was monitored by immunofluorescence staining of LC3. LC3-I and LC3-II formation differs markedly in radioresistant MDA-MB-231 versus radiosensitive HBL-100 cells. Western blot analyses of LC3-II/LC3-I ratio indicated marked induction of autophagy by IR in radioresistant MDA-MB-231 cells, but not in radiosensitive HBL-100 cells. Indirect immunofluorescence analysis of LC3-II positive vacuoles confirmed this differential effect. Pre-treatment with 3-methyladenine (3-MA) antagonized IR-induced autophagy. Likewise, pretreatment of radioresistant MDA-231 cells with autophagy inhibitors 3-MA or chloroquine (CQ) significantly reduced clonogenic survival of irradiated cells. Our data clearly indicate that radioresistant breast tumor cells show a strong post-irradiation induction of autophagy, which thus serves as a protective and pro-survival mechanism in radioresistance. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  8. The radiobiological response of the thyroid

    International Nuclear Information System (INIS)

    O'Connor, M.K.; Malone, J.F.; Moriarty, M.; Cullen, M.J.

    1980-01-01

    The response of sheep thyroid cells in culture to single doses of X or γ rays is described. In the absence of cellular proliferation the cells were unusually radioresistant, showing little sign of interphase death at doses up to 9 krad. The follicular morphology characteristic of thyroid cells in vivo was also very radioresistant. Iodide trapping was reduced to 50% of the control value by doses of the order of 2 krad. When proliferation was induced the cells could be assayed for post-irradiation survival using a clonogenic endpoint. The survival curves were sigmoid with a Do of 410 rad and a very low extrapolation number. (author)

  9. Hindbrain GLP-1 receptor mediation of cisplatin-induced anorexia and nausea.

    Science.gov (United States)

    De Jonghe, Bart C; Holland, Ruby A; Olivos, Diana R; Rupprecht, Laura E; Kanoski, Scott E; Hayes, Matthew R

    2016-01-01

    While chemotherapy-induced nausea and vomiting are clinically controlled in the acute (anorexia, nausea, fatigue, and other illness-type behaviors during the delayed phase (>24 h) of chemotherapy are largely uncontrolled. As the hindbrain glucagon-like peptide-1 (GLP-1) system contributes to energy balance and mediates aversive and stressful stimuli, here we examine the hypothesis that hindbrain GLP-1 signaling mediates aspects of chemotherapy-induced nausea and reductions in feeding behavior in rats. Specifically, hindbrain GLP-1 receptor (GLP-1R) blockade, via 4th intracerebroventricular (ICV) exendin-(9-39) injections, attenuates the anorexia, body weight reduction, and pica (nausea-induced ingestion of kaolin clay) elicited by cisplatin chemotherapy during the delayed phase (48 h) of chemotherapy-induced nausea. Additionally, the present data provide evidence that the central GLP-1-producing preproglucagon neurons in the nucleus tractus solitarius (NTS) of the caudal brainstem are activated by cisplatin during the delayed phase of chemotherapy-induced nausea, as cisplatin led to a significant increase in c-Fos immunoreactivity in NTS GLP-1-immunoreactive neurons. These data support a growing body of literature suggesting that the central GLP-1 system may be a potential pharmaceutical target for adjunct anti-emetics used to treat the delayed-phase of nausea and emesis, anorexia, and body weight loss that accompany chemotherapy treatments. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. Bovine lactoferrin counteracts Toll-like receptor mediated activation signals in antigen presenting cells.

    Directory of Open Access Journals (Sweden)

    Patrizia Puddu

    Full Text Available Lactoferrin (LF, a key element in mammalian immune system, plays pivotal roles in host defence against infection and excessive inflammation. Its protective effects range from direct antimicrobial activities against a large panel of microbes, including bacteria, viruses, fungi and parasites, to antinflammatory and anticancer activities. In this study, we show that monocyte-derived dendritic cells (MD-DCs generated in the presence of bovine LF (bLF fail to undergo activation by up-modulating CD83, co-stimulatory and major histocompatibility complex molecules, and cytokine/chemokine secretion. Moreover, these cells are weak activators of T cell proliferation and retain antigen uptake activity. Consistent with an impaired maturation, bLF-MD-DC primed T lymphocytes exhibit a functional unresponsiveness characterized by reduced expression of CD154 and impaired expression of IFN-γ and IL-2. The observed imunosuppressive effects correlate with an increased expression of molecules with negative regulatory functions (i.e. immunoglobulin-like transcript 3 and programmed death ligand 1, indoleamine 2,3-dioxygenase, and suppressor of cytokine signaling-3. Interestingly, bLF-MD-DCs produce IL-6 and exhibit constitutive signal transducer and activator of transcription 3 activation. Conversely, bLF exposure of already differentiated MD-DCs completely fails to induce IL-6, and partially inhibits Toll-like receptor (TLR agonist-induced activation. Cell-specific differences in bLF internalization likely account for the distinct response elicited by bLF in monocytes versus immature DCs, providing a mechanistic base for its multiple effects. These results indicate that bLF exerts a potent anti-inflammatory activity by skewing monocyte differentiation into DCs with impaired capacity to undergo activation and to promote Th1 responses. Overall, these bLF-mediated effects may represent a strategy to block excessive DC activation upon TLR-induced inflammation, adding further evidence for a critical role of bLF in directing host immune function.

  11. Effect of lipopolysaccharide and antidepressant drugs on glucocorticoid receptor-mediated gene transcription

    Czech Academy of Sciences Publication Activity Database

    Budziszewska, B.; Basta-Kaim, A.; Kubera, M.; Jaworska, L.; Leskiewicz, M.; Tetich, M.; Otczyk, M.; Zajícová, Alena; Holáň, Vladimír; Lasoń, W.

    2005-01-01

    Roč. 57, č. 4 (2005), s. 540-544 ISSN 1734-1140 Grant - others:State Committee for Scientific Research (KBN)(PL) 6P05A076 Institutional research plan: CEZ:AV0Z5052915 Keywords : glucocorticoid receptor * antidepressant drugs * interleukin-6 Subject RIV: EB - Genetics ; Molecular Biology

  12. Ultradian hormone stimulation induces glucocorticoid receptor-mediated pulses of gene transcription.

    Science.gov (United States)

    Stavreva, Diana A; Wiench, Malgorzata; John, Sam; Conway-Campbell, Becky L; McKenna, Mervyn A; Pooley, John R; Johnson, Thomas A; Voss, Ty C; Lightman, Stafford L; Hager, Gordon L

    2009-09-01

    Studies on glucocorticoid receptor (GR) action typically assess gene responses by long-term stimulation with synthetic hormones. As corticosteroids are released from adrenal glands in a circadian and high-frequency (ultradian) mode, such treatments may not provide an accurate assessment of physiological hormone action. Here we demonstrate that ultradian hormone stimulation induces cyclic GR-mediated transcriptional regulation, or gene pulsing, both in cultured cells and in animal models. Equilibrium receptor-occupancy of regulatory elements precisely tracks the ligand pulses. Nascent RNA transcripts from GR-regulated genes are released in distinct quanta, demonstrating a profound difference between the transcriptional programs induced by ultradian and constant stimulation. Gene pulsing is driven by rapid GR exchange with response elements and by GR recycling through the chaperone machinery, which promotes GR activation and reactivation in response to the ultradian hormone release, thus coupling promoter activity to the naturally occurring fluctuations in hormone levels. The GR signalling pathway has been optimized for a prompt and timely response to fluctuations in hormone levels, indicating that biologically accurate regulation of gene targets by GR requires an ultradian mode of hormone stimulation.

  13. VIP/PACAP receptor mediation of cutaneous active vasodilation during heat stress in humans.

    Science.gov (United States)

    Kellogg, Dean L; Zhao, Joan L; Wu, Yubo; Johnson, John M

    2010-07-01

    Vasoactive intestinal peptide (VIP) is implicated in cutaneous active vasodilation in humans. VIP and the closely related pituitary adenylate cyclase activating peptide (PACAP) act through several receptor types: VIP through VPAC1 and VPAC2 receptors and PACAP through VPAC1, VPAC2, and PAC1 receptors. We examined participation of VPAC2 and/or PAC1 receptors in cutaneous vasodilation during heat stress by testing the effects of their specific blockade with PACAP6-38. PACAP6-38 dissolved in Ringer's was administered by intradermal microdialysis at one forearm site while a control site received Ringer's solution. Skin blood flow was monitored by laser-Doppler flowmetry (LDF). Blood pressure was monitored noninvasively and cutaneous vascular conductance (CVC) calculated. A 5- to 10-min baseline period was followed by approximately 70 min of PACAP6-38 (100 microM) perfusion at one site in normothermia and a 3-min period of body cooling. Whole body heating was then performed to engage cutaneous active vasodilation and was maintained until CVC had plateaued at an elevated level at all sites for 5-10 min. Finally, 58 mM sodium nitroprusside was perfused through both microdialysis sites to effect maximal vasodilation. No CVC differences were found between control and PACAP6-38-treated sites during normothermia (19 +/- 3%max untreated vs. 20 +/- 3%max, PACAP6-38 treated; P > 0.05 between sites) or cold stress (11 +/- 2%max untreated vs. 10 +/- 2%max, PACAP6-38 treated, P > 0.05 between sites). PACAP6-38 attenuated the increase in CVC during whole body heating when compared with untreated sites (59 +/- 3%max untreated vs. 46 +/- 3%max, PACAP6-38 treated, P < 0.05). We conclude that VPAC2 and/or PAC1 receptor activation is involved in cutaneous active vasodilation in humans.

  14. β-Receptor-mediated increase in cerebral blood flow during hypoglycemia

    International Nuclear Information System (INIS)

    Hollinger, B.R.; Bryan, R.M.

    1987-01-01

    The authors tested the hypothesis that β-adrenergic receptor stimulation is involved with the increase in regional cerebral blood flow (rCBF) during hypoglycemia. Rats were surgically prepared with the use of halothane-nitrous oxide anesthesia. A plaster restraining cast was placed around the hindquarters, and anesthesia was discontinued. Hypoglycemia was produced by an intravenous injection of insulin; normoglycemic control rates were given saline. Propranolol was administered to some control and some hypoglycemic rats to block the β-adrenergic receptors. Regional CBF was measured using 4-[N-methyl- 14 C]iodoantipyrine. Regional CBF increased during hypoglycemia in rats that were not treated with propranolol. The increase varied from ∼60 to 200% depending on the brain region. During hypoglycemia, propranolol abolished the increase in rCBF in the hypothalamus, cerebellum, and pyramidal tract. In other regions the increase in rCBF was only 33-65% of the increase in hypoglycemic rats that were not treated with propranolol. They conclude that β-receptor stimulation plays a major role in the increase in rCBF during hypoglycemia

  15. Probenecid inhibits α-adrenergic receptor-mediated vasoconstriction in the human leg vasculature

    DEFF Research Database (Denmark)

    Nyberg, Michael Permin; Piil, Peter Bergmann; Kiehn, Oliver Thistrup

    2018-01-01

    to α1- and α2-adrenergic receptor stimulation in the human forearm and leg vasculature of young healthy male subjects (23±3 years). By use of immunolabeling and confocal microscopy, Panx1 channels were found to be expressed in vascular smooth muscle cells of arterioles in human leg skeletal muscle....... Probenecid treatment increased (Padrenergic receptor stimulation) by ≈15%, whereas the response to the α1-agonist phenylephrine was unchanged. Inhibition...

  16. β-Adrenergic receptor-mediated suppression of interleukin 2 receptors in human lymphocytes

    International Nuclear Information System (INIS)

    Feldman, R.D.; Hunninghake, G.W.; McArdle, W.L.

    1987-01-01

    Adrenergic receptor agonists are know to attenuate the proliferative response of human lymphocytes after activation; however, their mechanism of action is unknown. Since expression of interleukin 2 (IL-2) receptors is a prerequisite for proliferation, the effect of β-adrenergic receptor agonists on lymphocyte IL-2 receptors was studied on both mitogen-stimulated lymphocytes and IL-2-dependent T lymphocyte cell lines. In both cell types the β-adrenergic receptor agonist isoproterenol blocked the expression of IL-2 receptors, as determined with the IL-2 receptor anti-TAC antibody. To determine the effect of β-adrenergic agonists on expression of the high affinity IL-2 receptors, [ 125 I]IL-2 binding studies were performed at concentrations selective for high affinity sites. No significant effect of β-adrenergic agonists on high affinity IL-2 receptor sites could be detected. The data demonstrate that β-adrenergic receptor agonists down-regulate IL-2 receptors primarily affecting low affinity sites

  17. β2-adrenergic receptor-mediated negative regulation of group 2 innate lymphoid cell responses.

    Science.gov (United States)

    Moriyama, Saya; Brestoff, Jonathan R; Flamar, Anne-Laure; Moeller, Jesper B; Klose, Christoph S N; Rankin, Lucille C; Yudanin, Naomi A; Monticelli, Laurel A; Putzel, Gregory Garbès; Rodewald, Hans-Reimer; Artis, David

    2018-03-02

    The type 2 inflammatory response is induced by various environmental and infectious stimuli. Although recent studies identified group 2 innate lymphoid cells (ILC2s) as potent sources of type 2 cytokines, the molecular pathways controlling ILC2 responses are incompletely defined. Here we demonstrate that murine ILC2s express the β 2 -adrenergic receptor (β 2 AR) and colocalize with adrenergic neurons in the intestine. β 2 AR deficiency resulted in exaggerated ILC2 responses and type 2 inflammation in intestinal and lung tissues. Conversely, β 2 AR agonist treatment was associated with impaired ILC2 responses and reduced inflammation in vivo. Mechanistically, we demonstrate that the β 2 AR pathway is a cell-intrinsic negative regulator of ILC2 responses through inhibition of cell proliferation and effector function. Collectively, these data provide the first evidence of a neuronal-derived regulatory circuit that limits ILC2-dependent type 2 inflammation. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  18. Serotonin(2) receptors mediate respiratory recovery after cervical spinal cord hemisection in adult rats.

    Science.gov (United States)

    Zhou, S Y; Basura, G J; Goshgarian, H G

    2001-12-01

    The aim of the present study was to specifically investigate the involvement of serotonin [5-hydroxytryptamine (5-HT(2))] receptors in 5-HT-mediated respiratory recovery after cervical hemisection. Experiments were conducted on C(2) spinal cord-hemisected, anesthetized (chloral hydrate, 400 mg/kg ip), vagotomized, pancuronium- paralyzed, and artificially ventilated female Sprague-Dawley rats in which CO(2) levels were monitored and maintained. Twenty-four hours after spinal hemisection, the ipsilateral phrenic nerve displayed no respiratory-related activity indicative of a functionally complete hemisection. Intravenous administration of the 5-HT(2A/2C)-receptor agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI) induced respiratory-related activity in the phrenic nerve ipsilateral to hemisection under conditions in which CO(2) was maintained at constant levels and augmented the activity induced under conditions of hypercapnia. The effects of DOI were found to be dose dependent, and the recovery of activity could be maintained for up to 2 h after a single injection. DOI-induced recovery was attenuated by the 5-HT(2)-receptor antagonist ketanserin but not with the 5-HT(2C)-receptor antagonist RS-102221, suggesting that 5-HT(2A) and not necessarily 5-HT(2C) receptors may be involved in the induction of respiratory recovery after cervical spinal cord injury.

  19. Role of glucocorticoid receptor-mediated mechanisms in cocaine memory enhancement.

    Science.gov (United States)

    Stringfield, S J; Higginbotham, J A; Wang, R; Berger, A L; McLaughlin, R J; Fuchs, R A

    2017-09-01

    The basolateral amygdala (BLA) is a critical site for the reconsolidation of labile contextual cocaine memories following retrieval-induced reactivation/destabilization. Here, we examined whether glucocorticoid receptors (GR), which are abundant in the BLA, mediate this phenomenon. Rats were trained to lever press for cocaine reinforcement in a distinct environmental context, followed by extinction training in a different context. Rats were then briefly exposed to the cocaine-paired context (to elicit memory reactivation and reconsolidation) or their home cages (no reactivation control). Exposure to the cocaine-paired context elicited greater serum corticosterone concentrations than home cage stay. Interestingly, the GR antagonist, mifepristone (3-10 ng/hemisphere), administered into the BLA after memory reactivation produced a further, dose-dependent increase in serum corticosterone concentrations during the putative time of cocaine-memory reconsolidation but produced an inverted U-shaped dose-effect curve on subsequent cocaine-seeking behavior 72 h later. This effect was anatomically selective, dependent on memory reactivation (i.e., not observed after home cage exposure), and did not reflect protracted hyperactivity. However, the effect was also observed when mifepristone was administered after novelty stress that mimics drug context-induced hypothalamic-pituitary-adrenal (HPA) axis activation without explicit memory reactivation. Together, these findings suggest that, similar to explicit memory retrieval, a stressful event is sufficient to destabilize cocaine memories and permit their manipulation. Furthermore, BLA GR stimulation exerts inhibitory feedback upon HPA axis activation and thus suppresses cocaine-memory reconsolidation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. High Throughput Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale L

    2005-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  1. High Throughout Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale

    2003-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  2. High Throughput Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale

    2004-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  3. Molecular and Biochemical Effects of a Kola Nut Extract on Androgen Receptor-Mediated Pathways

    Directory of Open Access Journals (Sweden)

    Rajasree Solipuram

    2009-01-01

    Full Text Available The low incidence of prostate cancer in Asians has been attributed to chemopreventative properties of certain chemicals found in their diet. This study characterized the androgenic and chemopreventative properties of the Jamaican bush tea “Bizzy,” using androgen receptor positive and negative cell lines. Exposure of prostate cells to Biz-2 resulted in a growth inhibition (GI50 of 15 ppm in LNCaP cells and 3.6 ppm in DU145 cells. Biz-2 elicited a 2-fold increase in the mRNA of the anti-apoptotic gene Bcl2, with a 10-fold increase in that of the proapoptotic gene Bax. We observed a 2.4- to 7.5-fold change in apoptotic cells in both cell lines. Biz-2 at 10 ppm elicited a time- and dose-dependent stimulation of both the protein and mRNA levels of several androgen-regulated genes. Biz-2 caused a 36% decrease in PSA secretion and a significant increase in PSA mRNA. The relative binding affinity (IC50 of Biz-2 for AR was 2- to 5-fold lower than that of the synthetic androgen R1881. Biz-2 was found to be a specific ligand for the AR in that the natural ligand, DHT, and the anti-androgen, flutamide, displaced Biz-2 bound to AR and inhibited Biz-2-induced transcription and PSA secretion. This study provided evidence that Biz-2 extract possesses the ability to modulate prostate cancer cell biology in an AR-dependent manner.

  4. Kappa Opioid Receptors Mediate where Fear Is Expressed Following Extinction Training

    Science.gov (United States)

    Cole, Sindy; Richardson, Rick; McNally, Gavan P.

    2011-01-01

    Six experiments used a within-subjects renewal design to examine the involvement of kappa opioid receptors (KORs) in regulating the expression and recovery of extinguished fear. Rats were trained to fear a tone conditioned stimulus (CS) via pairings with foot shock in a distinctive context (A). This was followed by extinction training of the CS in…

  5. The CRF Family of Neuropeptides and their Receptors - Mediators of the Central Stress Response

    Science.gov (United States)

    Dedic, Nina; Chen, Alon; Deussing, Jan M.

    2018-01-01

    Background: Dysregulated stress neurocircuits, caused by genetic and/or environmental changes, underlie the development of many neuropsychiatric disorders. Corticotropin-releasing factor (CRF) is the major physiological activator of the hypothalamic-pituitary-adrenal (HPA) axis and conse-quently a primary regulator of the mammalian stress response. Together with its three family members, urocortins (UCNs) 1, 2, and 3, CRF integrates the neuroendocrine, autonomic, metabolic and behavioral responses to stress by activating its cognate receptors CRFR1 and CRFR2. Objective: Here we review the past and current state of the CRF/CRFR field, ranging from pharmacologi-cal studies to genetic mouse models and virus-mediated manipulations. Results: Although it is well established that CRF/CRFR1 signaling mediates aversive responses, includ-ing anxiety and depression-like behaviors, a number of recent studies have challenged this viewpoint by revealing anxiolytic and appetitive properties of specific CRF/CRFR1 circuits. In contrast, the UCN/CRFR2 system is less well understood and may possibly also exert divergent functions on physiol-ogy and behavior depending on the brain region, underlying circuit, and/or experienced stress conditions. Conclusion: A plethora of available genetic tools, including conventional and conditional mouse mutants targeting CRF system components, has greatly advanced our understanding about the endogenous mecha-nisms underlying HPA system regulation and CRF/UCN-related neuronal circuits involved in stress-related behaviors. Yet, the detailed pathways and molecular mechanisms by which the CRF/UCN-system translates negative or positive stimuli into the final, integrated biological response are not completely un-derstood. The utilization of future complementary methodologies, such as cell-type specific Cre-driver lines, viral and optogenetic tools will help to further dissect the function of genetically defined CRF/UCN neurocircuits in the context of adaptive and maladaptive stress responses. PMID:28260504

  6. Astrocytic β2-adrenergic receptors mediate hippocampal long-term memory consolidation

    KAUST Repository

    Gao, Virginia; Suzuki, Akinobu; Magistretti, Pierre J.; Lengacher, Sylvain; Pollonini, Gabriella; Steinman, Michael Q.; Alberini, Cristina M.

    2016-01-01

    Emotionally relevant experiences form strong and long-lasting memories by critically engaging the stress hormone/neurotransmitter noradrenaline, which mediates and modulates the consolidation of these memories. Noradrenaline acts through adrenergic receptors (ARs), of which β2- Adrenergic receptors (βARs) are of particular importance. The differential anatomical and cellular distribution of βAR subtypes in the brain suggests that they play distinct roles in memory processing, although much about their specific contributions and mechanisms of action remains to be understood. Here we show that astrocytic rather than neuronal β2ARs in the hippocampus play a key role in the consolidation of a fear-based contextual memory. These hippocampal β2ARs, but not β1ARs, are coupled to the training-dependent release of lactate from astrocytes, which is necessary for long- Term memory formation and for underlying molecular changes. This key metabolic role of astrocytic β2ARs may represent a novel target mechanism for stress-related psychopathologies and neurodegeneration.

  7. Rab GTPases Regulate Endothelial Cell Protein C Receptor-Mediated Endocytosis and Trafficking of Factor VIIa

    Science.gov (United States)

    Nayak, Ramesh C.; Keshava, Shiva; Esmon, Charles T.; Pendurthi, Usha R.; Rao, L. Vijaya Mohan

    2013-01-01

    Recent studies have established that factor VIIa (FVIIa) binds to the endothelial cell protein C receptor (EPCR). FVIIa binding to EPCR may promote the endocytosis of this receptor/ligand complex. Rab GTPases are known to play a crucial role in the endocytic and exocytic pathways of receptors or receptor/ligand complexes. The present study was undertaken to investigate the role of Rab GTPases in the intracellular trafficking of EPCR and FVIIa. CHO-EPCR cells and human umbilical vein endothelial cells (HUVEC) were transduced with recombinant adenoviral vectors to express wild-type, constitutively active, or dominant negative mutant of various Rab GTPases. Cells were exposed to FVIIa conjugated with AF488 fluorescent probe (AF488-FVIIa), and intracellular trafficking of FVIIa, EPCR, and Rab proteins was evaluated by immunofluorescence confocal microscopy. In cells expressing wild-type or constitutively active Rab4A, internalized AF488-FVIIa accumulated in early/sorting endosomes and its entry into the recycling endosomal compartment (REC) was inhibited. Expression of constitutively active Rab5A induced large endosomal structures beneath the plasma membrane where EPCR and FVIIa accumulated. Dominant negative Rab5A inhibited the endocytosis of EPCR-FVIIa. Expression of constitutively active Rab11 resulted in retention of accumulated AF488-FVIIa in the REC, whereas expression of a dominant negative form of Rab11 led to accumulation of internalized FVIIa in the cytoplasm and prevented entry of internalized FVIIa into the REC. Expression of dominant negative Rab11 also inhibited the transport of FVIIa across the endothelium. Overall our data show that Rab GTPases regulate the internalization and intracellular trafficking of EPCR-FVIIa. PMID:23555015

  8. Probing multivalency in ligand–receptor-mediated adhesion of soft, biomimetic interfaces

    Directory of Open Access Journals (Sweden)

    Stephan Schmidt

    2015-05-01

    Full Text Available Many biological functions at cell level are mediated by the glycocalyx, a dense carbohydrate-presenting layer. In this layer specific interactions between carbohydrate ligands and protein receptors are formed to control cell–cell recognition, cell adhesion and related processes. The aim of this work is to shed light on the principles of complex formation between surface anchored carbohydrates and receptor surfaces by measuring the specific adhesion between surface bound mannose on a concanavalin A (ConA layer via poly(ethylene glycol-(PEG-based soft colloidal probes (SCPs. Special emphasis is on the dependence of multivalent presentation and density of carbohydrate units on specific adhesion. Consequently, we first present a synthetic strategy that allows for controlled density variation of functional groups on the PEG scaffold using unsaturated carboxylic acids (crotonic acid, acrylic acid, methacrylic acid as grafting units for mannose conjugation. We showed by a range of analytic techniques (ATR–FTIR, Raman microscopy, zeta potential and titration that this synthetic strategy allows for straightforward variation in grafting density and grafting length enabling the controlled presentation of mannose units on the PEG network. Finally we determined the specific adhesion of PEG-network-conjugated mannose units on ConA surfaces as a function of density and grafting type. Remarkably, the results indicated the absence of a molecular-level enhancement of mannose/ConA interaction due to chelate- or subsite-binding. The results seem to support the fact that weak carbohydrate interactions at mechanically flexible interfaces hardly undergo multivalent binding but are simply mediated by the high number of ligand–receptor interactions.

  9. Basolateral amygdala GABA-A receptors mediate stress-induced memory retrieval impairment in rats.

    Science.gov (United States)

    Sardari, Maryam; Rezayof, Ameneh; Khodagholi, Fariba; Zarrindast, Mohammad-Reza

    2014-04-01

    The present study was designed to investigate the involvement of GABA-A receptors of the basolateral amygdala (BLA) in the impairing effect of acute stress on memory retrieval. The BLAs of adult male Wistar rats were bilaterally cannulated and memory retrieval was measured in a step-through type passive avoidance apparatus. Acute stress was evoked by placing the animals on an elevated platform for 10, 20 and 30 min. The results indicated that exposure to 20 and 30 min stress, but not 10 min, before memory retrieval testing (pre-test exposure to stress) decreased the step-through latency, indicating stress-induced memory retrieval impairment. Intra-BLA microinjection of a GABA-A receptor agonist, muscimol (0.005-0.02 μg/rat), 5 min before exposure to an ineffective stress (10 min exposure to stress) induced memory retrieval impairment. It is important to note that pre-test intra-BLA microinjection of the same doses of muscimol had no effect on memory retrieval in the rats unexposed to 10 min stress. The blockade of GABA-A receptors of the BLA by injecting an antagonist, bicuculline (0.4-0.5 μg/rat), 5 min before 20 min exposure to stress, prevented stress-induced memory retrieval. Pre-test intra-BLA microinjection of the same doses of bicuculline (0.4-0.5 μg/rat) in rats unexposed to 20 min stress had no effect on memory retrieval. In addition, pre-treatment with bicuculline (0.1-0.4 μg/rat, intra-BLA) reversed muscimol (0.02 μg/rat, intra-BLA)-induced potentiation on the effect of stress in passive avoidance learning. It can be concluded that pre-test exposure to stress can induce memory retrieval impairment and the BLA GABA-A receptors may be involved in stress-induced memory retrieval impairment.

  10. Insulin receptors mediate growth effects in cultured fetal neurons. I. Rapid stimulation of protein synthesis

    International Nuclear Information System (INIS)

    Heidenreich, K.A.; Toledo, S.P.

    1989-01-01

    In this study we have examined the effects of insulin on protein synthesis in cultured fetal chick neurons. Protein synthesis was monitored by measuring the incorporation of [3H]leucine (3H-leu) into trichloroacetic acid (TCA)-precipitable protein. Upon addition of 3H-leu, there was a 5-min lag before radioactivity occurred in protein. During this period cell-associated radioactivity reached equilibrium and was totally recovered in the TCA-soluble fraction. After 5 min, the incorporation of 3H-leu into protein was linear for 2 h and was inhibited (98%) by the inclusion of 10 micrograms/ml cycloheximide. After 24 h of serum deprivation, insulin increased 3H-leu incorporation into protein by approximately 2-fold. The stimulation of protein synthesis by insulin was dose dependent (ED50 = 70 pM) and seen within 30 min. Proinsulin was approximately 10-fold less potent than insulin on a molar basis in stimulating neuronal protein synthesis. Insulin had no effect on the TCA-soluble fraction of 3H-leu at any time and did not influence the uptake of [3H]aminoisobutyric acid into neurons. The isotope ratio of 3H-leu/14C-leu in the leucyl tRNA pool was the same in control and insulin-treated neurons. Analysis of newly synthesized proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that insulin uniformly increased the incorporation of 14C-leu into all of the resolved neuronal proteins. We conclude from these data that (1) insulin rapidly stimulates overall protein synthesis in fetal neurons independent of amino acid uptake and aminoacyl tRNA precursor pools; (2) stimulation of protein synthesis is mediated by the brain subtype of insulin receptor; and (3) insulin is potentially an important in vivo growth factor for fetal central nervous system neurons

  11. Pattern recognition receptor-mediated cytokine response in infants across 4 continents.

    Science.gov (United States)

    Smolen, Kinga K; Ruck, Candice E; Fortuno, Edgardo S; Ho, Kevin; Dimitriu, Pedro; Mohn, William W; Speert, David P; Cooper, Philip J; Esser, Monika; Goetghebuer, Tessa; Marchant, Arnaud; Kollmann, Tobias R

    2014-03-01

    Susceptibility to infection as well as response to vaccination varies among populations. To date, the underlying mechanisms responsible for these clinical observations have not been fully delineated. Because innate immunity instructs adaptive immunity, we hypothesized that differences between populations in innate immune responses may represent a mechanistic link to variation in susceptibility to infection or response to vaccination. Determine whether differences in innate immune responses exist among infants from different continents of the world. We determined the innate cytokine response following pattern recognition receptor (PRR) stimulation of whole blood from 2-year-old infants across 4 continents (Africa, North America, South America, and Europe). We found that despite the many possible genetic and environmental exposure differences in infants across 4 continents, innate cytokine responses were similar for infants from North America, South America, and Europe. However, cells from South African infants secreted significantly lower levels of cytokines than did cells from infants from the 3 other sites, and did so following stimulation of extracellular and endosomal but not cytosolic PRRs. Substantial differences in innate cytokine responses to PRR stimulation exist among different populations of infants that could not have been predicted. Delineating the underlying mechanism(s) for these differences will not only aid in improving vaccine-mediated protection but possibly also provide clues for the susceptibility to infection in different regions of the world. Copyright © 2013 The Authors. Published by Mosby, Inc. All rights reserved.

  12. Scavenger receptor-mediated endocytosis by sinusoidal cells in rat bone marrow

    International Nuclear Information System (INIS)

    Geoffroy, J.S.

    1987-01-01

    Endocytosis of serum albumin by sinusoidal endothelial cells in rat bone marrow was investigated initially at the ultrastructural level with subsequent biochemical investigation of the specificity mediating this event. Bovine serum albumin adsorbed to 20nm colloidal gold particles (AuBSA) was chosen as the electron microscopic probe. Morphological data strongly suggested that a receptor was involved in uptake of AuBSA. Confirmation of receptor involvement in the uptake of AuBSA by marrow sinusoidal endothelial cells was achieved utilizing an in situ isolated hind limb perfusion protocol in conjunction with unlabeled, radiolabeled, and radio-/colloidal gold labeled probes. The major findings of competition and saturation experiments were: (1) endocytosis of AuBSA was mediated by a receptor for modified/treated serum albumin; (2) endocytosis of formaldehyde-treated serum albumin was mediated by a binding site which may be the same or closely related to the site responsible for the uptake of AuBSA; and (3) endocytosis of native untreated albumin was not mediated by receptor and probably represents fluid-phase pinocitosis

  13. Presynaptic CRF1 Receptors Mediate the Ethanol Enhancement of GABAergic Transmission in the Mouse Central Amygdala

    Directory of Open Access Journals (Sweden)

    Zhiguo Nie

    2009-01-01

    Full Text Available Corticotropin-releasing factor (CRF is a 41-amino-acid neuropeptide involved in stress responses initiated from several brain areas, including the amygdala formation. Research shows a strong relationship between stress, brain CRF, and excessive alcohol consumption. Behavioral studies suggest that the central amygdala (CeA is significantly involved in alcohol reward and dependence. We recently reported that the ethanol augmentation of GABAergic synaptic transmission in rat CeA involves CRF1 receptors, because both CRF and ethanol significantly enhanced the amplitude of evoked GABAergic inhibitory postsynaptic currents (IPSCs in CeA neurons from wild-type (WT and CRF2 knockout (KO mice, but not in neurons of CRF1 KO mice. The present study extends these findings using selective CRF receptor ligands, gene KO models, and miniature IPSC (mIPSC analysis to assess further a presynaptic role for the CRF receptors in mediating ethanol effects in the CeA. In whole-cell patch recordings of pharmacologically isolated GABAAergic IPSCs from slices of mouse CeA, both CRF and ethanol augmented evoked IPSCs in a concentration-dependent manner, with low EC50s. A CRF1 (but not CRF2 KO construct and the CRF1-selective nonpeptide antagonist NIH-3 (LWH-63 blocked the augmenting effect of both CRF and ethanol on evoked IPSCs. Furthermore, the new selective CRF1 agonist stressin1, but not the CRF2 agonist urocortin 3, also increased evoked IPSC amplitudes. Both CRF and ethanol decreased paired-pulse facilitation (PPF of evoked IPSCs and significantly enhanced the frequency, but not the amplitude, of spontaneous miniature GABAergic mIPSCs in CeA neurons of WT mice, suggesting a presynaptic site of action. The PPF effect of ethanol was abolished in CeA neurons of CRF1 KO mice. The CRF1 antagonist NIH-3 blocked the CRF- and ethanol-induced enhancement of mIPSC frequency in CeA neurons. These data indicate that presynaptic CRF1 receptors play a critical role in permitting or mediating ethanol enhancement of GABAergic synaptic transmission in CeA, via increased vesicular GABA release, and thus may be a rational target for the treatment of alcohol abuse and alcoholism.

  14. Bradykinin B2 receptor-mediated phosphoinositide hydrolysis in bovine cultured tracheal smooth muscle cells.

    OpenAIRE

    Marsh, K. A.; Hill, S. J.

    1992-01-01

    1. Bovine tracheal smooth muscle cells were established in culture to study agonist-induced phosphoinositide (PI) hydrolysis in this tissue. 2. Bradykinin (0.1 nM-10 microM) evoked a concentration-dependent increase (log EC50 (M) = -9.4 +/- 0.2; n = 8) in the accumulation of total [3H]-inositol phosphates in cultured tracheal smooth muscle cells whereas the selective B1 receptor agonist des-Arg9-bradykinin (10 microM) was significantly less effective (16% of bradykinin maximal response; relat...

  15. Hippocampal GluA1-containing AMPA receptors mediate context-dependent sensitization to morphine.

    Science.gov (United States)

    Xia, Yan; Portugal, George S; Fakira, Amanda K; Melyan, Zara; Neve, Rachael; Lee, H Thomas; Russo, Scott J; Liu, Jie; Morón, Jose A

    2011-11-09

    Glutamatergic systems, including AMPA receptors (AMPARs), are involved in opiate-induced neuronal and behavioral plasticity, although the mechanisms underlying these effects are not fully understood. In the present study, we investigated the effects of repeated morphine administration on AMPAR expression, synaptic plasticity, and context-dependent behavioral sensitization to morphine. We found that morphine treatment produced changes of synaptic AMPAR expression in the hippocampus, a brain area that is critically involved in learning and memory. These changes could be observed 1 week after the treatment, but only when mice developed context-dependent behavioral sensitization to morphine in which morphine treatment was associated with drug administration environment. Context-dependent behavioral sensitization to morphine was also associated with increased basal synaptic transmission and disrupted hippocampal long-term potentiation (LTP), whereas these effects were less robust when morphine administration was not paired with the drug administration environment. Interestingly, some effects may be related to the prior history of morphine exposure in the drug-associated environment, since alterations of AMPAR expression, basal synaptic transmission, and LTP were observed in mice that received a saline challenge 1 week after discontinuation of morphine treatment. Furthermore, we demonstrated that phosphorylation of GluA1 AMPAR subunit plays a critical role in the acquisition and expression of context-dependent behavioral sensitization, as this behavior is blocked by a viral vector that disrupts GluA1 phosphorylation. These data provide evidence that glutamatergic signaling in the hippocampus plays an important role in context-dependent sensitization to morphine and supports further investigation of glutamate-based strategies for treating opiate addiction.

  16. Hippocampal GluA1-containing AMPA receptors mediate context-dependent sensitization to morphine

    OpenAIRE

    Xia, Yan; Portugal, George S.; Fakira, Amanda K.; Melyan, Zara; Neve, Rachael; Lee, H. Thomas; Russo, Scott J.; Liu, Jie; Morón, Jose A.

    2011-01-01

    Glutamatergic systems, including α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) are involved in opiate-induced neuronal and behavioral plasticity, although the mechanisms underlying these effects are not fully understood. In the present study, we investigated the effects of repeated morphine administration on AMPAR expression, synaptic plasticity, and context-dependent behavioral sensitization to morphine. We found that morphine treatment produced changes of synaptic...

  17. Alpha7 nicotinic receptor mediated protection against ethanol-induced cytotoxicity in PC12 cells.

    Science.gov (United States)

    Li, Y; King, M A; Grimes, J; Smith, N; de Fiebre, C M; Meyer, E M

    1999-01-16

    Ethanol caused a concentration-dependent loss of PC12 cells over a 24 h interval, accompanied by an increase in intracellular calcium. The specific alpha7 nicotinic receptor partial agonist DMXB attenuated both of these ethanol-induced actions at a concentration (3 microM) found previously to protect against apoptotic and necrotic cell loss. The alpha7 nicotinic receptor antagonist methylylaconitine blocked the neuroprotective action of DMXB when applied with but not 30 min after the agonist. These results indicate that activation of alpha7 nicotinic receptors may be therapeutically useful in preventing ethanol-neurotoxicity. Copyright 1999 Elsevier Science B.V.

  18. Alpha 7 nicotinic acetylcholine receptor-mediated protection against ethanol-induced neurotoxicity.

    Science.gov (United States)

    de Fiebre, NancyEllen C; de Fiebre, Christopher M

    2003-11-01

    The alpha(7)-selective nicotinic partial agonist 3-[2,4-dimethoxybenzylidene]anabaseine (DMXB) was examined for its ability to modulate ethanol-induced neurotoxicity in primary cultures of rat neurons. Primary cultures of hippocampal neurons were established from Long-Evans, embryonic day (E)-18 rat fetuses and maintained for 7 days. Ethanol (0-150 mM), DMXB (0-56 microM), or both were subsequently co-applied to cultures. Ethanol was added two additional times to the cultures to compensate for evaporation. After 5 days, neuronal viability was assessed with the MTT cell proliferation assay. Results demonstrated that ethanol reduces neuronal viability in a concentration-dependent fashion and that DMXB protects against this ethanol-induced neurotoxicity, also in a concentration-dependent fashion. These results support the suggestion that nicotinic partial agonists may be useful in treating binge drinking-induced neurotoxicity and may provide clues as to why heavy drinkers are usually smokers.

  19. Intrinsically disordered cytoplasmic domains of two cytokine receptors mediate conserved interactions with membranes

    DEFF Research Database (Denmark)

    Haxholm, Gitte Wolfsberg; Nikolajsen, Louise Fletcher; Olsen, Johan Gotthardt

    2015-01-01

    . This study presents the first comprehensive structural characterization of any cytokine receptor ICD and demonstrates that the human prolactin and growth hormone receptor ICDs are intrinsically disordered throughout their entire lengths. We show that they interact specifically with hallmark lipids...

  20. GABA(A) receptors mediate orexin-A induced stimulation of food intake.

    Science.gov (United States)

    Kokare, Dadasaheb M; Patole, Angad M; Carta, Anna; Chopde, Chandrabhan T; Subhedar, Nishikant K

    2006-01-01

    Although the role of orexins in sleep/wake cycle and feeding behavior is well established, underlying mechanisms have not been fully understood. An attempt has been made to investigate the role of GABA(A) receptors and their benzodiazepine site on the orexin-A induced response to feeding. Different groups of rats were food deprived overnight and next day injected intracerebroventricularly (icv) with vehicle (artificial CSF; 5 microl/rat) or orexin-A (20-50 nM/rat) and the animals were given free access to food. Cumulative food intake was measured during light phase of light/dark cycle at 1-, 2-, 4- and 6-h post-injection time points. Orexin-A (30-50 nM/rat, icv) stimulated food intake at all the time points (P GABA(A) receptor agonists muscimol (25 ng/rat, icv) and diazepam (0.5 mg/kg, ip) at subeffective doses significantly potentiated the hyperphagic effect of orexin-A (30 nM/rat, icv). However, the effect was negated by the GABA(A) receptor antagonist bicuculline (1 mg/kg, ip). Interestingly, benzodiazepine receptor antagonist flumazenil (5 ng/rat, icv), augmented the orexin-A (30 nM/rat, icv) induced hyperphagia; the effect may be attributed to the intrinsic activity of the agent. The results suggest that the hyperphagic effect of orexin-A, at least in part, is mediated by enhanced GABA(A) receptor activity.

  1. Acupuncture suppresses intravenous methamphetamine self-administration through GABA receptor's mediation.

    Science.gov (United States)

    Choi, Yi Jeong; Kim, Nam Jun; Zhao, Rong Jie; Kim, Da Hye; Yang, Chae Ha; Kim, Hee Young; Gwak, Young S; Jang, Eun Young; Kim, Jae Su; Lee, Yun Kyu; Lee, Hyun Jong; Lee, Sang Nam; Lim, Sung Chul; Lee, Bong Hyo

    2018-01-01

    Methamphetamine is one of the widely abused drugs. In spite of a number of studies, there is still little successful therapy to suppress the methamphetamine abuse. Acupuncture has shown to attenuate the reinforcing effects of psychostimulant. Based on, the present study investigated if acupuncture could suppress intravenous methamphetamine self-administration behavior. In addition, a possible neuronal mechanism was investigated. Male Sprague-Dawley rats weighing 270-300g were trained to intake food pellet. After catheter implantation, animal was trained to self-administer methamphetamine (0.05mg/kg) intravenously using fixed ratio 1 schedule in daily 2h session during 3 weeks. After training, rats who established baseline (infusion variation less than 20% of the mean for 3 consecutive days) received acupuncture treatment on the next day. Acupuncture was performed at each acupoint manually. In the second experiment, the selective antagonists of GABA A or GABA B receptor were given before acupuncture to investigate the possible neuronal involvement of GABA receptor pathway in the acupuncture effects. C-Fos expression was examined in the nucleus accumbens to support behavioral data. Acupuncture at HT7, but not at control acupoint LI5, reduced the self-administration behavior significantly. Also, the effects of acupuncture were blocked by the GABA receptor antagonists. C-Fos expression was shown to be parallel with the behavioral data. Results of this study have shown that acupuncture at HT7 suppressed methamphetamine self-administration through GABA receptor system, suggesting that acupuncture at HT7 can be a useful therapy for the treatment of methamphetamine abuse. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. A putative octopamine/tyramine receptor mediating appetite in a hungry fly

    Science.gov (United States)

    Ishida, Yuko; Ozaki, Mamiko

    2011-07-01

    In the blowfly Phormia regina, experience of simultaneous feeding with d-limonene exposure inhibits proboscis extension reflex (PER) due to decreased tyramine (TA) titer in the brain. To elucidate the molecular mechanism of TA signaling pathway related to the associated feeding behavior, we cloned cDNA encoding the octopamine/TA receptor (PregOAR/TAR). The deduced protein is composed of 607 amino acid residues and has 7 predicted transmembrane domains. Based on homology and phylogenetic analyses, this protein belongs to the OAR/TAR family. The PregOAR/TAR was mainly expressed in head, with low levels of expression in other tissues at adult stages. Gene expression profile is in agreement with a plethora of functions ascribed to TA in various insect tissues. The immunolabeled cell bodies and processes were localized in the medial protocerebrum, outer layer of lobula, antennal lobe, and subesophageal ganglion. These results suggest that decrease of TA level in the brain likely affects neurons expressing PregOAR/TAR, causing mediation of the sensitivity in the sensillum and/or output of motor neurons for PER.

  3. 17-AAG sensitized malignant glioma cells to death-receptor mediated apoptosis.

    Science.gov (United States)

    Siegelin, Markus David; Habel, Antje; Gaiser, Timo

    2009-02-01

    17-AAG is a selective HSP90-inhibitor that exhibited therapeutic activity in cancer. In this study three glioblastoma cell lines (U87, LN229 and U251) were treated with 17-AAG, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) or the combination of both. Treatment with subtoxic doses of 17-AAG in combination with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces rapid apoptosis in TRAIL-resistant glioma cells, suggesting that this combined treatment may offer an attractive strategy for treating gliomas. 17-AAG treatment down-regulated survivin through proteasomal degradation. In addition, over-expression of survivin attenuated cytotoxicity induced by the combination of 17-AAG and TRAIL. In summary, survivin is a key regulator of TRAIL-17-AAG mediated cell death in malignant glioma.

  4. Serotonin₂A/C receptors mediate the aggressive phenotype of TLX gene knockout mice.

    Science.gov (United States)

    Juárez, Pablo; Valdovinos, Maria G; May, Michael E; Lloyd, Blair P; Couppis, Maria H; Kennedy, Craig H

    2013-11-01

    Deleting the tailless (TLX) gene in mice produces a highly aggressive phenotype yet to be characterized in terms of heterozygous animals or neurotransmitter mechanisms. We sought to establish pharmacological control over aggression and study the role of serotonin (5-HT)(2A/C) receptors in mediating changes in aggression. We analyzed aggression in mice heterozygous (+/-) or homozygous (-/-) for the TLX gene and wild-types (+/+) using a resident-intruder paradigm. No +/+ mice were aggressive, 36% of +/- TLX and 100% of -/- TLX mice showed aggression. Dose-effect functions were established for clozapine (0.1-1.5mg/kg, ip), ketanserin (0.3-1.25 mg/kg, ip), and (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane [(±)DOI] (0.5-2.0 mg/kg, ip). Injecting clozapine decreased the frequency and duration of attacks for +/- TLX and -/- TLX mice. Clozapine did not decrease grooming in either +/- TLX or -/- TLX mice but may have increased locomotion for -/- TLX mice. Injecting ketanserin, a 5-HT(2A/C) receptor antagonist, produced differential decreases in frequency and latency to aggression between genotypes and corresponding increases in locomotor behavior. Injecting (±)DOI, a 5-HT(2A/C) receptor agonist, increased the frequency and duration of attacks, decreased the latency to attacks, and decreased locomotion in +/- and -/- TLX mice. Results of the current study suggest aggression displayed by TLX null and heterozygous mice involves 5-HT(2A/C) receptors. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Gamma interferon augments Fc gamma receptor-mediated dengue virus infection of human monocytic cells.

    OpenAIRE

    Kontny, U; Kurane, I; Ennis, F A

    1988-01-01

    It has been reported that anti-dengue antibodies at subneutralizing concentrations augment dengue virus infection of monocytic cells. This is due to the increased uptake of dengue virus in the form of virus-antibody complexes by cells via Fc gamma receptors. We analyzed the effects of recombinant human gamma interferon (rIFN-gamma) on dengue virus infection of human monocytic cells. U937 cells, a human monocytic cell line, were infected with dengue virus in the form of virus-antibody complexe...

  6. Characterization of murine melanocortin receptors mediating adipocyte lipolysis and examination of signalling pathways involved

    DEFF Research Database (Denmark)

    Møller, Cathrine Laustrup; Raun, Kirsten; Jacobsen, Marianne Lambert

    2011-01-01

    hormone (a-MSH) generated from proopiomelanocortin (POMC), as well as synthetic MSH analogues to stimulate lipolysis in murine 3T3-L1 adipocytes it is shown that MC2R and MC5R are lipolytic mediators in differentiated 3T3-L1 adipocytes. Involvement of cAMP, phosphorylated extracellular signal...

  7. Ku proteins function as corepressors to regulate farnesoid X receptor-mediated gene expression

    International Nuclear Information System (INIS)

    Ohno, Masae; Kunimoto, Masaaki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2009-01-01

    The farnesoid X receptor (FXR; NR1H4) is a member of the nuclear receptor superfamily and regulates the expression of genes involved in enterohepatic circulation and the metabolism of bile acids. Based on functional analyses, nuclear receptors are divided into regions A-F. To explore the cofactors interacting with FXR, we performed a pull-down assay using GST-fused to the N-terminal A/B region and the C region, which are required for the ligand-independent transactivation and DNA-binding, respectively, of FXR, and nuclear extracts from HeLa cells. We identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and Ku70 as FXR associated factors. These proteins are known to have an important role in DNA repair, recombination, and transcription. DNA-PKcs mainly interacted with the A/B region of FXR, whereas the Ku proteins interacted with the C region and with the D region (hinge region). Chromatin immunoprecipitation assays revealed that the Ku proteins associated with FXR on the bile salt export pump (BSEP) promoter. Furthermore, we demonstrated that ectopic expression of the Ku proteins decreased the promoter activity and expression of BSEP gene mediated by FXR. These results suggest that the Ku proteins function as corepressors for FXR.

  8. Voltage dependent anion channel-1 regulates death receptor mediated apoptosis by enabling cleavage of caspase-8

    International Nuclear Information System (INIS)

    Chacko, Alex D; Liberante, Fabio; Paul, Ian; Longley, Daniel B; Fennell, Dean A

    2010-01-01

    Activation of the extrinsic apoptosis pathway by tumour necrosis factor related apoptosis inducing ligand (TRAIL) is a novel therapeutic strategy for treating cancer that is currently under clinical evaluation. Identification of molecular biomarkers of resistance is likely to play an important role in predicting clinical anti tumour activity. The involvement of the mitochondrial type 1 voltage dependent anion channel (VDAC1) in regulating apoptosis has been highly debated. To date, a functional role in regulating the extrinsic apoptosis pathway has not been formally excluded. We carried out stable and transient RNAi knockdowns of VDAC1 in non-small cell lung cancer cells, and stimulated the extrinsic apoptotic pathway principally by incubating cells with the death ligand TRAIL. We used in-vitro apoptotic and cell viability assays, as well as western blot for markers of apoptosis, to demonstrate that TRAIL-induced toxicity is VDAC1 dependant. Confocal microscopy and mitochondrial fractionation were used to determine the importance of mitochondria for caspase-8 activation. Here we show that either stable or transient knockdown of VDAC1 is sufficient to antagonize TRAIL mediated apoptosis in non-small cell lung cancer (NSCLC) cells. Specifically, VDAC1 is required for processing of procaspase-8 to its fully active p18 form at the mitochondria. Loss of VDAC1 does not alter mitochondrial sensitivity to exogenous caspase-8-cleaved BID induced mitochondrial depolarization, even though VDAC1 expression is essential for TRAIL dependent activation of the intrinsic apoptosis pathway. Furthermore, expression of exogenous VDAC1 restores the apoptotic response to TRAIL in cells in which endogenous VDAC1 has been selectively silenced. Expression of VDAC1 is required for full processing and activation of caspase-8 and supports a role for mitochondria in regulating apoptosis signaling via the death receptor pathway

  9. Natural Modulators of Endosomal Toll-Like Receptor-Mediated Psoriatic Skin Inflammation

    Directory of Open Access Journals (Sweden)

    Chao-Yang Lai

    2017-01-01

    Full Text Available Psoriasis is a chronic inflammatory autoimmune disease that can be initiated by excessive activation of endosomal toll-like receptors (TLRs, particularly TLR7, TLR8, and TLR9. Therefore, inhibitors of endosomal TLR activation are being investigated for their ability to treat this disease. The currently approved biological drugs adalimumab, etanercept, infliximab, ustekinumab, ixekizumab, and secukizumab are antibodies against effector cytokines that participate in the initiation and development of psoriasis. Several immune modulatory oligonucleotides and small molecular weight compounds, including IMO-3100, IMO-8400, and CPG-52364, that block the interaction between endosomal TLRs and their ligands are under clinical investigation for their effectiveness in the treatment of psoriasis. In addition, several chemical compounds, including AS-2444697, PF-05387252, PF-05388169, PF-06650833, ML120B, and PHA-408, can inhibit TLR signaling. Although these compounds have demonstrated anti-inflammatory activity in animal models, their therapeutic potential for the treatment of psoriasis has not yet been tested. Recent studies demonstrated that natural compounds derived from plants, fungi, and bacteria, including mustard seed, Antrodia cinnamomea extract, curcumin, resveratrol, thiostrepton, azithromycin, and andrographolide, inhibited psoriasis-like inflammation induced by the TLR7 agonist imiquimod in animal models. These natural modulators employ different mechanisms to inhibit endosomal TLR activation and are administered via different routes. Therefore, they represent candidate psoriasis drugs and might lead to the development of new treatment options.

  10. Dopamine D3 receptors mediate the discriminative stimulus effects of quinpirole in free-feeding rats.

    Science.gov (United States)

    Baladi, Michelle G; Newman, Amy H; France, Charles P

    2010-01-01

    The discriminative stimulus effects of dopamine (DA) D3/D2 receptor agonists are thought to be mediated by D2 receptors. To maintain responding, access to food is often restricted, which can alter neurochemical and behavioral effects of drugs acting on DA systems. This study established stimulus control with quinpirole in free-feeding rats and tested the ability of agonists to mimic and antagonists to attenuate the effects of quinpirole. The same antagonists were studied for their ability to attenuate quinpirole-induced yawning and hypothermia. DA receptor agonists apomorphine and lisuride, but not amphetamine and morphine, occasioned responding on the quinpirole lever. The discriminative stimulus effects of quinpirole were attenuated by the D3 receptor-selective antagonist N-{4-[4-(2,3-dichlorophenyl)-piperazin-1-yl]-trans-but-2-enyl}-4-pyridine-2-yl-benzamide HCl (PG01037) and the nonselective D3/D2 receptor antagonist raclopride, but not by the D2 receptor-selective antagonist 3-[4-(4-chlorophenyl)-4-hydroxypiperidin-1-yl]methyl-1H-indole (L-741,626); the potencies of PG01037 and raclopride to antagonize this effect of quinpirole paralleled their potencies to antagonize the ascending limb of the quinpirole yawning dose-response curve (thought to be mediated by D3 receptors). L-741,626 selectively antagonized the descending limb of the quinpirole yawning dose-response curve, and both L-741,626 and raclopride, but not PG01037, antagonized the hypothermic effects of quinpirole (thought to be mediated by D2 receptors). Food restriction (10 g/day/7 days) significantly decreased quinpirole-induced yawning without affecting the quinpirole discrimination. Many discrimination studies on DA receptor agonists use food-restricted rats; together with those studies, the current experiment using free-feeding rats suggests that feeding conditions affecting the behavioral effects of direct-acting DA receptor agonists might also have an impact on the effects of indirect-acting agonists such as cocaine and amphetamine.

  11. Receptor-mediated internalization of [3H]-neurotensin in synaptosomal preparations from rat neostriatum.

    Science.gov (United States)

    Nguyen, Ha Minh Ky; Cahill, Catherine M; McPherson, Peter S; Beaudet, Alain

    2002-06-01

    Following its binding to somatodendritic receptors, the neuropeptide neurotensin (NT) internalizes via a clathrin-mediated process. In the present study, we investigated whether NT also internalizes presynaptically using synaptosomes from rat neostriatum, a region in which NT1 receptors are virtually all presynaptic. Binding of [(3)H]-NT to striatal synaptosomes in the presence of levocabastine to block NT2 receptors is specific, saturable, and has NT1 binding properties. A significant fraction of the bound radioactivity is resistant to hypertonic acid wash indicating that it is internalized. Internalization of [(3)H]-NT, like that of [(125)I]-transferrin, is blocked by sucrose and low temperature, consistent with endocytosis occurring via a clathrin-dependent pathway. However, contrary to what was reported at the somatodendritic level, neither [(3)H]-NT nor [(125)I]-transferrin internalization in synaptosomes is sensitive to the endocytosis inhibitor phenylarsine oxide. Moreover, treatment of synaptosomes with monensin, which prevents internalized receptors from recycling to the plasma membrane, reduces [(3)H]-NT binding and internalization, suggesting that presynaptic NT1 receptors, in contrast to somatodendritic ones, are recycled back to the plasma membrane. Taken together, these results suggest that NT internalizes in nerve terminals via an endocytic pathway that is related to, but is mechanistically distinct from that responsible for NT internalization in nerve cell bodies.

  12. The α7-nicotinic acetylcholine receptor mediates the sensitivity of gastric cancer cells to taxanes.

    Science.gov (United States)

    Tu, Chao-Chiang; Huang, Chien-Yu; Cheng, Wan-Li; Hung, Chin-Sheng; Uyanga, Batzorig; Wei, Po-Li; Chang, Yu-Jia

    2016-04-01

    Gastric cancer is difficult to cure because most patients are diagnosed at an advanced disease stage. Systemic chemotherapy remains an important therapy for gastric cancer, but both progression-free survival and disease-free survival associated with various combination regimens are limited because of refractoriness and chemoresistance. Accumulating evidence has revealed that the homomeric α7-nicotinic acetylcholine receptor (A7-nAChR) promotes human gastric cancer by driving cancer cell proliferation, migration, and metastasis. Therefore, A7-nAChR may serve as a potential therapeutic target for gastric cancer. However, the role of A7-nAChR in taxane therapy for gastric cancer was unclear. Cells were subjected to A7-nAChR knockdown (A7-nAChR KD) using short interfering RNA (siRNA). The anti-proliferative effects of taxane were assessed via 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL), and cell cycle distribution assays. A7-nAChR-KD cells exhibited low resistance to docetaxel and paclitaxel treatment, as measured by the MTT assay. Following paclitaxel treatment, the proportion of apoptotic cells was higher among A7-nAChR-KD cells than among scrambled control cells, as measured by cell cycle distribution and TUNEL assays. Further molecular analyses showed a reduction in the pAKT levels and a dramatic increase in the Bad levels in paclitaxel-treated A7-nAChR-KD cells but not in scrambled control cells. Following paclitaxel treatment, the level of Bax was slightly increased in both cell populations, whereas Poly (ADP-ribose) polymerase (PARP) cleavage was increased only in A7-nAChR-KD cells. These findings indicate that A7-nAChR-KD cells are more sensitive to paclitaxel treatment. We conclude that A7-nAChR may be a key biomarker for assessing the chemosensitivity of gastric cancer cells to taxane.

  13. Toll-like receptor mediated activation is possibly involved in immunoregulating properties of cow's milk hydrolysates

    NARCIS (Netherlands)

    Kiewiet, M. B. Gea; Dekkers, Renske; Gros, Marjan; van Neerven, R. J. Joost; Groeneveld, Andre; de Vos, Paul; Faas, Marijke M.

    2017-01-01

    Immunomodulating proteins and peptides are formed during the hydrolysis of cow's milk proteins. These proteins are potential ingredients in functional foods used for the management of a range of immune related problems, both in infants and adults. However, the mechanism behind these effects is

  14. Glucocorticoid Receptor-Mediated Repression of Pro-Inflammatory Genes in Rheumatoid Arthritis

    Science.gov (United States)

    2015-10-01

    Braun TP, Zhu X, Szumowski M, Scott GD, Grossberg AJ, et al. 2011. Central nervous system inflam - mation induces muscle atrophy via activation of the...glucocorticoid receptor and nutritional sensor mTOR in skeletal muscle . Cell Metab. 13:170–82 133. Lutzner N, Kalbacher H, Krones-Herzig A, Rosl F. 2012...suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704

  15. Ursodeoxycholic Acid Induces Death Receptor-mediated Apoptosis in Prostate Cancer Cells

    Science.gov (United States)

    Lee, Won Sup; Jung, Ji Hyun; Panchanathan, Radha; Yun, Jeong Won; Kim, Dong Hoon; Kim, Hye Jung; Kim, Gon Sup; Ryu, Chung Ho; Shin, Sung Chul; Hong, Soon Chan; Choi, Yung Hyun; Jung, Jin-Myung

    2017-01-01

    Background Bile acids have anti-cancer properties in a certain types of cancers. We determined anticancer activity and its underlying molecular mechanism of ursodeoxycholic acid (UDCA) in human DU145 prostate cancer cells. Methods Cell viability was measured with an MTT assay. UDCA-induced apoptosis was determined with flow cytometric analysis. The expression levels of apoptosis-related signaling proteins were examined with Western blotting. Results UDCA treatment significantly inhibited cell growth of DU145 in a dose-dependent manner. It induced cellular shrinkage and cytoplasmic blebs and accumulated the cells with sub-G1 DNA contents. Moreover, UDCA activated caspase 8, suggesting that UDCA-induced apoptosis is associated with extrinsic pathway. Consistent to this finding, UDCA increased the expressions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, death receptor 4 (DR4) and death receptor 5 (DR5), and TRAIL augmented the UDCA-induced cell death in DU145 cells. In addition, UDCA also increased the expressions of Bax and cytochrome c and decreased the expression of Bcl-xL in DU145 cells. This finding suggests that UDCA-induced apoptosis may be involved in intrinsic pathway. Conclusions UDCA induces apoptosis via extrinsic pathway as well as intrinsic pathway in DU145 prostate cancer cells. UDCA may be a promising anti-cancer agent against prostate cancer. PMID:28382282

  16. Adrenergic β2-receptors mediates visceral hypersensitivity induced by heterotypic intermittent stress in rats.

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    Chunhua Zhang

    Full Text Available Chronic visceral pain in patients with irritable bowel syndrome (IBS has been difficult to treat effectively partially because its pathophysiology is not fully understood. Recent studies show that norepinephrine (NE plays an important role in the development of visceral hypersensitivity. In this study, we designed to investigate the role of adrenergic signaling in visceral hypersensitivity induced by heterotypical intermittent stress (HIS. Abdominal withdrawal reflex scores (AWRs used as visceral sensitivity were determined by measuring the visceromoter responses to colorectal distension. Colon-specific dorsal root ganglia neurons (DRGs were labeled by injection of DiI into the colon wall and were acutely dissociated for whole-cell patch-clamp recordings. Blood plasma level of NE was measured using radioimmunoassay kits. The expression of β2-adrenoceptors was measured by western blotting. We showed that HIS-induced visceral hypersensitivity was attenuated by systemic administration of a β-adrenoceptor antagonist propranolol, in a dose-dependent manner, but not by a α-adrenoceptor antagonist phentolamine. Using specific β-adrenoceptor antagonists, HIS-induced visceral hypersensitivity was alleviated by β2 adrenoceptor antagonist but not by β1- or β3-adrenoceptor antagonist. Administration of a selective β2-adrenoceptor antagonist also normalized hyperexcitability of colon-innervating DRG neurons of HIS rats. Furthermore, administration of β-adrenoceptor antagonist suppressed sustained potassium current density (IK without any alteration of fast-inactivating potassium current density (IA. Conversely, administration of NE enhanced the neuronal excitability and produced visceral hypersensitivity in healthy control rats, and blocked by β2-adrenoceptor antagonists. In addition, HIS significantly enhanced the NE concentration in the blood plasma but did not change the expression of β2-adrenoceptor in DRGs and the muscularis externa of the colon. The present study might provide a potential molecular target for therapy of visceral hypersensitivity in patents with IBS.

  17. Cerebellar Kainate Receptor-Mediated Facilitation of Glutamate Release Requires Ca2+-Calmodulin and PKA

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    Rafael Falcón-Moya

    2018-06-01

    Full Text Available We elucidated the mechanisms underlying the kainate receptor (KAR-mediated facilitatory modulation of synaptic transmission in the cerebellum. In cerebellar slices, KA (3 μM increased the amplitude of evoked excitatory postsynaptic currents (eEPSCs at synapses between axon terminals of parallel fibers (PF and Purkinje neurons. KA-mediated facilitation was antagonized by NBQX under condition where AMPA receptors were previously antagonized. Inhibition of protein kinase A (PKA suppressed the effect of KA on glutamate release, which was also obviated by the prior stimulation of adenylyl cyclase (AC. KAR-mediated facilitation of synaptic transmission was prevented by blocking Ca2+ permeant KARs using philanthotoxin. Furthermore, depletion of intracellular Ca2+ stores by thapsigargin, or inhibition of Ca2+-induced Ca2+-release by ryanodine, abrogated the synaptic facilitation by KA. Thus, the KA-mediated modulation was conditional on extracellular Ca2+ entry through Ca2+-permeable KARs, as well as and mobilization of Ca2+ from intracellular stores. Finally, KAR-mediated facilitation was sensitive to calmodulin inhibitors, W-7 and calmidazolium, indicating that the increased cytosolic [Ca2+] sustaining KAR-mediated facilitation of synaptic transmission operates through a downstream Ca2+/calmodulin coupling. We conclude that, at cerebellar parallel fiber-Purkinje cell synapses, presynaptic KARs mediate glutamate release facilitation, and thereby enhance synaptic transmission through Ca2+-calmodulin dependent activation of adenylyl cyclase/cAMP/protein kinase A signaling.

  18. Fc-receptor-mediated phagocytosis is regulated by mechanical properties of the target

    Science.gov (United States)

    Beningo, Karen A.; Wang, Yu-li

    2002-01-01

    Phagocytosis is an actin-based process used by macrophages to clear particles greater than 0.5 microm in diameter. In addition to its role in immunological responses, phagocytosis is also necessary for tissue remodeling and repair. To prevent catastrophic autoimmune reactions, phagocytosis must be tightly regulated. It is commonly assumed that the recognition/selection of phagocytic targets is based solely upon receptor-ligand binding. Here we report an important new criterion, that mechanical parameters of the target can dramatically affect the efficiency of phagocytosis. When presented with particles of identical chemical properties but different rigidity, macrophages showed a strong preference to engulf rigid objects. Furthermore, phagocytosis of soft particles can be stimulated with the microinjection of constitutively active Rac1 but not RhoA, and with lysophosphatidic acid, an agent known to activate the small GTP-binding proteins of the Rho family. These data suggest a Rac1-dependent mechanosensory mechanism for phagocytosis, which probably plays an important role in a number of physiological and pathological processes from embryonic development to autoimmune diseases.

  19. Identification of a novel modulator of thyroid hormone receptor-mediated action.

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    Bernhard G Baumgartner

    Full Text Available BACKGROUND: Diabetes is characterized by reduced thyroid function and altered myogenesis after muscle injury. Here we identify a novel component of thyroid hormone action that is repressed in diabetic rat muscle. METHODOLOGY/PRINCIPAL FINDINGS: We have identified a gene, named DOR, abundantly expressed in insulin-sensitive tissues such as skeletal muscle and heart, whose expression is highly repressed in muscle from obese diabetic rats. DOR expression is up-regulated during muscle differentiation and its loss-of-function has a negative impact on gene expression programmes linked to myogenesis or driven by thyroid hormones. In agreement with this, DOR enhances the transcriptional activity of the thyroid hormone receptor TR(alpha1. This function is driven by the N-terminal part of the protein. Moreover, DOR physically interacts with TR( alpha1 and to T(3-responsive promoters, as shown by ChIP assays. T(3 stimulation also promotes the mobilization of DOR from its localization in nuclear PML bodies, thereby indicating that its nuclear localization and cellular function may be related. CONCLUSIONS/SIGNIFICANCE: Our data indicate that DOR modulates thyroid hormone function and controls myogenesis. DOR expression is down-regulated in skeletal muscle in diabetes. This finding may be of relevance for the alterations in muscle function associated with this disease.

  20. Modulation of Toll-like receptor-mediated activation of Microglia

    NARCIS (Netherlands)

    Putten, C. M.-T. van der

    2015-01-01

    Microglia are the resident macrophages of the central nervous system (CNS). Like other tissue macrophages, microglia have many different functions under physiological as well as pathological conditions. Microglia can contribute to the initiation, progression and resolution of disease processes and

  1. Modeling receptor-mediated endocytosis of polymer-functionalized iron oxide nanoparticles by human macrophages

    Czech Academy of Sciences Publication Activity Database

    Lunov, O.; Zablotskyy, Vitaliy A.; Syrovets, T.; Röcker, C.; Tron, K.; Nienhaus, G.U.; Simmet, T.

    2011-01-01

    Roč. 32, č. 2 (2011), s. 547-555 ISSN 0142-9612 Institutional research plan: CEZ:AV0Z10100522 Keywords : macrophage * nanoparticle * bioabsorption * modeling * antisense * MRI (magnetic resonance imaging) Subject RIV: BM - Solid Matter Physics ; Magnetism Impact factor: 7.404, year: 2011 http://www.sciencedirect.com/science/article/pii/S014296121001149X

  2. Mechanisms of G Protein-Coupled Estrogen Receptor-Mediated Spinal Nociception

    DEFF Research Database (Denmark)

    Deliu, Elena; Brailoiu, G. Cristina; Arterburn, Jeffrey B.

    2012-01-01

    . Cytosolic calcium concentration elevates faster and with higher amplitude following G-1 intracellular microinjections compared to extracellular exposure, suggesting subcellular GPER functionality. Thus, GPER activation results in spinal nociception, and the downstream mechanisms involve cytosolic calcium......Human and animal studies suggest that estrogens are involved in the processing of nociceptive sensory information and analgesic responses in the central nervous system. Rapid pronociceptive estrogenic effects have been reported, some of which likely involve G protein-coupled estrogen receptor (GPER......) activation. Membrane depolarization and increases in cytosolic calcium and reactive oxygen species (ROS) levels are markers of neuronal activation, underlying pain sensitization in the spinal cord. Using behavioral, electrophysiological, and fluorescent imaging studies, we evaluated GPER involvement...

  3. An in vitro approach for prioritization and evaluation of chemical effects on glucocorticoid receptor mediated adipogenesis.

    Science.gov (United States)

    Hartman, Jessica K; Beames, Tyler; Parks, Bethany; Doheny, Daniel; Song, Gina; Efremenko, Alina; Yoon, Miyoung; Foley, Briana; Deisenroth, Chad; McMullen, Patrick D; Clewell, Rebecca A

    2018-05-18

    Rising obesity rates worldwide have socio-economic ramifications. While genetics, diet, and lack of exercise are major contributors to obesity, environmental factors may enhance susceptibility through disruption of hormone homeostasis and metabolic processes. The obesogen hypothesis contends that chemical exposure early in development may enhance adipocyte differentiation, thereby increasing the number of adipocytes and predisposing for obesity and metabolic disease. We previously developed a primary human adipose stem cell (hASC) assay to evaluate the effect of environmental chemicals on PPARG-dependent adipogenesis. Here, the assay was modified to determine the effects of chemicals on the glucocorticoid receptor (GR) pathway. In differentiation cocktail lacking the glucocorticoid agonist dexamethasone (DEX), hASCs do not differentiate into adipocytes. In the presence of GR agonists, adipocyte maturation was observed using phenotypic makers for lipid accumulation, adipokine secretion, and expression of key genes. To evaluate the role of environmental compounds on adipocyte differentiation, progenitor cells were treated with 19 prioritized compounds previously identified by ToxPi as having GR-dependent bioactivity, and multiplexed assays were used to confirm a GR-dependent mode of action. Five chemicals were found to be strong agonists. The assay was also modified to evaluate GR-antagonists, and 8/10 of the hypothesized antagonists inhibited adipogenesis. The in vitro bioactivity data was put into context with extrapolated human steady state concentrations (Css) and clinical exposure data (Cmax). These data support using a human adipose-derived stem cell differentiation assay to test the potential of chemicals to alter human GR-dependent adipogenesis. Copyright © 2017. Published by Elsevier Inc.

  4. Valerian inhibits rat hepatocarcinogenesis by activating GABA(A receptor-mediated signaling.

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    Anna Kakehashi

    Full Text Available Valerian is widely used as a traditional medicine to improve the quality of sleep due to interaction of several active components with the γ-aminobutyric acid (GABA A receptor (GABA(AR system. Recently, activation of GABA signaling in stem cells has been reported to suppress cell cycle progression in vivo. Furthermore, possible inhibitory effects of GABA(AR agonists on hepatocarcinogenesis have been reported. The present study was performed to investigate modulating effects of Valerian on hepatocarcinogenesis using a medium-term rat liver bioassay. Male F344 rats were treated with one of the most powerful Valerian species (Valeriana sitchensis at doses of 0, 50, 500 and 5000 ppm in their drinking water after initiation of hepatocarcinogenesis with diethylnitrosamine (DEN. Formation of glutathione S-transferase placental form positive (GST-P(+ foci was significantly inhibited by Valerian at all applied doses compared with DEN initiation control rats. Generation of 8-hydroxy-2'-deoxyguanosine in the rat liver was significantly suppressed by all doses of Valerian, likely due to suppression of Nrf2, CYP7A1 and induction of catalase expression. Cell proliferation was significantly inhibited, while apoptosis was induced in areas of GST-P(+ foci of Valerian groups associated with suppression of c-myc, Mafb, cyclin D1 and induction of p21(Waf1/Cip1, p53 and Bax mRNA expression. Interestingly, expression of the GABA(AR alpha 1 subunit was observed in GST-P(+ foci of DEN control rats, with significant elevation associated with Valerian treatment. These results indicate that Valerian exhibits inhibitory effects on rat hepatocarcinogenesis by inhibiting oxidative DNA damage, suppressing cell proliferation and inducing apoptosis in GST-P(+ foci by activating GABA(AR-mediated signaling.

  5. Chlorotoxin Fused to IgG-Fc Inhibits Glioblastoma Cell Motility via Receptor-Mediated Endocytosis

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    Tomonari Kasai

    2012-01-01

    Full Text Available Chlorotoxin is a 36-amino acid peptide derived from Leiurus quinquestriatus (scorpion venom, which has been shown to inhibit low-conductance chloride channels in colonic epithelial cells. Chlorotoxin also binds to matrix metalloproteinase-2 and other proteins on glioma cell surfaces. Glioma cells are considered to require the activation of matrix metalloproteinase-2 during invasion and migration. In this study, for targeting glioma, we designed two types of recombinant chlorotoxin fused to human IgG-Fcs with/without a hinge region. Chlorotoxin fused to IgG-Fcs was designed as a dimer of 60 kDa with a hinge region and a monomer of 30 kDa without a hinge region. The monomeric and dimeric forms of chlorotoxin inhibited cell proliferation at 300 nM and induced internalization in human glioma A172 cells. The monomer had a greater inhibitory effect than the dimer; therefore, monomeric chlorotoxin fused to IgG-Fc was multivalently displayed on the surface of bionanocapsules to develop a drug delivery system that targeted matrix metalloproteinase-2. The target-dependent internalization of bionanocapsules in A172 cells was observed when chlorotoxin was displayed on the bionanocapsules. This study indicates that chlorotoxin fused to IgG-Fcs could be useful for the active targeting of glioblastoma cells.

  6. Dopamine D(1) receptor-mediated control of striatal acetylcholine release by endogenous dopamine.

    Science.gov (United States)

    Acquas, E; Di Chiara, G

    1999-10-27

    The role of dopamine D(1) and D(2) receptors in the control of acetylcholine release in the dorsal striatum by endogenous dopamine was investigated by monitoring with microdialysis the effect of the separate or combined administration of the dopamine D(1) receptor antagonist, SCH 39166 ¿(-)-trans-6,7,7a,8,9, 13b-exahydro-3-chloro-2-hydroxy-N-methyl-5H-benzo-[d]-nap hto-[2, 1b]-a