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Sample records for irradiated human lymphoblastoid

  1. Cytogenetic characterization of low-dose hyper-radiosensitivity in Cobalt-60 irradiated human lymphoblastoid cells.

    PubMed

    Joshi, Gnanada S; Joiner, Michael C; Tucker, James D

    2014-12-01

    The dose-effect relationships of cells exposed to ionizing radiation are frequently described by linear quadratic (LQ) models over an extended dose range. However, many mammalian cell lines, when acutely irradiated in G2 at doses ≤0.3Gy, show hyper-radiosensitivity (HRS) as measured by reduced clonogenic cell survival, thereby indicating greater cell lethality than is predicted by extrapolation from high-dose responses. We therefore hypothesized that the cytogenetic response in G2 cells to low doses would also be steeper than predicted by LQ extrapolation from high doses. We tested our hypothesis by exposing four normal human lymphoblastoid cell lines to 0-400cGy of Cobalt-60 gamma radiation. The cytokinesis block micronucleus assay was used to determine the frequencies of micronuclei and nucleoplasmic bridges. To characterize the dependence of the cytogenetic damage on dose, univariate and multivariate regression analyses were used to compare the responses in the low- (HRS) and high-dose response regions. Our data indicate that the slope of the response for all four cell lines at ≤20cGy during G2 is greater than predicted by an LQ extrapolation from the high-dose responses for both micronuclei and bridges. These results suggest that the biological consequences of low-dose exposures could be underestimated and may not provide accurate risk assessments following such exposures.

  2. Identification of low-dose responsive metabolites in X-irradiated human B lymphoblastoid cells and fibroblasts

    PubMed Central

    Tsuyama, Naohiro; Mizuno, Hajime; Katafuchi, Atsushi; Abe, Yu; Kurosu, Yumiko; Yoshida, Mitsuaki; Kamiya, Kenji; Sakai, Akira

    2015-01-01

    Ionizing radiation (IR) induces cellular stress responses, such as signal transduction, gene expression, protein modification, and metabolite change that affect cellular behavior. We analyzed X-irradiated human Epstein-Barr virus-transformed B lymphoblastoid cells and normal fibroblasts to search for metabolites that would be suitable IR-responsive markers by Liquid Chromotography–Mass spectrometry (LC–MS). Mass spectra, as analyzed with principal component analysis, showed that the proportion of peaks with IR-induced change was relatively small compared with the influence of culture time. Dozens of peaks that had either been upregulated or downregulated by IR were extracted as candidate IR markers. The IR-changed peaks were identified by comparing mock-treated groups to 100 mGy-irradiated groups that had recovered after 10 h, and the results indicated that the metabolites involved in nucleoside synthesis increased and that some acylcarnitine levels decreased in B lymphoblastoids. Some peaks changed by as much as 20 mGy, indicating the presence of an IR-sensitive signal transduction/metabolism control mechanism in these cells. On the other hand, we could not find common IR-changed peaks in fibroblasts of different origin. These data suggest that cell phenotype-specific pathways exist, even in low-dose responses, and could determine cell behavior. PMID:25227127

  3. Synergistic Effects of Incubation in Rotating Bioreactors and Cumulative Low Dose 60Co γ-ray Irradiation on Human Immortal Lymphoblastoid Cells

    NASA Astrophysics Data System (ADS)

    Wei, Lijun; Han, Fang; Yue, Lei; Zheng, Hongxia; Yu, Dan; Ma, Xiaohuan; Cheng, Huifang; Li, Yu

    2012-11-01

    The complex space environments can influence cell structure and function. The research results on space biology have shown that the major mutagenic factors in space are microgravity and ionizing radiation. In addition, possible synergistic effects of radiation and microgravity on human cells are not well understood. In this study, human immortal lymphoblastoid cells were established from human peripheral blood lymphocytes and the cells were treated with low dose (0.1, 0.15 and 0.2 Gy) cumulative 60Co γ-irradiation and simulated weightlessness [obtained by culturing cells in the Rotating Cell Culture System (RCCS)]. The commonly used indexes of cell damage such as micronucleus rate, cell cycle and mitotic index were studied. Previous work has proved that Gadd45 (growth arrest and DNA-damage-inducible protein 45) gene increases with a dose-effect relationship, and will possibly be a new biological dosimeter to show irradiation damage. So Gadd45 expression is also detected in this study. The micronucleus rate and the expression of Gadd45α gene increased with irradiation dose and were much higher after incubation in the rotating bioreactor than that in the static irradiation group, while the cell proliferation after incubation in the rotating bioreactor decreased at the same time. These results indicate synergetic effects of simulated weightlessness and low dose irradiation in human cells. The cell damage inflicted by γ-irradiation increased under simulated weightlessness. Our results suggest that during medium- and long-term flight, the human body can be damaged by cumulative low dose radiation, and the damage will even be increased by microgravity in space.

  4. Identification of differentially transcribed genes in human lymphoblastoid cells irradiated with 0.5 Gy of gamma-ray and the involvement of low dose radiation inducible CHD6 gene in cell proliferation and radiosensitivity.

    PubMed

    Wang, H P; Long, X H; Sun, Z Z; Rigaud, O; Xu, Q Z; Huang, Y C; Sui, J L; Bai, B; Zhou, P K

    2006-03-01

    To identify candidate genes specifically involved in response to low-dose irradiation in human lymphoblastoid cells; to better clarify the role of the human chromodomain helicase DNA binding protein 6 gene (CHD6), one of these genes, in cell proliferation and radiosensitivity. DNA microarray technology was used to analyse global transcriptional profile in human lymphoblastoid AHH-1 cells at 4 h after exposure to 0.5 Gy of gamma-ray. Gene expression changes were confirmed by semi-quantitative reverse transcription--polymerase chain reaction (RT-PCR) and Northern blot. RNA interfering technology was employed to knock-down the CHD6 gene in A549 cells. Colony-forming ability was used to analyse radiosensitivity. The microarray assay revealed a set of 0.5 Gy-responsive genes, including 30 up-regulated genes and 45 down-regulated genes. The up-regulated genes include a number of genes involved in: signal transduction pathways, e.g., STAT3, CAMKK2, SIRT1, CREM, MAPK3K7IP2 and GPR56; transcription or DNA-binding, e.g., CHD6, CRSP3, SNURF, SH2 domain binding protein 1 and MIZF. Some of the down-regulated genes are involved in: cytoskeleton and cell movement (WASF2, LCP1, MSN, NIPSNAP1, KIF2C); DNA replication and repair (MCM2, MCM3, MCM7 and XRCC-4). Radiation-increased expression of CHD6 was also found in A549 cells and HeLa cells. The sustained CHD6 induction was restricted to relatively low doses (0.2 Gy or 0.5 Gy), no change occurring after 4 Gy irradiation. Silencing of CHD6 mediated by siRNA increased the growth rate of A549 cells by 40 approximately 60%. Most importantly, silencing CHD6 led to an increased radioresistance of A459 cells to radiation doses up to 2 Gy, but barely affected the sensitivity of cells at 4 and 8 Gy. This study has identified a set of genes responsive to 0.5 Gy of gamma-rays. CDH6 gene can be specifically up-regulated by low dose irradiation, and its inducible expression could be involved in a low dose hypersensitive response.

  5. The role of mitochondria in the radiation-induced bystander effect in human lymphoblastoid cells.

    PubMed

    Rajendran, Sountharia; Harrison, Scott H; Thomas, Robert A; Tucker, James D

    2011-02-01

    Cells without intact mitochondrial DNA have been shown to lack the bystander effect, which is an energy-dependent process. We hypothesized that cells harboring mutations in mitochondrial genes responsible for ATP synthesis would show a decreased bystander effect compared to normal cells. Radiation-induced bystander effects were analyzed in two normal and four mitochondrial mutant human lymphoblastoid cells. Medium from previously irradiated cells (conditioned medium) was transferred to unirradiated cells from the respective cell lines and evaluated for the bystander effect using the cytokinesis-block micronucleus assay. Unlike normal cells that were used as a control, mitochondrial mutant cells neither generated nor responded to the bystander signals. The bystander effect was inhibited in normal cells by adding the mitochondrial inhibitors rotenone and oligomycin to the culture medium. Time-controlled blocking of the bystander effect by inhibitors was found to occur either for prolonged exposure to the inhibitor prior to irradiation with an immediate and subsequent removal of the inhibitors or immediate post-application of the inhibitor. Adding the inhibitors just prior to irradiation and removing them immediately after irradiation was uneventful. Fully functional mitochondrial metabolic capability may therefore be essential for the bystander effect.

  6. Heterogeneity of a human T-lymphoblastoid cell line

    SciTech Connect

    Snow, K.; Judd, W.

    1987-08-01

    A human T-lymphoblastoid cell line (Jurkat) was cloned, and four resulting sublines were characterized in a variety of ways with the objective of gaining information on heterogeneity in cell lines. Within a few weeks of cloning, distinct cellular morphologies and growth patterns became apparent in the four sublines. Growth rate measurements made over 3 months did not show any significant differences between the sublines. Surface protein profiles obtained by radioimmunoprecipitation using antisera in conjunction with extracts from (/sup 35/S)Met and /sup 125/I-labeled cells revealed differences between the sublines. Analysis of total cell DNA showed that one of the sublines possessed only half the chromosome complement of the other sublines and the parental line. Karyotyping confirmed this result and, in addition, demonstrated that chromosome numbers fluctuated around a mean value for each subline. Karyotypic variability became apparent within 2 months of cloning and tended to increase with time in culture. G-banding analysis showed that the analyzed cell populations contained distinctive cytogenetic aberrations. Properties of the cloned sublines were monitored over a 9-month period. One of the sublines that had shown heterogeneous morphology even after 6 weeks maintained the heterogeneity throughout this time. Another subline underwent a marked change in morphology (round to irregular) and growth habit (single cells to large clumps) with increasing time in culture. Interestingly, several alterations to surface proteins accompanied these growth changes. A third subline had relatively stable morphology and chromosome number throughout the 9-month period. The modal chromosome number was hypotetraploid for three sublines and the parent line, but was diploid for another subline.

  7. Network signatures of cellular immortalization in human lymphoblastoid cell lines

    SciTech Connect

    Shim, Sung-Mi; Jung, So-Young; Nam, Hye-Young; Kim, Hye-Ryun; Lee, Mee-Hee; Kim, Jun-Woo; Han, Bok-Ghee; Jeon, Jae-Pil

    2013-11-15

    Highlights: •We identified network signatures of LCL immortalization from transcriptomic profiles. •More than 41% of DEGs are possibly regulated by miRNAs in LCLs. •MicroRNA target genes in LCLs are involved in apoptosis and immune-related functions. •This approach is useful to find functional miRNA targets in specific cell conditions. -- Abstract: Human lymphoblastoid cell line (LCL) has been used as an in vitro cell model in genetic and pharmacogenomic studies, as well as a good model for studying gene expression regulatory machinery using integrated genomic analyses. In this study, we aimed to identify biological networks of LCL immortalization from transcriptomic profiles of microRNAs and their target genes in LCLs. We first selected differentially expressed genes (DEGs) and microRNAs (DEmiRs) between early passage LCLs (eLCLs) and terminally differentiated late passage LCLs (tLCLs). The in silico and correlation analysis of these DEGs and DEmiRs revealed that 1098 DEG–DEmiR pairs were found to be positively (n = 591 pairs) or negatively (n = 507 pairs) correlated with each other. More than 41% of DEGs are possibly regulated by miRNAs in LCL immortalizations. The target DEGs of DEmiRs were enriched for cellular functions associated with apoptosis, immune response, cell death, JAK–STAT cascade and lymphocyte activation while non-miRNA target DEGs were over-represented for basic cell metabolisms. The target DEGs correlated negatively with miR-548a-3p and miR-219-5p were significantly associated with protein kinase cascade, and the lymphocyte proliferation and apoptosis, respectively. In addition, the miR-106a and miR-424 clusters located in the X chromosome were enriched in DEmiR–mRNA pairs for LCL immortalization. In this study, the integrated transcriptomic analysis of LCLs could identify functional networks of biologically active microRNAs and their target genes involved in LCL immortalization.

  8. Perspectives on fast-neutron mutagenesis of human lymphoblastoid cells.

    PubMed

    Kronenberg, A

    1991-10-01

    The effects of low-fluence exposures to (Pu, Be) neutrons (En = 4.2 MeV) have been studied in a sensitive human B-lymphoblastoid cell line, TK6. Mutations were scored for two genetic loci, hypoxanthine phosphoribosyltransferase (hgprt) and thymidine kinase (tk), as a function of dose and dose rate. For exposures limited to less than one cell cycle, the mutation frequency for the hgprt locus was 1.92 X 10(-7)/cGy. When exposures were protracted over multiple cell generations, mutation yields were increased to 6.07 X 10(-7)/cGy. Similar yields were obtained for the induction of tk-deficient mutants with a normal cell generation time (tk-ng) when exposures were carried out at very low dose rates over multiple cell generations. In the series of data presented here, the results obtained for short-duration neutron exposures are compared with data obtained for monoenergetic heavy charged particles of defined linear energy transfer (LET) produced at the BEVALAC accelerator at Lawrence Berkeley Laboratory. TK6 cells have been exposed to beams ranging in atomic number from 20Ne to 40Ar over an energy range from 330 to 670 MeV/amu. Mutation induction was evaluated for both loci for a subset of these beams. The results obtained with 20Ne ions of 425 MeV/amu (LET = 32 keV/microns) and 28Si ions of 670 MeV/amu (LET = 50 keV/microns) closely resemble the mutation yields obtained for brief exposures to (Pu, Be) neutrons. The nature of alterations in DNA structure induced within the tk locus of tk-ng mutants is reviewed for a series of neutron-induced mutants and a series of mutants induced by exposure to 40Ar ions (470 MeV/amu, LET = 95 keV/microns). The mutational spectra for these two types of mutants were similar and were dominated by allele loss mutations. Multilocus deletions inclusive of the c-erbA1 locus were common among tk-deficient mutants induced by these densely ionizing radiations. For the mutants induced by 40Ar ions, it is likely that the mutations were produced by

  9. Radiation quality and mutagenesis in human lymphoblastoid cells.

    PubMed

    Liber, Howard L; Idate, Rupa; Warner, Christy; Bailey, Susan M

    2014-10-01

    An interesting problem associated with studying the effects of low doses of high atomic number and energy (HZE) particles, as found in space, is that not all cells will necessarily be similarly traversed during exposure, a scenario that greatly complicates the measurement of end points that require time to develop, gene-locus mutation being a perfect example. The standard protocol for measuring mutations at the heterozygous thymidine kinase locus in human lymphoblastoid cells involves waiting three days after treatment for newly induced mutants to fully express, at which time cells are then plated in the presence of the selective agent, and mutants are counted three weeks later. This approach is acceptable as long as all cells are uniformly affected, as is the case with low-linear energy transfer (LET) ionizing radiation. However, for HZE particles some fraction of cells may not be traversed or perhaps would receive fewer than the average number of "hits", and they would continue to grow at or closer to the normal rate, thus outpacing cells that received more damage. As a result, at three days post-treatment, more heavily damaged cells will have been "diluted" by the less damaged ones, and thus the measured mutant frequency (MF) will underestimate actual mutant frequency. We therefore developed a modified approach for measuring mutation that eliminates this problem and demonstrates that the mutagenicity of 1 GeV/n Fe ions are underestimated by a factor of two when using the standard MF protocol. Furthermore, we determined the mutagenic effects of a variety of heavy ions, all of which induced mutations in a linear fashion. We found that the maximal yield of mutations (i.e., highest relative biological efficiency) was about 7.5 times higher at an LET of 70 keV/μ (400 MeV/n Si) than for gamma rays. Nontargeted mutagenicity after treatment with ionizing radiation was also investigated. For each particular ion/energy examined and in agreement with many previous studies

  10. Efficacy of human lymphoblastoid interferon in the therapy of resistant condyloma acuminata.

    PubMed

    Gall, S A; Hughes, C E; Mounts, P; Segriti, A; Weck, P K; Whisnant, J K

    1986-05-01

    The efficacy and tolerance of human lymphoblastoid interferon (Wellferon) were studied in an open label trial of 17 patients with resistant and persistent condyloma acuminata. Patients were treated intramuscularly with 5 X 10(6) U (5 MU)/m2 daily for 28 days followed by thrice weekly injections for two weeks. Sixteen patients were considered evaluable; eight experienced complete clearance, seven had significant reduction (greater than 50%) in lesion size, and one showed no response during the course of this trial. Biologic side effects of interferon occurred in all patients during initial dosing and diminished during thrice weekly therapy. Intramuscular injections and associated side effects were tolerated well. This study shows that systemic human lymphoblastoid interferon is active in treating severe recurrent genital warts in women with a history of recalcitrant disease.

  11. Radiation Induced Bystander Effects in Human Lymphoblastoid Cells

    DTIC Science & Technology

    2003-12-01

    34 observ6 peut etre caus6 par les interactions cellulaires via les prot~ines s~cr~toires 1ib~r~es par les cellules irradi~es en agissant sur les...l’accident du r~acteur de Chernobyl. Nous avons formulk l’hypoth~se que l’effet "bystander" observ6 pouvait 6tre une consdquence d’interactions cellulaires ...qui seraient indicatifs d’expositions biologiques ou chimiques. 11 est pr~vu que certains de ces marqueurs seront communs aux trois agents stressants

  12. Human iPSC-derived neurons and lymphoblastoid cells for personalized medicine research in neuropsychiatric disorders

    PubMed Central

    Gurwitz, David

    2016-01-01

    The development and clinical implementation of personalized medicine crucially depends on the availability of high-quality human biosamples; animal models, although capable of modeling complex human diseases, cannot reflect the large variation in the human genome, epigenome, transcriptome, proteome, and metabolome. Although the biosamples available from public biobanks that store human tissues and cells may represent the large human diversity for most diseases, these samples are not always sufficient for developing biomarkers for patient-tailored therapies for neuropsychiatric disorders. Postmortem human tissues are available from many biobanks; nevertheless, collections of neuronal human cells from large patient cohorts representing the human diversity remain scarce. Two tools are gaining popularity for personalized medicine research on neuropsychiatric disorders: human induced pluripotent stem cell-derived neurons and human lymphoblastoid cell lines. This review examines and contrasts the advantages and limitations of each tool for personalized medicine research. PMID:27757061

  13. Human iPSC-derived neurons and lymphoblastoid cells for personalized medicine research in neuropsychiatric disorders.

    PubMed

    Gurwitz, David

    2016-09-01

    The development and clinical implementation of personalized medicine crucially depends on the availability of high-quality human biosamples; animal models, although capable of modeling complex human diseases, cannot reflect the large variation in the human genome, epigenome, transcriptome, proteome, and metabolome. Although the biosamples available from public biobanks that store human tissues and cells may represent the large human diversity for most diseases, these samples are not always sufficient for developing biomarkers for patient-tailored therapies for neuropsychiatric disorders. Postmortem human tissues are available from many biobanks; nevertheless, collections of neuronal human cells from large patient cohorts representing the human diversity remain scarce. Two tools are gaining popularity for personalized medicine research on neuropsychiatric disorders: human induced pluripotent stem cell-derived neurons and human lymphoblastoid cell lines. This review examines and contrasts the advantages and limitations of each tool for personalized medicine research.

  14. Use of RAPD to detect sodium arsenite-induced DNA damage in human lymphoblastoid cells.

    PubMed

    Lee, Yuan-Cho; Yang, Vivian C; Wang, Tsu-Shing

    2007-09-24

    Inorganic arsenic is a known human carcinogen, yet its mechanism of action remains unclear. Our previous study showed that arsenite significantly induces oxidative DNA adducts and DNA-protein cross-links in several mammalian cell lines. In the present study, we used the random amplified polymorphic DNA (RAPD) assay to evaluate the possible target in the genomic DNA of human lymphoblastoid cells that were exposed to sodium arsenite. Treatment with both 10 and 80 microM arsenite for 4h induced significant changes in RAPD profiles compared with the control pattern. Two 10-mer RAPD primers (D11 and F1) produced the most distinguishable banding profiles between arsenite-treated and control genomic DNA. The sequencing of four arsenite-sensitive RAPD bands showed that the RB1CC1 and PACE4 genes might be the DNA targets of sodium arsenite treatment. We propose that arsenite may induce sequence- or gene-specific damage and then change the RAPD profile in human lymphoblastoid cells. The results of our study also show that RAPD combined with other techniques is a good tool for detecting alterations in genomic DNA and for the direct screening of new molecular markers related to arsenite-induced carcinogenesis.

  15. Analysis of cellular response by exposure to acute or chronic radiation in human lymphoblastoid TK-6 cells

    NASA Astrophysics Data System (ADS)

    Ohnishi, T.; Yasumoto, J.; Takahashi, A.; Ohnishi, K.

    To clarify the biological effects of low-dose rate radiation on human health for long-term stay in space, we analyzed the induction of apoptosis and apoptosis-related gene expression after irradiation with different dose-rate in human lymphoblastoid TK-6 cells harboring wild-type p53 gene. We irradiated TK-6 cells by X-ray at 1.5 Gy (1 Gy/min) and then sampled at 25 hr after culturing. We also irradiated by gamma-ray at 1.5 Gy (1 mGy/min) and then sampled immediately or 25 hr after irradiation. For DNA ladder analysis, we extracted DNA from these samples and electrophoresed with 2% agarose gel. In addition, we extracted mRNA from these samples for DNA-array analysis. mRNA from non-irradiated cells was used as a control. After labeling the cDNA against mRNA with [α -33P]-dCTP and hybridizing onto DNA array (Human Apoptosis Expression Array, R&D Systems), we scanned the profiles of the spots by a phosphorimager (BAS5000, FUJI FILM) and calculated using a NIH Image program. The data of each DNA-array were normalized with eight kinds of house keeping genes. We analyzed the expression level of apoptosis-related genes such as p53-related, Bcl-2 family, Caspase family and Fas-related genes. DNA ladders were obviously detected in the cells exposed to a high dose-rate radiation. We detected the induction of the gene expression of apoptosis-promotive genes. In contrast, almost no apoptosis was observed in the cells exposed to the chronic radiation at a low dose-rate. In addition, we detected the induction of the gene expression of apoptosis-suppressive genes as compared with apoptosis promotive-genes immediately after chronic irradiation. These results lead the importance of biological meaning of exposure to radiation at low dose-rate from an aspect of carcinogenesis. Finally, the effects of chronic irradiation become a highly important issue in space radiation biology for human health.

  16. Leukemia patient-derived lymphoblastoid cell lines exhibit increased induction of leukemia-associated transcripts following high-dose irradiation.

    PubMed

    Spencer, A; Granter, N

    1999-09-01

    Improvement in diagnostic cytogenetic techniques has led to the recognition of an increasing number of leukemia-associated chromosomal translocations and inversions. These genetic lesions frequently are associated with the disruption of putative transcription factors and the production of hybrid transcripts that are implicated in leukemogenesis. Epidemiologic evidence suggests that some, but not all, individuals with a history of gamma-irradiation exposure are at increased risk of developing chronic myeloid leukemia (CML). CML is characterized by the Philadelphia chromosome and transcription of the resulting hybrid BCR-ABL gene. Utilizing the leukemia-associated BCR-ABL p210 transcript as a marker, we sought differences in the induction of illegitimate genetic recombination following high-dose gamma-irradiation of karyotypically normal lymphoblastoid cell lines (LCL) derived from individuals with and without a history of myeloid leukemias. Six LCL [4 leukemia patient derived [2 acute myeloid leukemia and 2 CML] and 2 from normal individuals were analyzed with reverse transcriptase polymerase chain reaction for BCR-ABL under stringent conditions following exposure to 0, 50, or 100 Gy of LET gamma-irradiation delivered via a Varian linear accelerator at 4 MV. Transcripts identical to disease-associated b2a2 and b3a2 transcripts were detected both spontaneously (background illegitimate genetic recombination) and following gamma-irradiation. Background BCR-ABL positivity was demonstrable in 4 of the 6 LCL, with no significant difference in detection between leukemic- and nonleukemic-derived LCL. Overall, increasing gamma-irradiation dose resulted in an increased frequency of BCR-ABL transcript detection (0 Gy vs 50 Gy vs 100 Gy,p = 0.0023, Chi-square test). Within the leukemic- but not the nonleukemic-derived LCL there was significantly greater BCR-ABL positivity after gamma-irradiation compared to unirradiated equivalents. Furthermore, the BCR-ABL positivity of both

  17. Cytotoxic effect of anti-idiotype antibody-chlorambucil conjugates against human lymphoblastoid cells.

    PubMed Central

    Tung, E; Goust, J M; Chen, W Y; Kang, S S; Wang, I Y; Wang, A C

    1983-01-01

    The secreted IgMs of two human lymphoblastoid cell lines, RPMI-6410 and RPMI-8392, were purified. Antisera against these two IgMs were raised in rabbits and made idiotypically specific to the respective antigens through various absorption procedures. By immunofluorescence and radioimmunoassay techniques, the purified anti-idiotype antibodies were found to react also with the membrane Igs of the respective cell lines, but not with those of other cell lines. The purified anti-idiotype antibodies were then coupled with Chlorambucil to form antibody-drug conjugates, whose effectiveness in the in-vitro killing of target cells was evaluated by a chromium-release cytotoxicity assay. The results showed that these anti-idiotype antibody-Chlorambucil conjugates were specifically cytotoxic to lymphoblastoid cells that bore membrane Igs carrying the respective idiotypic determinant(s). Furthermore, the conjugates were far more effective in causing cytolysis to the target cells than either Chlorambucil or the anti-idiotype antibodies alone. PMID:6350169

  18. Cytotoxic effect of anti-idiotype antibody-chlorambucil conjugates against human lymphoblastoid cells.

    PubMed

    Tung, E; Goust, J M; Chen, W Y; Kang, S S; Wang, I Y; Wang, A C

    1983-09-01

    The secreted IgMs of two human lymphoblastoid cell lines, RPMI-6410 and RPMI-8392, were purified. Antisera against these two IgMs were raised in rabbits and made idiotypically specific to the respective antigens through various absorption procedures. By immunofluorescence and radioimmunoassay techniques, the purified anti-idiotype antibodies were found to react also with the membrane Igs of the respective cell lines, but not with those of other cell lines. The purified anti-idiotype antibodies were then coupled with Chlorambucil to form antibody-drug conjugates, whose effectiveness in the in-vitro killing of target cells was evaluated by a chromium-release cytotoxicity assay. The results showed that these anti-idiotype antibody-Chlorambucil conjugates were specifically cytotoxic to lymphoblastoid cells that bore membrane Igs carrying the respective idiotypic determinant(s). Furthermore, the conjugates were far more effective in causing cytolysis to the target cells than either Chlorambucil or the anti-idiotype antibodies alone.

  19. Autophagy is the predominant process induced by arsenite in human lymphoblastoid cell lines

    SciTech Connect

    Bolt, Alicia M.; Byrd, Randi M.; Klimecki, Walter T.

    2010-05-01

    Arsenic is a widespread environmental toxicant with a diverse array of molecular targets and associated diseases, making the identification of the critical mechanisms and pathways of arsenic-induced cytotoxicity a challenge. In a variety of experimental models, over a range of arsenic exposure levels, apoptosis is a commonly identified arsenic-induced cytotoxic pathway. Human lymphoblastoid cell lines (LCL) have been used as a model system in arsenic toxicology for many years, but the exact mechanism of arsenic-induced cytotoxicity in LCL is still unknown. We investigated the cytotoxicity of sodium arsenite in LCL 18564 using a set of complementary markers for cell death pathways. Markers indicative of apoptosis (phosphatidylserine externalization, PARP cleavage, and sensitivity to caspase inhibition) were uniformly negative in arsenite exposed cells. Interestingly, electron microscopy, acidic vesicle fluorescence, and expression of LC3 in LCL 18564 identified autophagy as an arsenite-induced process that was associated with cytotoxicity. Autophagy, a cellular programmed response that is associated with both cellular stress adaptation as well as cell death appears to be the predominant process in LCL cytotoxicity induced by arsenite. It is unclear, however, whether LCL autophagy is an effector mechanism of arsenite cytotoxicity or alternatively a cellular compensatory mechanism. The ability of arsenite to induce autophagy in lymphoblastoid cell lines introduces a potentially novel mechanistic explanation of the well-characterized in vitro and in vivo toxicity of arsenic to lymphoid cells.

  20. Ciprofloxacin-induced inhibition of topoisomerase II in human lymphoblastoid cells.

    PubMed Central

    Bredberg, A; Brant, M; Jaszyk, M

    1991-01-01

    The antibacterial activities of the fluorinated 4-quinolones (e.g., ciprofloxacin) have been ascribed to a marked inhibition of bacterial DNA gyrase. In contrast, the influence on purified mammalian DNA enzymes, including topoisomerases, has been reported to be several orders of magnitude weaker, occurring at concentrations higher than 100 micrograms of ciprofloxacin per ml. In this study, using a nondenaturing filter elution method, a marked induction of double-strand DNA breaks in human lymphoblastoid cells exposed to 80 micrograms of ciprofloxacin per ml was seen. The proportion of single-strand versus double-strand DNA breaks was similar to that seen with the topoisomerase II inhibitory antitumor agent VP-16. The cellular recovery was more rapid after treatment with ciprofloxacin than after treatment with VP-16, displaying a normal elution profile within 15 min at 37 degrees C (60 min for VP-16). These data indicate that ciprofloxacin has an effect on intracellularly located topoisomerase II in humans. PMID:1645508

  1. Induction of apoptosis by epigallocatechin-3-gallate in human lymphoblastoid B cells

    SciTech Connect

    Noda, Chiseko He, Jinsong; Takano, Tomoko; Tanaka, Chisato; Kondo, Toshinori; Tohyama, Kaoru; Yamamura, Hirohei; Tohyama, Yumi

    2007-11-03

    (-)-Epigallocatechin-3-gallate (EGCG), a major constituent of green tea polyphenols, has been shown to suppress cancer cell proliferation and induce apoptosis. In this study we investigated its efficacy and the mechanism underlying its effect using human B lymphoblastoid cell line Ramos, and effect of co-treatment with EGCG and a chemotherapeutic agent on apoptotic cell death. EGCG induced dose- and time-dependent apoptotic cell death accompanied by loss of mitochondrial transmembrane potential, release of cytochrome c into the cytosol, and cleavage of pro-caspase-9 to its active form. EGCG also enhanced production of intracellular reactive oxygen species (ROS). Pretreatment with diphenylene iodonium chloride, an inhibitor of NAD(P)H oxidase and an antioxidant, partially suppressed both EGCG-induced apoptosis and production of ROS, implying that oxidative stress is involved in the apoptotic response. Furthermore, we showed that combined-treatment with EGCG and a chemotherapeutic agent, etoposide, synergistically induced apoptosis in Ramos cells.

  2. Upregulation of TFAM and mitochondria copy number in human lymphoblastoid cells.

    PubMed

    Chakrabarty, Sanjiban; D'Souza, Reena Reshma; Kabekkodu, Shama Prasada; Gopinath, Puthiya M; Rossignol, Rodrigue; Satyamoorthy, Kapaettu

    2014-03-01

    Mitochondria are central to several physiological and pathological conditions in humans. In the present study, we performed copy number analysis of nuclear encoded mitochondrial genes, in peripheral blood mononuclear cells (PBMCs) and its representative lymphoblastoid cells (LCLs). We have observed hyper diploid copies of mitochondrial transcription factor A (TFAM) gene in the LCLs along with increased mtDNA copy number, mitochondrial mass, intracellular ROS and mitochondrial membrane potential, suggesting elevated mitochondrial biogenesis in LCLs. Gene expression analysis confirmed TFAM over-expression in LCLs when compared to PBMC. Based on our observation, we suggest that increased copy number of TFAM gene upregulates its expression, increases mtDNA copy numbers and protects it from oxidative stress induced damage in the transformed LCLs.

  3. GDF-15 gene expression alterations in human lymphoblastoid cells and peripheral blood lymphocytes following exposure to ionizing radiation

    PubMed Central

    Li, Shuang; Zhang, Qing-Zhao; Zhang, De-Qin; Feng, Jiang-Bin; Luo, Qun; Lu, Xue; Wang, Xin-Ru; Li, Kun-Peng; Chen, De-Qing; Mu, Xiao-Feng; Gao, Ling; Liu, Qing-Jie

    2017-01-01

    The identification of rapid, sensitive and high-throughput biomarkers is imperative in order to identify individuals harmed by radiation accidents, and accurately evaluate the absorbed doses of radiation. DNA microarrays have previously been used to evaluate the alterations in growth/differentiation factor 15 (GDF15) gene expression in AHH-1 human lymphoblastoid cells, following exposure to γ-rays. The present study aimed to characterize the relationship between the dose of ionizing radiation and the produced effects in GDF-15 gene expression in AHH-1 cells and human peripheral blood lymphocytes (HPBLs). GDF-15 mRNA and protein expression levels following exposure to γ-rays and neutron radiation were assessed by reverse transcription-quantitative polymerase chain reaction and western blot analysis in AHH-1 cells. In addition, alterations in GDF-15 gene expression in HPBLs following ex vivo irradiation were evaluated. The present results demonstrated that GDF-15 mRNA and protein expression levels in AHH-1 cells were significantly upregulated following exposure to γ-ray doses ranging between 1 and 10 Gy, regardless of the dose rate. A total of 48 h following exposure to neutron radiation, a dose-response relationship was identified in AHH-1 cells at γ-ray doses between 0.4 and 1.6 Gy. GDF-15 mRNA levels in HPBLs were significantly upregulated following exposure to γ-ray doses between 1 and 8 Gy, within 4–48 h following irradiation. These results suggested that significant time- and dose-dependent alterations in GDF-15 mRNA and protein expression occur in AHH-1 cells and HPBLs in the early phases following exposure to ionizing radiation. In conclusion, alterations in GDF-15 gene expression may have potential as a biomarker to evaluate radiation exposure. PMID:28440431

  4. Effects of Simulated Microgravity on the Expression Profile of Microrna in Human Lymphoblastoid Cells

    NASA Astrophysics Data System (ADS)

    Zhang, Ye; Wu, Honglu; Ramesh, Govindarajan; Rohde, Larry; Story, Michael; Mangala, Lingegowda

    2012-07-01

    EFFECTS OF SIMULATED MICROGRAVITY ON THE EXPRESSION PROFILE OF MICRORNA IN HUMAN LYMPHOBLASTOID CELLS Lingegowda S. Mangala1,2, Ye Zhang1,3, Zhenhua He2, Kamal Emami1, Govindarajan T. Ramesh4, Michael Story 5, Larry H. Rohde2, and Honglu Wu1 1 NASA Johnson Space Center, Houston, Texas, USA 2 University of Houston Clear Lake, Houston, Texas, USA 3 Wyle Integrated Science and Engineering Group, Houston, Texas, USA 4 Norfolk State University, Norfolk, VA, USA 5 University of Texas, Southwestern Medical Center, Dallas, Texas, USA This study explores the changes in expression of microRNA (miRNA) and related genes under simulated microgravity conditions. In comparison to static 1g, microgravity has been shown to alter global gene expression patterns and protein levels in cultured cells or animals. miRNA has recently emerged as an important regulator of gene expression, possibly regulating as many as one-third of all human genes. However, very little is known about the effect of altered gravity on miRNA expression. To test the hypothesis that the miRNA expression profile would be altered in zero gravity resulting in altered regulation of gene expression leading to metabolic or functional changes in cells, we cultured TK6 human lymphoblastoid cells in a High Aspect Ratio Vessel (HARV; bioreactor) for 72 h either in the rotating condition to model microgravity in space or in the static condition as a control. Expression of several miRNA was changed significantly in the simulated microgravity condition including miR-150, miR-34a, miR-423-5p, miR-22 and miR-141, miR-618 and miR-222. To confirm whether this altered miRNA expression correlates with gene expression and functional changes of the cells, we performed DNA microarray and validated the related genes using q-RT PCR. Network and pathway analysis of gene and miRNA expression profiles indicates that the regulation of cell communication and catalytic activities, as well as pathways involved in immune response_IL-15

  5. Expression of genes and proteins in human cultured lymphoblastoid cells during spaceflight

    NASA Astrophysics Data System (ADS)

    Takahashi, Akihisa; Suzuki, Hiromi; Omori, Katsunori; Seki, Masaya; Hashizume, Toko; Shimazu, Toru; Ishioka, Noriaki; Ohnishi, Takeo

    2012-07-01

    The space environment contains two major biologically significant influences: space radiations and microgravity. The aim of this study was to clarify the biological effects of space radiations, microgravity and a space environment on the gene and protein expression. Space experiments were performed with human cultured lymphoblastoid cell lines at the first life science experiment to be conducted on the Japanese Experimental Module "Kibo" of the International Space Station (ISS). Under one gravity or microgravity condition, the cells were grown in the cell biology experimental facility (CBEF) of the ISS for 8 days without experiencing the stress during launching and landing because the cells were frozen during these periods. Ground control samples also were cultured for 8 days in the CBEF on the ground during the spaceflight. Gene and protein expression was analyzed by using DNA chip (a 44k whole human genome microarray, Agilent Technologies Inc.) and protein chip (Panorama ^{TM} Ab MicroArray, Sigma-Aldrich Co.), respectively. We already reported the behavior of p53-dependent regulated genes and proteins after exposure to space radiations, microgravity, and the space environment during spaceflight. Next stage, we will profile the expression except for the p53 gene status and discuss the biological meaning during spaceflight

  6. P53 alters the cytotoxicity and genotoxicity for oxidized graphene in human B-lymphoblastoid cells

    NASA Astrophysics Data System (ADS)

    Petibone, Dayton Matthew

    Widespread use of oxidized graphene nanomaterials in industry, medicine, and consumer products raises concern about potential adverse impacts on human health. The p53 tumor suppressor protein is crucial to maintaining cellular and genetic stability to prevent carcinogenesis. Here, we show that oxygen functionalized graphene (f-G) absorption and p53 functional status correlate with cytotoxicity and genotoxicity in human B-lymphoblastoid cells. Trends in f-G absorption by were dose-dependent. Cells with functional p53 exposed to f-G arrested in G0/G1 phase of the cell cycle, suppressed f-G induced reactive oxygen species (ROS), and had elevated apoptosis. While compared to p53 competent cells, the p53 deficient cells exposed to f-G accumulated in S-phase of the cell cycle, had elevated ROS levels, and evaded apoptosis. The f-G genotoxicity was evident as increased loss-of-heterozygosity mutants independent of p53 status, and structural chromosome damage in p53 deficient cells. These findings have broad implications for the safety and efficacy of oxidized graphene nanomaterials in industrial, consumer products and biomedical applications.

  7. Arsenite-induced autophagy is associated with proteotoxicity in human lymphoblastoid cells.

    PubMed

    Bolt, Alicia M; Zhao, Fei; Pacheco, Samantha; Klimecki, Walter T

    2012-10-15

    Epidemiological studies of arsenic-exposed populations have provided evidence that arsenic exposure in humans is associated with immunosuppression. Previously, we have reported that arsenite-induced toxicity is associated with the induction of autophagy in human lymphoblastoid cell lines (LCL). Autophagy is a cellular process that functions in the degradation of damaged cellular components, including protein aggregates formed by misfolded or damaged proteins. Accumulation of misfolded or damaged proteins in the endoplasmic reticulum (ER) lumen causes ER stress and activates the unfolded protein response (UPR). In an effort to investigate the mechanism of autophagy induction by arsenite in the LCL model, we examined the potential contribution of ER stress and activation of the UPR. LCL exposed to sodium arsenite for 8-days induced expression of UPR-activated genes, including CHOP and GRP78, at the RNA and the protein level. Evidence for activation of the three arms of the UPR was observed. The arsenite-induced activation of the UPR was associated with an accumulation of protein aggregates containing p62 and LC3, proteins with established roles in the sequestration and autophagic clearance of protein aggregates. Taken together, these data provide evidence that arsenite-induced autophagy is associated with the generation of ER stress, activation of the UPR, and formation of protein aggregates that may be targeted to the lysosome for degradation.

  8. Proliferation-dependent positioning of individual centromeres in the interphase nucleus of human lymphoblastoid cell lines.

    PubMed

    Ollion, Jean; Loll, François; Cochennec, Julien; Boudier, Thomas; Escudé, Christophe

    2015-07-01

    The cell nucleus is a highly organized structure and plays an important role in gene regulation. Understanding the mechanisms that sustain this organization is therefore essential for understanding genome function. Centromeric regions (CRs) of chromosomes have been known for years to adopt specific nuclear positioning patterns, but the significance of this observation is not yet completely understood. Here, using a combination of fluorescence in situ hybridization and immunochemistry on fixed human cells and high-throughput imaging, we directly and quantitatively investigated the nuclear positioning of specific human CRs. We observe differential attraction of individual CRs toward both the nuclear border and the nucleoli, the former being enhanced in nonproliferating cells and the latter being enhanced in proliferating cells. Similar positioning patterns are observed in two different lymphoblastoid cell lines. Moreover, the positioning of CRs differs from that of noncentromeric regions, and CRs display specific orientations within chromosome territories. These results suggest the existence of not-yet-characterized mechanisms that drive the nuclear positioning of CRs and therefore pave the way toward a better understanding of how CRs affect nuclear organization.

  9. The TP53 dependence of radiation-induced chromosome instability in human lymphoblastoid cells

    NASA Technical Reports Server (NTRS)

    Schwartz, Jeffrey L.; Jordan, Robert; Evans, Helen H.; Lenarczyk, Marek; Liber, Howard

    2003-01-01

    The dose and TP53 dependence for the induction of chromosome instability were examined in cells of three human lymphoblastoid cell lines derived from WIL2 cells: TK6, a TP53-normal cell line, NH32, a TP53-knockout created from TK6, and WTK1, a WIL2-derived cell line that spontaneously developed a TP53 mutation. Cells of each cell line were exposed to (137)Cs gamma rays, and then surviving clones were isolated and expanded in culture for approximately 35 generations before the frequency and characteristics of the instability were analyzed. The presence of dicentric chromosomes, formed by end-to-end fusions, served as a marker of chromosomal instability. Unexposed TK6 cells had low levels of chromosomal instability (0.002 +/- 0.001 dicentrics/cell). Exposure of TK6 cells to doses as low as 5 cGy gamma rays increased chromosome instability levels nearly 10-fold to 0.019 +/- 0.008 dicentrics/cell. There was no further increase in instability levels beyond 5 cGy. In contrast to TK6 cells, unexposed cultures of WTK1 and NH32 cells had much higher levels of chromosome instability of 0.034 +/- 0.007 and 0.041 +/- 0.009, respectively, but showed little if any effect of radiation on levels of chromosome instability. The results suggest that radiation exposure alters the normal TP53-dependent cell cycle checkpoint controls that recognize alterations in telomere structure and activate apoptosis.

  10. The TP53 dependence of radiation-induced chromosome instability in human lymphoblastoid cells

    NASA Technical Reports Server (NTRS)

    Schwartz, Jeffrey L.; Jordan, Robert; Evans, Helen H.; Lenarczyk, Marek; Liber, Howard

    2003-01-01

    The dose and TP53 dependence for the induction of chromosome instability were examined in cells of three human lymphoblastoid cell lines derived from WIL2 cells: TK6, a TP53-normal cell line, NH32, a TP53-knockout created from TK6, and WTK1, a WIL2-derived cell line that spontaneously developed a TP53 mutation. Cells of each cell line were exposed to (137)Cs gamma rays, and then surviving clones were isolated and expanded in culture for approximately 35 generations before the frequency and characteristics of the instability were analyzed. The presence of dicentric chromosomes, formed by end-to-end fusions, served as a marker of chromosomal instability. Unexposed TK6 cells had low levels of chromosomal instability (0.002 +/- 0.001 dicentrics/cell). Exposure of TK6 cells to doses as low as 5 cGy gamma rays increased chromosome instability levels nearly 10-fold to 0.019 +/- 0.008 dicentrics/cell. There was no further increase in instability levels beyond 5 cGy. In contrast to TK6 cells, unexposed cultures of WTK1 and NH32 cells had much higher levels of chromosome instability of 0.034 +/- 0.007 and 0.041 +/- 0.009, respectively, but showed little if any effect of radiation on levels of chromosome instability. The results suggest that radiation exposure alters the normal TP53-dependent cell cycle checkpoint controls that recognize alterations in telomere structure and activate apoptosis.

  11. Genome-wide survey of interindividual differences of RNA stability in human lymphoblastoid cell lines

    PubMed Central

    Duan, Jubao; Shi, Jianxin; Ge, Xijin; Dölken, Lars; Moy, Winton; He, Deli; Shi, Sandra; Sanders, Alan R.; Ross, Jeff; Gejman, Pablo V.

    2013-01-01

    The extent to which RNA stability differs between individuals and its contribution to the interindividual expression variation remain unknown. We conducted a genome-wide analysis of RNA stability in seven human HapMap lymphoblastoid cell lines (LCLs) and analyzed the effect of DNA sequence variation on RNA half-life differences. Twenty-six percent of the expressed genes exhibited RNA half-life differences between LCLs at a false discovery rate (FDR) < 0.05, which accounted for ~ 37% of the gene expression differences between individuals. Nonsense polymorphisms were associated with reduced RNA half-lives. In genes presenting interindividual RNA half-life differences, higher coding GC3 contents (G and C percentages at the third-codon positions) were correlated with increased RNA half-life. Consistently, G and C alleles of single nucleotide polymorphisms (SNPs) in protein coding sequences were associated with enhanced RNA stability. These results suggest widespread interindividual differences in RNA stability related to DNA sequence and composition variation. PMID:23422947

  12. Arsenite exposure in human lymphoblastoid cell lines induces autophagy and coordinated induction of lysosomal genes.

    PubMed

    Bolt, Alicia M; Douglas, Randi M; Klimecki, Walter T

    2010-11-30

    Chronic exposure to inorganic arsenic is associated with diverse, complex diseases, making the identification of the mechanism underlying arsenic-induced toxicity a challenge. An increasing body of literature from epidemiological and in vitro studies has demonstrated that arsenic is an immunotoxicant, but the mechanism driving arsenic-induced immunotoxicity is not well established. We have previously demonstrated that in human lymphoblastoid cell lines (LCLs), arsenic-induced cell death is strongly associated with the induction of autophagy. In this study we utilized genome-wide gene expression analysis and functional assays to characterize arsenic-induced effects in seven LCLs that were exposed to an environmentally relevant, minimally cytotoxic, concentration of arsenite (0.75 μM) over an eight-day time course. Arsenic exposure resulted in inhibition of cellular growth and induction of autophagy (measured by expansion of acidic vesicles) over the eight-day exposure duration. Gene expression analysis revealed that arsenic exposure increased global lysosomal gene expression, which was associated with increased functional activity of the lysosome protease, cathepsin D. The arsenic-induced expansion of the lysosomal compartment in LCL represents a novel target that may offer insight into the immunotoxic effects of arsenic.

  13. The impact of FANCD2 deficiency on formaldehyde-induced toxicity in human lymphoblastoid cell lines.

    PubMed

    Ren, Xuefeng; Ji, Zhiying; McHale, Cliona M; Yuh, Jessica; Bersonda, Jessica; Tang, Maycky; Smith, Martyn T; Zhang, Luoping

    2013-01-01

    Formaldehyde (FA), a major industrial chemical and ubiquitous environmental pollutant, has recently been classified by the International Agency for Research on Cancer as a human leukemogen. The major mode of action of FA is thought to be the formation of DNA-protein cross-links (DPCs). Repair of DPCs may be mediated by the Fanconi anemia pathway; however, data supporting the involvement of this pathway are limited, particularly in human hematopoietic cells. Therefore, we assessed the role of FANCD2, a critical component of the Fanconi anemia pathway, in FA-induced toxicity in human lymphoblast cell models of FANCD2 deficiency (PD20 cells) and FANCD2 sufficiency (PD20-D2 cells). After treatment of the cells with 0-150 μM FA for 24 h, DPCs were increased in a dose-dependent manner in both cell lines, with greater increases in FANCD2-deficient PD20 cells. FA also induced cytotoxicity, micronuclei, chromosome aberrations, and apoptosis in a dose-dependent manner in both cell lines, with greater increases in cytotoxicity and apoptosis in PD20 cells. Increased levels of γ-ATR and γ-H2AX in both cell lines suggested the recognition of FA-induced DNA damage; however, the induction of BRCA2 was compromised in FANCD2-deficient PD20 cells, potentially reducing the capacity to repair DPCs. Together, these findings suggest that FANCD2 protein and the Fanconi anemia pathway are essential to protect human lymphoblastoid cells against FA toxicity. Future studies are needed to delineate the role of this pathway in mitigating FA-induced toxicity, particularly in hematopoietic stem cells, the target cells in leukemia.

  14. Integration sites of Epstein-Barr virus genome on chromosomes of human lymphoblastoid cell lines

    SciTech Connect

    Wuu, K.D.; Chen, Y.J.; Wang-Wuu, S.

    1994-09-01

    Epstein-Barr virus (EBV) is the pathogen of infectious mononucleosis. The viral genome is present in more than 95% of the African cases of Burkitt lymphoma and it is usually maintained in episomal form in the tumor cells. Viral integration has been described only for Nanalwa which is a Burkitt lymphoma cell line lacking episomes. In order to examine the role of EBV in the immortalization of human Blymphocytes, we investigated whether the EBV integration into the human genome is essential. If the integration does occur, we would like to know whether the integration is randomly distributed or whether the viral DNA integrates preferentially at certain sites. Fourteen in vitro immortalized human lymphoblastoid cell lines (LCLs) were examined by fluorescence in situ hybridization (FISH) with a biotinylated EBV BamHI w DNA fragment as probe. The episomal form of EBV DNA was found in all cells of these cell lines, while only about 65% of the cells have the integrated viral DNA. This might suggest that integration is not a pre-requisite for cell immortalization. Although all chromosomes, except Y, have been found with integrated viral genome, chromsomes 1 and 5 are the most frequent EBV DNA carrier (p<0.05). Nine chromosome bands, namely, 1p31, 1q31, 2q32, 3q13, 3q26, 5q14, 6q24, 7q31 and 12q21, are preferential targets for EBV integration (p<0.001). Eighty percent of the total 938 EBV hybridization signals were found to be at G-band-positive area. This suggests that the mechanism of EBV integration might be different from that of the retroviruses, which specifically integrate to G-band-negative areas. Thus, we conclude that the integration of EBV to host genome is non-random and it may have something to do with the structure of chromosome and DNA sequences.

  15. Arsenite-induced autophagy is associated with proteotoxicity in human lymphoblastoid cells

    SciTech Connect

    Bolt, Alicia M.; Zhao, Fei; Pacheco, Samantha; Klimecki, Walter T.

    2012-10-15

    Epidemiological studies of arsenic-exposed populations have provided evidence that arsenic exposure in humans is associated with immunosuppression. Previously, we have reported that arsenite-induced toxicity is associated with the induction of autophagy in human lymphoblastoid cell lines (LCL). Autophagy is a cellular process that functions in the degradation of damaged cellular components, including protein aggregates formed by misfolded or damaged proteins. Accumulation of misfolded or damaged proteins in the endoplasmic reticulum (ER) lumen causes ER stress and activates the unfolded protein response (UPR). In an effort to investigate the mechanism of autophagy induction by arsenite in the LCL model, we examined the potential contribution of ER stress and activation of the UPR. LCL exposed to sodium arsenite for 8-days induced expression of UPR-activated genes, including CHOP and GRP78, at the RNA and the protein level. Evidence for activation of the three arms of the UPR was observed. The arsenite-induced activation of the UPR was associated with an accumulation of protein aggregates containing p62 and LC3, proteins with established roles in the sequestration and autophagic clearance of protein aggregates. Taken together, these data provide evidence that arsenite-induced autophagy is associated with the generation of ER stress, activation of the UPR, and formation of protein aggregates that may be targeted to the lysosome for degradation. -- Highlights: ► Arsenite induces endoplasmic reticulum stress and the unfolded protein response. ► Arsenite induces the formation of protein aggregates that contain p62 and LC3-II. ► Time-course data suggests that arsenite-induced autophagy precedes ER stress.

  16. A combination of in vitro comet assay and micronucleus test using human lymphoblastoid TK6 cells.

    PubMed

    Kimura, Aoi; Miyata, Atsuro; Honma, Masamitsu

    2013-09-01

    The comet assay has been widely used as a genotoxicity test for detecting primary DNA damage in individual cells. The micronucleus (MN) test is also a well-established assay for detecting clastogenicity and aneugenicity. A combination of the comet assay (COM) and MN test is capable of detecting a variety of genotoxic potentials as an in vitro screening system. Although the in vitro MN test has a robust protocol and Organisation for Economic Co-operation and Development (OECD) test guideline, the in vitro COM does not. To establish a robust protocol for the COM and to compare its sensitivity with that of the MN, we conducted COM and MN concurrently for five genotoxic agents (ethyl methanesulfonate, methyl methanesulfonate, hydrogen peroxide, gamma-rays and mitomycin C) and one non-genotoxic agent (triton X-100), using human lymphoblastoid TK6 cells. Relative cell count (RCC), relative population doubling (RPD), relative increase in cell count (RICC) and relative cell viability determined by trypan blue dye-exclusion assay (TBDE) were employed as cytotoxic measurements. However, the relative cell viability determined by TBDE just after the treatment was not an appropriate parameter of cytotoxicity for the genotoxic agents because it remained constant even at the highest doses, which showed severe cytotoxicity by RCC, RPD and RICC. The results of the COM showed qualitative agreement (positive or negative) with those of the MN except for mitomycin C, which is an interstrand cross-linker. The COM always required higher doses than the MN to detect the genotoxic potential of the genotoxic agents under the test conditions applied here. The doses that induced a comet tail always yielded <50% RICC, and do not accord to the OECD test guideline for MN because of their high cytotoxicity. These results are helpful for interpreting the results of the COM and MN in in vitro genotoxic hazard assessments. Further investigation is required to standardise the COM.

  17. Effects of Modeled Microgravity on Expression Profiles of Micro RNA in Human Lymphoblastoid Cells

    NASA Technical Reports Server (NTRS)

    Mangala, Lingegowda S.; Emami, Kamal; Story, Michael; Ramesh, Govindarajan; Rohde, Larry; Wu, Honglu

    2010-01-01

    Among space radiation and other environmental factors, microgravity or an altered gravity is undoubtedly the most significant stress experienced by living organisms during flight. In comparison to the static 1g, microgravity has been shown to alter global gene expression patterns and protein levels in cultured cells or animals. Micro RNA (miRNA) has recently emerged as an important regulator of gene expression, possibly regulating as many as one-third of all human genes. miRNA represents a class of single-stranded noncoding regulatory RNA molecules ( 22 nt) that control gene expressions by inhibiting the translation of mRNA to proteins. However, very little is known on the effect of altered gravity on miRNA expression. We hypothesized that the miRNA expression profile will be altered in zero gravity resulting in regulation of the gene expression and functional changes of the cells. To test this hypothesis, we cultured TK6 human lymphoblastoid cells in Synthecon s Rotary cell culture system (bioreactors) for 72 h either in the rotating (10 rpm) to model the microgravity in space or in the static condition. The cell viability was determined before and after culturing the cells in the bioreactor using both trypan blue and guava via count. Expressions of a panel of 352 human miRNA were analyzed using the miRNA PCRarray. Out of 352 miRNAs, expressions of 75 were significantly altered by a change of greater than 1.5 folds and seven miRNAs were altered by a fold change greater than 2 under the rotating culture condition. Among these seven, miR-545 and miR-517a were down regulated by 2 folds, whereas miR-150, miR-302a, miR-139-3p, miR-515-3p and miR-564 were up regulated by 2 to 8 folds. To confirm whether this altered miRNA expression correlates with gene expression and functional changes of the cells, we performed DNA Illumina Microarray Analysis and validated the related genes using q-RT PCR.

  18. Effects of soluble and particulate Cr(VI) on genome-wide DNA methylation in human B lymphoblastoid cells.

    PubMed

    Lou, Jianlin; Wang, Yu; Chen, Junqiang; Ju, Li; Yu, Min; Jiang, Zhaoqiang; Feng, Lingfang; Jin, Lingzhi; Zhang, Xing

    2015-10-01

    Several previous studies highlighted the potential epigenetic effects of Cr(VI), especially DNA methylation. However, few studies have compared the effects of Cr(VI) on DNA methylation profiles between soluble and particulate chromate in vitro. Accordingly, Illumina Infinium Human Methylation 450K BeadChip array was used to analyze DNA methylation profiles of human B lymphoblastoid cells exposed to potassium dichromate or lead chromate, and the cell viability was also studied. Array based DNA methylation analysis showed that the impacts of Cr(VI) on DNA methylation were limited, only about 40 differentially methylated CpG sites, with an overlap of 15CpG sites, were induced by both potassium dichromate and lead chromate. The results of mRNA expression showed that after Cr(VI) treatment, mRNA expression changes of four genes (TBL1Y, FZD5, IKZF2, and KIAA1949) were consistent with their DNA methylation alteration, but DNA methylation changes of other six genes did not correlate with mRNA expression. In conclusion, both of soluble and particulate Cr(VI) could induce a small amount of differentially methylated sites in human B lymphoblastoid cells, and the correlations between DNA methylation changes and mRNA expression varied between different genes. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Effects of inorganic and organic arsenic compounds on growth and apoptosis of human T-lymphoblastoid leukemia cells.

    PubMed

    Hikita, Eri; Arai, Mariko; Tanaka, Sachiko; Onda, Kenji; Utsumi, Hiroya; Yuan, Bo; Toyoda, Hiroo; Hirano, Toshihiko

    2011-12-01

    To investigate the effects of inorganic and organic arsenic compounds on human T-lymphoblastoid leukemia cells. Cell proliferation was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5¬diphenyltetrazolium bromide (MTT) assay. Apoptotic cell morphology was examined by cell staining with Hoechst 33342. Cellular caspase-3/7 activities were measured after arsenic treatment. The inhibitory concentration by 50% (IC(50)) values of As(2)O(3) towards MOLT-4 and daunorubicin- resistant MOLT-4/DNR cell proliferation were 0.87 and 0.92 μM, while the values for arsenic acid were 69.1 and 116.6 μM, respectively. These arsenic compounds also inhibited mitogen-induced proliferation of human peripheral blood mononuclear cells. Six organic arsenic compounds did not inhibit leukemia cell proliferation. As(2)O(3) and arsenic acid induced apoptotic cell morphology and increased caspase-3/7 activity in the leukemia cells. Ascorbic acid and buthionine sulfoxide enhanced, while N-acetyl-L-cysteine abated, the suppressive effects of inorganic arsenic compounds on leukemia cell proliferation. As(2)O(3) and arsenic acid inhibit proliferation and induce apoptosis in MOLT-4 and daunorubicine-resistant MOLT-4/DNR cells via glutathione-depletion and subsequent caspase-3/7 activation. Organic arsenic compounds have no inhibitory activity on the leukemia cell proliferation. Inorganic arsenic compounds are suggested as useful agents for treatment of T-lymphoblastoid leukemia.

  20. Low Dose Radiation Response Curves, Networks and Pathways in Human Lymphoblastoid Cells Exposed from 1 to 10 cGy of Acute Gamma Radiation

    SciTech Connect

    Wyrobek, A. J.; Manohar, C. F.; Nelson, D. O.; Furtado, M. R.; Bhattacharya, M. S.; Marchetti, F.; Coleman, M.A.

    2011-04-18

    We investigated the low dose dependency of the transcriptional response of human cells to characterize the shape and biological functions associated with the dose response curve and to identify common and conserved functions of low dose expressed genes across cells and tissues. Human lymphoblastoid (HL) cells from two unrelated individuals were exposed to graded doses of radiation spanning the range of 1-10 cGy were analyzed by transcriptome profiling, qPCR and bioinformatics, in comparison to sham irradiated samples. A set of {approx}80 genes showed consistent responses in both cell lines; these genes were associated with homeostasis mechanisms (e.g., membrane signaling, molecule transport), subcellular locations (e.g., Golgi, and endoplasmic reticulum), and involved diverse signal transduction pathways. The majority of radiation-modulated genes had plateau-like responses across 1-10 cGy, some with suggestive evidence that transcription was modulated at doses below 1 cGy. MYC, FOS and TP53 were the major network nodes of the low-dose response in HL cells. Comparison our low dose expression findings in HL cells with those of prior studies in mouse brain after whole body exposure, in human keratinocyte cultures, and in endothelial cells cultures, indicates that certain components of the low dose radiation response are broadly conserved across cell types and tissues, independent of proliferation status.

  1. Low dose radiation response curves, networks and pathways in human lymphoblastoid cells exposed from 1 to 10cGy of acute gamma radiation.

    PubMed

    Wyrobek, A J; Manohar, C F; Krishnan, V V; Nelson, D O; Furtado, M R; Bhattacharya, M S; Marchetti, F; Coleman, M A

    2011-06-17

    We investigated the low dose dependency of the transcriptional response of human cells to characterize the shape and biological functions associated with the dose-response curve and to identify common and conserved functions of low dose expressed genes across cells and tissues. Human lymphoblastoid (HL) cells from two unrelated individuals were exposed to graded doses of radiation spanning the range of 1-10cGy were analyzed by transcriptome profiling, qPCR and bioinformatics, in comparison to sham irradiated samples. A set of ∼80 genes showed consistent responses in both cell lines; these genes were associated with homeostasis mechanisms (e.g., membrane signaling, molecule transport), subcellular locations (e.g., Golgi, and endoplasmic reticulum), and involved diverse signal transduction pathways. The majority of radiation-modulated genes had plateau-like responses across 1-10cGy, some with suggestive evidence that transcription was modulated at doses below 1cGy. MYC, FOS and TP53 were the major network nodes of the low-dose-response in HL cells. Comparison our low dose expression findings in HL cells with those of prior studies in mouse brain after whole body exposure, in human keratinocyte cultures, and in endothelial cells cultures, indicates that certain components of the low dose radiation response are broadly conserved across cell types and tissues, independent of proliferation status. Copyright © 2011 Elsevier B.V. All rights reserved.

  2. Human NK cells activated by EBV+ lymphoblastoid cells overcome anti-apoptotic mechanisms of drug resistance in haematological cancer cells

    PubMed Central

    Sánchez-Martínez, Diego; Azaceta, Gemma; Muntasell, Aura; Aguiló, Nacho; Núñez, David; Gálvez, Eva M; Naval, Javier; Anel, Alberto; Palomera, Luis; Vilches, Carlos; Marzo, Isabel; Villalba, Martín; Pardo, Julián

    2015-01-01

    Natural killer (NK) cells recognize and eliminate transformed or infected cells that have downregulated MHC class-I and express specific activating ligands. Recent evidence indicates that allogeneic NK cells are useful to eliminate haematological cancer cells independently of MHC-I expression. However, it is unclear if transformed cells expressing mutations that confer anti-apoptotic properties and chemoresistance will be susceptible to NK cells. Allogeneic primary human NK cells were activated using different protocols and prospectively tested for their ability to eliminate diverse mutant haematological and apoptotic-resistant cancer cell lines as well as patient-derived B-cell chronic lymphocytic leukemia cells with chemotherapy multiresistance. Here, we show that human NK cells from healthy donors activated in vitro with Epstein Barr virus positive (EBV+)-lymphoblastoid cells display an enhanced cytotoxic and proliferative potential in comparison to other protocols of activation such a K562 cells plus interleukin (IL)2. This enhancement enables them to kill more efficiently a variety of haematological cancer cell lines, including a panel of transfectants that mimic natural mutations leading to oncogenic transformation and chemoresistance (e.g., overexpression of Bcl-2, Bcl-XL and Mcl-1 or downregulation of p53, Bak/Bax or caspase activity). The effect was also observed against blasts from B-cell chronic lymphocytic leukemia patients showing multi-resistance to chemotherapy. Our findings demonstrate that particular in vitro activated NK cells may overcome anti-apoptotic mechanisms and oncogenic alterations frequently occurring in transformed cells, pointing toward the use of EBV+-lymphoblastoid cells as a desirable strategy to activate NK cells in vitro for the purpose of treating haematological neoplasia with poor prognosis. PMID:25949911

  3. IsomiR expression profiles in human lymphoblastoid cell lines exhibit population and gender dependencies.

    PubMed

    Loher, Phillipe; Londin, Eric R; Rigoutsos, Isidore

    2014-09-30

    For many years it was believed that each mature microRNA (miRNA) existed as a single entity with fixed endpoints and a 'static' and unchangeable primary sequence. However, recent evidence suggests that mature miRNAs are more 'dynamic' and that each miRNA precursor arm gives rise to multiple isoforms, the isomiRs. Here we report on our identification of numerous and abundant isomiRs in the lymphoblastoid cell lines (LCLs) of 452 men and women from five different population groups. Unexpectedly, we find that these isomiRs exhibit an expression profile that is population-dependent and gender-dependent. This is important as it indicates that the LCLs of each gender/population combination have their own unique collection of mature miRNA transcripts. Moreover, each identified isomiR has its own characteristic abundance that remains consistent across biological replicates indicating that these are not degradation products. The primary sequences of identified isomiRs differ from the known miRBase miRNA either at their 5´-endpoint (leads to a different 'seed' sequence and suggests a different targetome), their 3´-endpoint, or both simultaneously. Our analysis of Argonaute PAR-CLIP data from LCLs supports the association of many of these newly identified isomiRs with the Argonaute silencing complex and thus their functional roles through participation in the RNA interference pathway.

  4. Cell culture-induced aberrant methylation of the imprinted IG DMR in human lymphoblastoid cell lines.

    PubMed

    Saferali, Aabida; Grundberg, Elin; Berlivet, Soizik; Beauchemin, Hugues; Morcos, Lisanne; Polychronakos, Constantin; Pastinen, Tomi; Graham, Jinko; McNeney, Brad; Naumova, Anna K

    2010-01-01

    DNA methylation patterns are often poorly conserved through cell culturing. To determine the effect of cell immortalization and culture on DNA methylation profiles, we analyzed methylation in the differentially methylated regions (DMR) of five imprinted domains: the intergenic (IG) DMR on chromosome 14q32; potassium voltage-gated channel, KQT-like subfamily, member 1, (KCNQ1); small nuclear ribonucleoprotein polypeptide N (SNRPN), mesoderm specific transcript homolog (MEST); and H19 in lymphoblastoid cell lines (LCLs). In the IG DMR we found an aberrant methylation pattern that was consistent through all the cell lines tested and significantly different from that of noncultured peripheral blood cells. Using a generalized linear mixed model to compare methylation profiles, we show that recently derived LCLs significantly differ from the CEPH LCLs. This implies a gradual cell-culture related deterioration of DNA methylation in the IG DMR with at least two steps that may be identified: loss of methylation at CG sites 1 and 8; and loss of allelic differences in DNA methylation. The IG DMR methylation profile also confirms the high level of clonality of the CEPH LCLs. We conclude that non-transformed primary cells may be less susceptible to epigenetic anomalies and therefore may provide a more accurate reflection of gene expression in vivo.

  5. IsomiR expression profiles in human lymphoblastoid cell lines exhibit population and gender dependencies

    PubMed Central

    Loher, Phillipe; Londin, Eric R.; Rigoutsos, Isidore

    2014-01-01

    For many years it was believed that each mature microRNA (miRNA) existed as a single entity with fixed endpoints and a ‘static’ and unchangeable primary sequence. However, recent evidence suggests that mature miRNAs are more ‘dynamic’ and that each miRNA precursor arm gives rise to multiple isoforms, the isomiRs. Here we report on our identification of numerous and abundant isomiRs in the lymphoblastoid cell lines (LCLs) of 452 men and women from five different population groups. Unexpectedly, we find that these isomiRs exhibit an expression profile that is population-dependent and gender-dependent. This is important as it indicates that the LCLs of each gender/population combination have their own unique collection of mature miRNA transcripts. Moreover, each identified isomiR has its own characteristic abundance that remains consistent across biological replicates indicating that these are not degradation products. The primary sequences of identified isomiRs differ from the known miRBase miRNA either at their 5´-endpoint (leads to a different ‘seed’ sequence and suggests a different targetome), their 3´-endpoint, or both simultaneously. Our analysis of Argonaute PAR-CLIP data from LCLs supports the association of many of these newly identified isomiRs with the Argonaute silencing complex and thus their functional roles through participation in the RNA interference pathway. PMID:25229428

  6. Characterization of human lymphoblastoid cell lines as a novel in vitro test system to predict the immunotoxicity of xenobiotics.

    PubMed

    Markovič, Tijana; Gobec, Martina; Gurwitz, David; Mlinarič-Raščan, Irena

    2015-02-17

    Evaluating immunomodulatory effects of xenobiotics is an important component of the toxicity studies. Herein we report on the establishment of a novel invitro test system for the immunotoxicity screening of xenobiotics based on human lymphoblastoid cell lines (LCLs). Four immunotoxic compounds; tributyltin chloride, cyclosporine A, benzo(a)pyrene and verapamil hydrochloride, as well as three immune-inert compounds; urethane, furosemide and mannitol were selected for characterization. The treatment of LCLs with immunosuppressive compounds resulted in reduced viability. The IC50 values determined in human LCLs were in agreement with the data obtained for human peripheral mononuclear cells. Since cytokine production reflects lymphocytes responses to external stimuli, we evaluated the functional responses of LCLs by monitoring their pro-inflammatory and immunoregulatory cytokine production. Our findings prove that LCLs allowed for reliable differentiation between immunomodulatory and immune-inert compounds. Hence, pre-treatment with immunomodulatory compounds led to a decrease in the production of pro-inflammatory TNFα, IL-6 and immunoregulatory IL-2, IL-4, IL-10 and IFNγ cytokines, when compared to untreated ionomycin/PMA stimulated cells. Moreover, testing a panel of ten LCLs derived from unrelated healthy individuals reflects inter-individual variability in response to immunomodulatory xenobiotics. In conclusion, LCLs provide a novel alternative method for the testing of the immunotoxic effects of xenobiotics.

  7. Modulation of the DNA damage response in UV-exposed human lymphoblastoid cells through genetic-versus functional-inactivation of the p53 tumor suppressor.

    PubMed

    Léger, Caroline; Drobetsky, Elliot A

    2002-10-01

    The global cellular response to UV-induced DNA damage has been analyzed in the p53-proficient human lymphoblastoid strain TK6 versus two isogenic derivatives wherein p53 activity was abrogated by diverse experimental approaches: (i) NH32, carrying a homozygous genetic knockout of p53; and (ii) TK6-5E, expressing the human papillomavirus E6 oncoprotein which binds and functionally inactivates p53 protein. Although widely employed as such, the extent to which intracellular E6 expression faithfully models the p53 deficient state still remains uncertain. Following irradiation with UV (either monochromatic 254 nm UV or broad-spectrum simulated sunlight), relative to wild-type TK6, p53-null NH32 exhibited virtually identical clonogenic survival and kinetics of G1-S progression but was nonetheless profoundly resistant to apoptosis. In addition, there were significant qualitative and quantitative differences between NH32 and TK6 with respect to UV mutagenesis at the endogenous hypoxanthine phosphoribosyltransferase (hprt) locus. However, important disparities were observed between genetically p53-deficient NH32 and E6-expressing TK6-5E regarding the manner in which they responded to UV-induced genotoxic stress in relation to wild-type TK6. Indeed, although NH32 and TK6-5E behaved similarly with respect to UV mutagenesis at the hprt locus, there were significant differences between these strains in clonogenic survival, apoptosis, and G1-S progression. Using a well-defined isogenic system, our data clearly reveal the influence of p53 inactivation on the global response of human cells to UV-induced DNA damage, and highlight an important caveat in the field of p53 biology by directly demonstrating that this influence varies substantially depending upon whether p53 function is abrogated genetically, or through E6 oncoprotein expression.

  8. Genetic instability in lymphoblastoid cell lines expressing biallelic and monoallelic variants in the human MUTYH gene.

    PubMed

    Grasso, Francesca; Giacomini, Elisa; Sanchez, Massimo; Degan, Paolo; Gismondi, Viviana; Mazzei, Filomena; Varesco, Liliana; Viel, Alessandra; Bignami, Margherita

    2014-07-15

    The MUTYH DNA glycosylase counteracts mutagenesis by removing adenine misincorporated opposite DNA 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG). Biallelic germline mutations in MUTYH cause the autosomal recessive MUTYH-associated polyposis (MAP). The impact on genetic instability of the p.Tyr179Cys and p.Arg245His MUTYH variants was evaluated in lymphoblastoid cell lines (LCLs) derived from MAP patients and their relatives in comparison to wild-type LCLs. No difference in MUTYH expression was identified between wild type and LCLs with the p.Tyr179Cys, while the p.Arg245His mutation was associated with an unstable MUTYH protein. LCLs homozygous for the p.Tyr179Cys or the p.Arg245His variant contained increased DNA 8-oxodG levels and exhibited a mutator phenotype at the PIG-A gene. The extent of the increased spontaneous mutation frequency was 3-fold (range 1.6- to 4.6-fold) in four independent LCLs carrying the p.Tyr179Cys variant, while a larger increase (6-fold) was observed in two p.Arg245His LCLs. A similar hypermutability and S-phase delay following treatment with KBrO3 was observed in LCLs homozygous for either variant. When genetic instability was investigated in monoallelic p.Arg245His carriers, mutant frequencies showed an increase which is intermediate between wild-type and homozygous cells, whereas the mutator effect in heterozygous p.Tyr179Cys LCLs was similar to that in homozygotes. These findings indicate that the type of MUTYH mutation can affect the extent of genome instability associated with MUTYH inactivation. In addition, the mild spontaneous mutator phenotype observed in monoallelic carriers highlights the biological importance of this gene in the protection of the genome against endogenous DNA damage. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  9. Constant splice-isoform ratios in human lymphoblastoid cells support the concept of a splico-stat.

    PubMed

    Kramer, Marcel; Huse, Klaus; Menzel, Uwe; Backhaus, Oliver; Rosenstiel, Philip; Schreiber, Stefan; Hampe, Jochen; Platzer, Matthias

    2011-03-01

    Splicing generates mature transcripts from genes in pieces in eukaryotic cells. Overwhelming evidence has accumulated that alternative routes in splicing are possible for most human and mammalian genes, thereby allowing formation of different transcripts from one gene. No function has been assigned to the majority of identified alternative splice forms, and it has been assumed that they compose inert or tolerated waste from aberrant or noisy splicing. Here we demonstrate that five human transcription units (WT1, NOD2, GNAS, RABL2A, RABL2B) have constant splice-isoform ratios in genetically diverse lymphoblastoid cell lines independent of the type of alternative splicing (exon skipping, alternative donor/acceptor, tandem splice sites) and gene expression level. Even splice events that create premature stop codons and potentially trigger nonsense-mediated mRNA decay are found at constant fractions. The analyzed alternative splicing events were qualitatively but not quantitatively conserved in corresponding chimpanzee cell lines. Additionally, subtle splicing at tandem acceptor splice sites (GNAS, RABL2A/B) was highly constrained and strongly depends on the upstream donor sequence content. These results also demonstrate that unusual and unproductive splice variants are produced in a regulated manner. © 2011 by the Genetics Society of America

  10. Impact of 1.8-GHz radiofrequency radiation (RFR) on DNA damage and repair induced by doxorubicin in human B-cell lymphoblastoid cells.

    PubMed

    Zhijian, Chen; Xiaoxue, Li; Yezhen, Lu; Shijie, Chen; Lifen, Jin; Jianlin, Lou; Deqiang, Lu; Jiliang, He

    2010-01-01

    In the present in vitro study, a comet assay was used to determine whether 1.8-GHz radiofrequency radiation (RFR, SAR of 2W/kg) can influence DNA repair in human B-cell lymphoblastoid cells exposed to doxorubicin (DOX) at the doses of 0microg/ml, 0.05microg/ml, 0.075microg/ml, 0.10microg/ml, 0.15microg/ml and 0.20microg/ml. The combinative exposures to RFR with DOX were divided into five categories. DNA damage was detected at 0h, 6h, 12h, 18h and 24h after exposure to DOX via the comet assay, and the percent of DNA in the tail (% tail DNA) served as the indicator of DNA damage. The results demonstrated that (1) RFR could not directly induce DNA damage of human B-cell lymphoblastoid cells; (2) DOX could significantly induce DNA damage of human B-cell lymphoblastoid cells with the dose-effect relationship, and there were special repair characteristics of DNA damage induced by DOX; (3) E-E-E type (exposure to RFR for 2h, then simultaneous exposure to RFR and DOX, and exposure to RFR for 6h, 12h, 18h and 24h after exposure to DOX) combinative exposure could obviously influence DNA repair at 6h and 12h after exposure to DOX for four DOX doses (0.075microg/ml, 0.10microg/ml, 0.15microg/ml and 0.20microg/ml) in human B-cell lymphoblastoid cells. Copyright 2009 Elsevier B.V. All rights reserved.

  11. Biomarkers of Exposure and Effect in Human Lymphoblastoid TK6 Cells Following [13C2]-Acetaldehyde Exposure

    PubMed Central

    Swenberg, James A.

    2013-01-01

    The dose-response relationship for biomarkers of exposure (N2-ethylidene-dG adducts) and effect (cell survival and micronucleus formation) was determined across 4.5 orders of magnitude (50nM–2mM) using [13C2]-acetaldehyde exposures to human lymphoblastoid TK6 cells for 12h. There was a clear increase in exogenous N 2-ethylidene-dG formation at exposure concentrations ≥ 1µM, whereas the endogenous adducts remained nearly constant across all exposure concentrations, with an average of 3.0 adducts/107 dG. Exogenous adducts were lower than endogenous adducts at concentrations ≤ 10µM and were greater than endogenous adducts at concentrations ≥ 250µM. When the endogenous and exogenous adducts were summed together, statistically significant increases in total adduct formation over the endogenous background occurred at 50µM. Cell survival and micronucleus formation were monitored across the exposure range and statistically significant decreases in cell survival and increases in micronucleus formation occurred at ≥ 1000µM. This research supports the hypothesis that endogenously produced reactive species, including acetaldehyde, are always present and constitute the majority of the observed biological effects following very low exposures to exogenous acetaldehyde. These data can replace default assumptions of linear extrapolation to very low doses of exogenous acetaldehyde for risk prediction. PMID:23425604

  12. Discovery of genetic biomarkers contributing to variation in drug response of cytidine analogues using human lymphoblastoid cell lines

    PubMed Central

    2014-01-01

    Background Two cytidine analogues, gemcitabine and cytosine arabinoside (AraC), are widely used in the treatment of a variety of cancers with a large individual variation in response. To identify potential genetic biomarkers associated with response to these two drugs, we used a human lymphoblastoid cell line (LCL) model system with extensive genomic data, including 1.3 million SNPs and 54,000 basal expression probesets to perform genome-wide association studies (GWAS) with gemcitabine and AraC IC50 values. Results We identified 11 and 27 SNP loci significantly associated with gemcitabine and AraC IC50 values, respectively. Eleven candidate genes were functionally validated using siRNA knockdown approach in multiple cancer cell lines. We also characterized the potential mechanisms of genes by determining their influence on the activity of 10 cancer-related signaling pathways using reporter gene assays. Most SNPs regulated gene expression in a trans manner, except 7 SNPs in the PIGB gene that were significantly associated with both the expression of PIGB and gemcitabine cytotoxicity. Conclusion These results suggest that genetic variation might contribute to drug response via either cis- or trans- regulation of gene expression. GWAS analysis followed by functional pharmacogenomics studies might help identify novel biomarkers contributing to variation in response to these two drugs and enhance our understanding of underlying mechanisms of drug action. PMID:24483146

  13. Noninvolvement of the X chromosome in radiation-induced chromosome translocations in the human lymphoblastoid cell line TK6

    SciTech Connect

    Jordan, R.; Schwartz, J.L. )

    1994-03-01

    Fluorescence in situ hybridization procedures were used to examine the influence of chromosome locus on the frequency and type of chromosome aberrations induced by [sup 60]Co [gamma] rays in the human lymphoblastoid cell line TK6. Aberrations involving the X chromosome were compared to those involving the similarly sized autosome chromosome 7. When corrected for DNA content, acentric fragments were induced with equal frequency in the X and 7 chromosomes. Dose-dependent increases in chromosomal interchanges involving chromosome 7 were noted, and the frequencies of balanced translocations and dicentrics produced were approximately equal. Chromosome interchanges involving the X chromosome were rare and showed no apparent dose dependence. Thus, while chromosomes 7 and X are equally sensitive to the induction of chromosome breaks, the X chromosome is much less likely to interact with autosomes than chromosome 7. The noninvolvement of the X chromosome in translocations with autosomes may reflect a more peripheral and separate location for the X chromosome in the mammalian nucleus. 20 refs., 2 figs., 1 tab.

  14. Chromosome loss caused by DNA fragmentation induced in main nuclei and micronuclei of human lymphoblastoid cells treated with colcemid.

    PubMed

    Yamamoto, Mika; Wakata, Akihiro; Aoki, Yoshinobu; Miyamae, Yoichi; Kodama, Seiji

    2014-04-01

    Aneuploidy, a change in the number of chromosomes, plays an essential role in tumorigenesis. Our previous study demonstrated that a loss of a whole chromosome is induced in human lymphocytes by colcemid, a well-known aneugen. Here, to clarify the mechanism for colcemid-induced chromosome loss, we investigated the relationship between chromosome loss and DNA fragmentation in human lymphoblastoid cells treated with colcemid (an aneugen) compared with methyl methanesulfonate (MMS; a clastogen). We analyzed the number of fluorescence in situ hybridization (FISH) signals targeted for a whole chromosome 2 in cytokinesis-blocked binucleated TK6 cells and WTK-1 cells treated with colcemid and MMS, and concurrently detected DNA fragmentation by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Results revealed that DNA fragmentation occurred in 60% of all binucleated TK6 cells harboring colcemid-induced chromosome loss (30% of micronuclei and 30% of main nuclei). DNA fragmentation was observed in colcemid-induced micronuclei containing a whole chromosome but not in MMS-induced micronuclei containing chromosome fragments. In contrast, colcemid-induced nondisjunction had no effect on induction of DNA fragmentation, suggesting that DNA fragmentation was triggered by micronuclei containing a whole chromosome but not by micronuclei containing chromosome fragments or nondisjunction. In addition, the frequency of binucleated cells harboring chromosome loss with DNA fragmentation in micronuclei or main nuclei was higher in wild-type p53 TK6 cells than in mutated-p53 WTK-1 cells treated with colcemid. Taken together, these present and previous results suggest that colcemid-induced chromosome loss is caused by DNA fragmentation, which is triggered by a micronucleus with a whole chromosome and controlled by the p53-dependent pathway. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Oxidative metabolism of flunarizine and cinnarizine by microsomes from B-lymphoblastoid cell lines expressing human cytochrome P450 enzymes.

    PubMed

    Kariya, S; Isozaki, S; Uchino, K; Suzuki, T; Narimatsu, S

    1996-11-01

    The oxidative metabolism of cinnarizine [(E)-1-(diphenylmethyl)-4-(3-phenyl-2-propyl)piperazine, CZ] and flunarizine [(E)-1-[bis(4-fluorophenyl)methyl]-4-(3-phenyl-2-propyl)piperazine, FZ] was examined in microsomes from lymphoblastoid cells that expressed human cytochrome P450 (CYP) enzymes. Among 10 kinds of CYP enzymes examined, only CYP2D6 catalyzed p-hydroxylation of the cinnamyl phenyl ring of CZ (C-2 formation) and FZ (F-2 formation), and only CYP2B6 exhibited activity for p-hydroxylation (C-4 formation) of the diphenylmethyl group of CZ at a substrate concentration of 50 microM. On the other hand, CYP2C9 together with CYP1A1, -1A2 and/or -2A6 mediated N-desalkylation at the 1- and 4-positions of the piperazine ring of the two drugs that formed C-1 and C-3 from CZ and F-1 and F-3 from FZ, respectively, whereas CYP2C8, -2C19, -2E1 or -3A4 did not show detectable activity for these reactions under the conditions used. We then examined kinetics for the oxidative metabolism of CZ and FZ using CYP2B6 and -2D6 that have considerable activities. CYP2D6 with Km values of 2 to 4 microM had intrinsic clearance values (Vmax/Km) of 0.31 and 0.14 ml/min/nmol CYP for C-2 and F-2 formation, respectively, while CYP2B6 with a Km value of 17 microM exhibited the clearance value of 0.10 ml/min/nmol CYP for C-4 formation. These results suggest that CYP2D6 mainly mediates p-hydroxylation of the cinnamyl phenyl rings of CZ and FZ, and CYP2B6 mediates that of the diphenylmethyl group of CZ.

  16. The patterns of histone modifications in the vicinity of transcription factor binding sites in human lymphoblastoid cell lines.

    PubMed

    Nie, Yumin; Liu, Hongde; Sun, Xiao

    2013-01-01

    Transcription factor (TF) binding at specific DNA sequences is the fundamental step in transcriptional regulation and is highly dependent on the chromatin structure context, which may be affected by specific histone modifications and variants, known as histone marks. The lack of a global binding map for hundreds of TFs means that previous studies have focused mainly on histone marks at binding sites for several specific TFs. We therefore studied 11 histone marks around computationally-inferred and experimentally-determined TF binding sites (TFBSs), based on 164 and 34 TFs, respectively, in human lymphoblastoid cell lines. For H2A.Z, methylation of H3K4, and acetylation of H3K27 and H3K9, the mark patterns exhibited bimodal distributions and strong pairwise correlations in the 600-bp region around enriched TFBSs, suggesting that these marks mainly coexist within the two nucleosomes proximal to the TF sites. TFs competing with nucleosomes to access DNA at most binding sites, contributes to the bimodal distribution, which is a common feature of histone marks for TF binding. Mark H3K79me2 showed a unimodal distribution on one side of TFBSs and the signals extended up to 4000 bp, indicating a longer-distance pattern. Interestingly, H4K20me1, H3K27me3, H3K36me3 and H3K9me3, which were more diffuse and less enriched surrounding TFBSs, showed unimodal distributions around the enriched TFBSs, suggesting that some TFs may bind to nucleosomal DNA. Besides, asymmetrical distributions of H3K36me3 and H3K9me3 indicated that repressors might establish a repressive chromatin structure in one direction to repress gene expression. In conclusion, this study demonstrated the ranges of histone marks associated with TF binding, and the common features of these marks around the binding sites. These findings have epigenetic implications for future analysis of regulatory elements.

  17. Genetic instability on chromosome 16 in a Human B lymphoblastoid cell line

    SciTech Connect

    Smith, L.E.; Grosovsky, A.J. )

    1993-11-01

    Mutagenesis at the aprt locus in TK6 human lymphoblasts has been found to occur at an unusually high rate (1.2 [times] 10[sup [minus]9]) for a homozygous diploid locus. Evaluation of linked microsatellite polymorphisms demonstrated that loss of heterozygosity (LOH) accompanies conventional intragenic sequence alterations in each APRT[sup [minus

  18. RECEPTOR FOR SOLUBLE C3 AND C3b ON HUMAN LYMPHOBLASTOID (RAJI) CELLS

    PubMed Central

    Theofilopoulos, Argyrios N.; Bokisch, Viktor A.; Dixon, Frank J.

    1974-01-01

    This study describes the presence of a receptor for fluid phase human C3 and C3b on Raji cell membranes. The binding of C3 and C3b was demonstrated indirectly by a fluoresceinated anti-C3 serum and directly by using radioiodinated proteins. No other complement proteins or serum factors were needed to mediate binding of C3 and C3b to the receptor. The possibility of enzymatic cleavage of C3 before or after its attachment on the cell membrane was ruled out by the demonstration of antigenically intact C3 on Raji cells. Inhibition and dissociation of Raji cell-EAC1423 rosettes by C3 and C3b indicated that both of these proteins bind to the same receptor site or closely associated receptor sites on Raji cells. C3b-bearing Raji cells were immune adherence negative, indicating that C3b binding to the receptor is brought about through the immune adherence region of the molecule and not the C3d portion. The C3 receptor on Raji cell membranes is uniformly distributed and can move on the membrane plane. Approximately 4 x 105 molecules of C3 or C3b bind per Raji cell. The receptor had a higher affinity for C3 than C3b, as was shown by uptake experiments and inhibition of Raji cell-EAC1423 rosette formation. Apart from the described receptor for C3 and C3b another specific receptor for C3b inactivator-cleaved C3b (C3d) bound to red cells was shown to be present on Raji cells. Raji cells cultured in medium containing fresh normal human serum and cobra venom factor were lysed. Similar results were obtained when C3b-bearing Raji cells were cultured in medium with fresh normal human serum. The lytic effect could be abolished by inactivating serum C3 proactivator (C3PA) and required C6. It was concluded that C3b bound to the Raji cell membrane activates the complement system through the alternate pathway and results in membrane damage and cytolysis. It is postulated that cell destruction by this mechanism may play an important role in vivo in controlling cell growth. PMID:4591176

  19. Mutational spectrum of ICR-191 at the hprt locus in human lymphoblastoid cells.

    PubMed

    Taft, S A; Liber, H L; Skopek, T R

    1994-01-01

    Human TK6 lymphoblasts were treated with the acridine derivative ICR-191, and mutants at the hprt locus were isolated. Mutant hprt cDNA was reverse-transcribed from mRNA, amplified by polymerase chain reaction (PCR), and sequenced. Additions of single G:C base pairs (+1 frameshift mutations) in repetitive G:C sequences were found in 82% (32/39) of the mutants. Sixteen of the +1 frameshifts analyzed were located in a single sequence of six consecutive guanine bases in exon 3. The remaining +1 frameshifts occurred at six different GGG sequences (14 mutants) and a single GGGG sequence (2 mutants) in other hprt exons. The repetitive guanine sequences that underwent frameshift mutagenesis were located in both the transcribed and nontranscribed strands of hprt. No single base deletions (-1 frameshift mutations) were observed. Base substitutions were observed in 13% (5/39) of the clones analyzed and occurred at both G:C and A:T bases. Loss of exon 4 from the cDNA was also observed in 5% (2/39) of the mutants. Hprt mutants containing seven consecutive guanines (produced from a +1 frameshift in a GGGGGG sequence) were treated with ICR-191 and wild-type revertants selected in CHAT medium. Revertants were recovered at a frequency of approximately 10(-7) and contained the wild-type sequence (GGGGGG) in all clones analyzed. The observed frequency of ICR-191-induced-1 frameshift reversion in the GGGGGGG sequence was approximately 500-fold lower than the estimated frequency of +1 frameshifts observed in the wild-type GGGGGG sequence following the same ICR-191 treatment. These results suggest that ICR-191 produces predominantly +1 frameshift mutations at the hprt locus in human cells.

  20. In vitro adverse effects of iron ore dusts on human lymphoblastoid cells in culture.

    PubMed

    Wang, He; Wang, Jing J; Sanderson, Barbara J S

    2013-01-01

    The aim of this study was to investigate the adverse effects produced by four types of iron (Fe) ore dust using cultured human cells. Genotoxicity and cytotoxicity induced by Fe ore dusts were determined by assays including cytokinesis block micronucleus (CBMN), population growth, and methyl tetrazolium (MTT). Four iron ore dusts were tested, namely, 1002 Limonite & Goethite (1002), HG2 hematite (HG2), HG1 Soutlem Pit (HG1), and HG4. WIL2 -NS cells were incubated for 10 h with extracts from a range of concentrations (0, 75, or 150 μg/ml) of Fe ore dust. Significant decreases in percent cell viability were seen at 150 μg/ml HG2 and 1002 as measured by MTT, with viability that decreased to 75 and 73%, respectively, compared to untreated controls. The cell population regrew to a different extent after Fe ore dust was removed, except for HG1, where population remained declined. An approximately twofold significant increase in the frequency of micronucleated binucleated cells (MNBNC) was seen with 1002, HG2, and HG1 at 150 μg/ml. A significant rise in apoptosis induction was observed at 150 μg/ml HG1. Data indicate that Fe ore dusts at 150 μg/ml produced cytotoxicity and genotoxicity.

  1. Modulation of a human lymphoblastoid B cell line by cyclic AMP. Ig secretion and phosphatidylcholine metabolism

    SciTech Connect

    Shearer, W.T.; Patke, C.L.; Gilliam, E.B.; Rosenblatt, H.M.; Barron, K.S.; Orson, F.M.

    1988-09-01

    A transformed human B cell line, LA350, was found to be sensitive to cAMP-elevating agents by responding with rapid (0 to 2 h) severalfold elevations of intracellular cAMP to treatment with cholera toxin, isobutylmethylxanthine (IBMX), forskolin, and dibutyryl cAMP (all p less than 0.001). These cAMP-elevating agents also produced significant inhibitions of subsequent (48 to 72 h) Ig secretion by the same B cells as measured by a reverse hemolytic plaque assay and an enzyme-linked immunoadsorbent assay for IgM (both p less than 0.001). PMA- and IBMX-treated cells were particularly responsive to the effects of cholera toxin, showing a doubling of cAMP content and profound decrease in Ig production (p less than 0.001). Because our previous studies had correlated activation of the metabolic turnover of the phosphatidylcholine (PC) fraction of membrane phospholipids with enhanced Ig secretion, we examined the sensitivity of PC metabolism to cAMP in control and PMA-stimulated cells. Formation of PC was found to be inhibited by forskolin and IBMX (both p less than 0.002) but breakdown of PC was stimulated (p less than 0.001). These findings imply that as the enzymatic products of PC, choline phosphate and diacylglycerol, are depleted due to the combined effects of cAMP upon synthesis and turnover of PC, there is a decrease in Ig secretion. Since diacylglycerol activates protein kinase C, it appears reasonable that Ig secretion is at least partially regulated by cAMP-responsive alterations in PC metabolism produced by protein kinase C-induced phosphorylation. We conclude that the early cAMP-sensitive changes in PC metabolism in this activated B cell line may signal for subsequent alterations in Ig secretion.

  2. The effect of zinc sulphate and zinc carnosine on genome stability and cytotoxicity in the WIL2-NS human lymphoblastoid cell line.

    PubMed

    Sharif, Razinah; Thomas, Philip; Zalewski, Peter; Graham, Robin D; Fenech, Michael

    2011-02-28

    Zinc (Zn) is an essential cofactor required by numerous enzymes that are essential for cell metabolism and the maintenance of DNA integrity. We investigated the effect of Zn deficiency or excess on genomic instability events and determined the optimal concentration of two Zn compounds that minimize DNA-damage events. The effects of Zn sulphate (ZnSO(4)) and Zn carnosine (ZnC) on cell proliferation were investigated in the WIL2-NS human lymphoblastoid cell line. DNA damage was determined by the use of both the comet assay and the cytokinesis-block micronucleus cytome (CBMN-Cyt) assay. Zn-deficient medium (0μM) was produced using Chelex treatment, and the two Zn compounds (i.e. ZnSO(4) and ZnC) were tested at concentrations of 0.0, 0.4, 4.0, 16.0, 32.0 and 100.0μM. Results from an MTT assay showed that cell growth and viability were decreased in Zn-depleted cells (0μM) as well as at 32μM and 100μM for both Zn compounds (P<0.0001). DNA strand-breaks, as measured by the comet assay, were found to be increased in Zn-depleted cells compared with the other treatment groups (P<0.05). The CBMN-Cyt assay showed a significant increase in the frequency of both apoptotic and necrotic cells under Zn-deficient conditions (P<0.0001). Elevated frequencies of micronuclei (MNi), nucleoplasmic bridges (NPBs) and nuclear buds (NBuds) were induced in Zn-depleted cells (P<0.0001), whereas genome damage was reduced in supplemented cultures for both Zn compounds at 4μM and 16μM, possibly suggesting that these concentrations may be optimal for genome stability. The potential protective effect of ZnSO(4) and ZnC was also investigated following exposure to 1.0Gy γ-radiation. Culture in medium containing these compounds at 4-32μM prior to irradiation displayed significantly reduced frequencies of MNi, NPBs and NBuds compared with cells maintained in 0μM medium (P<0.0001). Expression of γ-H2AX and 8-oxoguanine glycosylase measured by western blotting was increased in Zn

  3. Molecular characterization of the GM 4672 human lymphoblastoid cell line and analysis of its use as a fusion partner in the generation of human-human hybridoma autoantibodies.

    PubMed

    Rioux, J D; Rauch, J; Zdarsky, E; Newkirk, M M

    1993-07-01

    The GM 4672 lymphoblastoid cell line has been used in cell hybridization experiments with peripheral blood lymphocytes (PBLs) in order to generate human-human hybridomas that secrete immunoglobulins directed against a number of different autoantigens. The GM 4672 cells were fused with PBLs isolated from patients with rheumatoid arthritis or systemic lupus erythematosus, or from normal individuals, and the resulting hybridomas were screened for reactivity to platelets, erythrocytes, DNA, cardiolipin, human IgG-Fc, phosphatidylethanolamine, and for lupus anticoagulant activity. This report analyzes the results from 149 fusion experiments completed over a period of nine years. Fifty to sixty-six percent of the fusion experiments resulted in immunoglobulin-secreting clones, with an average of 15 clones/fusion. The hybridoma antibodies were predominantly of the IgM heavy chain isotype, and 67% expressed kappa light chains. Although most hybridoma antibodies (78%) recognized a single autoantigen, 22% recognized more than one autoantigen and were considered polyreactive. In addition, the light and heavy chain variable regions of the antibody secreted by the GM 4672 cell line were amplified by the polymerase chain reaction technique and sequenced. The GM 4672 light chain was encoded by a VkI gene and used a Jk4 minigene. The GM 4672 heavy chain was derived for the rearrangement of a gene from the VH4 subgroup and used a JH4 minigene. The 8 amino acid long diversity region was generated by the fusion of the DK1 and DLR2 genes. The hybridomas generated in fusion experiments, when examined, did not appear to secrete antibodies using the immunoglobulin variable regions derived from the GM 4672 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

  4. Activation of 3-nitrobenzanthrone and its metabolites to DNA-damaging species in human B lymphoblastoid MCL-5 cells.

    PubMed

    Arlt, Volker M; Cole, Kathleen J; Phillips, David H

    2004-03-01

    3-Nitrobenzanthrone (3-NBA) is one of the most potent mutagens in the Ames Salmonella typhimurium assay and a suspected human carcinogen recently identified in diesel exhaust and in airborne particulate matter. 3-Aminobenzanthrone (3-ABA), 3-acetylaminobenzanthrone (3-Ac-ABA) and N-acetyl-N-hydroxy-3-aminobenzanthrone (N-Ac-N-OH-ABA) have been identified as 3-NBA metabolites. In the present study we investigated the genotoxic effects of 3-NBA and its metabolites in the human B lymphoblastoid cell line MCL-5. DNA strand breaks were measured using the Comet assay, chromosomal damage was assessed using the micronucleus assay and DNA adduct formation was determined by 32P-post-labelling analysis. DNA strand-breaking activity was observed with each compound in a concentration-dependent manner (1-50 microM, 2 h incubation time). At 50 microM median comet tail lengths (CTLs) were 25.0 microm for 3-NBA, 48.0 microm for 3-ABA, 54.5 microm for 3-Ac-ABA and 65.0 microm for N-Ac-N-OH-ABA. Median CTLs in control incubations were in the range 7.7-13.1 micro m. Moreover, the strand-breaking activity of 3-NBA was more pronounced in the presence of a DNA repair inhibitor, hydroxyurea. Depending on the concentration used (1-20 microM, 24 h incubation time), 3-NBA and its metabolites also showed clastogenic activity in the micronucleus assay. 3-NBA and N-Ac-N-OH-ABA were the most active at low concentrations; at 1 microM the total number of micronuclei per 500 binucleate cells was 4.7 +/- 0.6 in both cases, compared with 1.7-3.0 for controls (P < 0.05). Furthermore, multiple DNA adducts were detected with each compound (1 microM, 24 h incubation time), essentially similar to those found recently in vivo in rats treated with 3-NBA or its metabolites. DNA adduct levels ranged from 1.3 to 42.8 and from 2.0 to 39.8 adducts/10(8) nt using the nuclease P1 and butanol enrichment procedures, respectively. DNA binding was highest for N-Ac-N-OH-ABA, followed by 3-NBA, and much lower for 3-ABA

  5. High-potentiality preliminary selection criteria and transformation time-dependent factors analysis for establishing Epstein-Barr virus transformed human lymphoblastoid cell lines.

    PubMed

    Chang, I-C; Wu, J-Y; Lu, H-I; Ko, H-W; Kuo, J-L; Wang, C-Y; Shen, P-S; Hwang, S-M

    2006-12-01

    Infection of freshly isolated and cryopreserved lymphocytes with Epstein-Barr virus (EBV) leads to the establishment of human B lymphoblastoid cell lines (LCLs). Techniques for optimal infection of the lymphocytes are vital for the establishment of a human biobank. The present study found that more than half (58-86%) of such established LCLs had transport times of less than 48 h, cell densities exceeding 10(6) cells/ml and cell viabilities greater than 90%. After EBV infection, 3306 freshly isolated lymphocytes required 30.0 +/- 0.1 days to become LCLs. Conversely, 1210 cryopreserved lymphocytes required 36.2 +/- 0.4 days. Cell density and viability of the culture affected transformation time in freshly isolated lymphocytes. On the other hand, blood transport time, cryopreservation time and initial cell viability were major factors in cryopreserved specimens. These results contribute to general information concerning the establishment of a human biobank for EBV infected cells.

  6. Mutagenic adaptive response to high-LET radiation in human lymphoblastoid cells exposed to X-rays.

    PubMed

    Varès, Guillaume; Wang, Bing; Tanaka, Kaoru; Kakimoto, Ayana; Eguchi-Kasai, Kyomi; Nenoi, Mitsuru

    2011-01-10

    The ability of cells to adapt low-dose or low-dose rate radiation is well known. High-LET radiation has unique characteristics, and the data concerning low doses effects and high-LET radiation remain fragmented. In this study, we assessed in vitro the ability of low doses of X-rays to induce an adaptive response (AR) to a subsequent challenging dose of heavy-ion radiation. Lymphoblastoid cells (TK6, AHH-1, NH32) were exposed to priming 0.02-0.1Gy X-rays, followed 6h later by challenging 1Gy heavy-ion radiation (carbon-ion: 20 and 40keV/μm, neon-ion: 150keV/μm). Pre-exposure of p53-competent cells resulted in decreased mutation frequencies at hypoxanthine-guanine phosphoribosyl transferase locus and different H2AX phosphorylation kinetics, as compared to cells exposed to challenging radiation alone. This phenomenon did not seem to be linked with cell cycle effects or radiation-induced apoptosis. Taken together, our results suggested the existence of an AR to mutagenic effects of heavy-ion radiation in lymphoblastoid cells and the involvement of double-strand break repair mechanisms.

  7. Comparative study of the metabolism of drug substrates by human cytochrome P450 3A4 expressed in bacterial, yeast and human lymphoblastoid cells.

    PubMed

    Andrews, J; Abd-Ellah, M F; Randolph, N L; Kenworthy, K E; Carlile, D J; Friedberg, T; Houston, J B

    2002-11-01

    1. The aim was to compare the metabolic activity of human CYP3A4 expressed in bacteria (E. coli), yeast (S. cerevisiae) and human lymphoblastoid cells (hBl), with the native CYP3A4 activity observed in a panel of human livers. 2. Three CYP3A4 substrates were selected for study: dextromethorphan (DEM), midazolam (MDZ) and diazepam (DZ). The substrate metabolism in each of the four systems was characterized by deriving the kinetic parameters K(m) or S(50), V(max) and intrinsic clearance (CL(int)) or maximum clearance (CL(max)) from the kinetic profiles; the latter differing by 100-fold across the three substrates. 3. The K(m) or S(50) for the formation of metabolites 3-methoxymorphinan (MEM), 1'-hydroxymidazolam (1'-OH MDZ) and 3-hydroxydiazepam (3HDZ) compared well in all systems. For CYP3A4-mediated metabolism of DEM, MDZ and DZ, the V(max) for hBl microsomes were generally 2-9-fold higher than the respective yeast and human liver microsomes and E. coli membrane preparations, resulting in greater CL(int) or CL(max). In the case of 3HDZ formation, non-linear kinetics were observed for E. coli, hBl microsomes and human liver microsomes, whereas the kinetics observed for S. cerevisiae were linear. 4. The use of native human liver microsomes for drug metabolic studies will always be preferable. However, owing to the limited availability of human tissues, we find it is reasonable to use any of the recombinant systems described herein, since all three recombinant systems gave good predictions of the native human liver enzyme activities.

  8. Studying the protein expression in human B lymphoblastoid cells exposed to 1.8-GHz (GSM) radiofrequency radiation (RFR) with protein microarray.

    PubMed

    Zhijian, Chen; Xiaoxue, Li; Wei, Zheng; Yezhen, Lu; Jianlin, Lou; Deqiang, Lu; Shijie, Chen; Lifen, Jin; Jiliang, He

    2013-03-29

    In the present study, the protein microarray was used to investigate the protein expression in human B-cell lymphoblastoid cells intermittently exposed to 1.8-GHz GSM radiofrequency radiation (RFR) at the specific absorption rate (SAR) of 2.0 W/kg for 24 h. The differential expression of 27 proteins was found, which were related to DNA damage repair, apoptosis, oncogenesis, cell cycle and proliferation (ratio >1.5-fold, P<0.05). The results validated with Western blot assay indicated that the expression of RPA32 was significantly down-regulated (P<0.05) while the expression of p73 was significantly up-regulated in RFR exposure group (P<0.05). Because of the crucial roles of those proteins in DNA repair and cell apoptosis, the results of present investigation may explain the biological effects of RFR on DNA damage/repair and cell apoptosis. Copyright © 2013 Elsevier Inc. All rights reserved.

  9. Endogenous antigen presentation by autoantigen-transfected Epstein-Barr virus-lymphoblastoid cells. I. Generation of human thyroid peroxidase-reactive T cells and their T cell receptor repertoire.

    PubMed Central

    Martin, A; Magnusson, R P; Kendler, D L; Concepcion, E; Ben-Nun, A; Davies, T F

    1993-01-01

    To develop a model for endogenous thyroid autoantigen presentation, we transfected EBV-transformed B lymphoblastoid cell lines (EBV-LCL), established from patients with autoimmune thyroid disease and normal controls, with cDNA for the human thyroid autoantigen thyroid peroxidase (hTPO). hTPO-antigen presentation to patient peripheral blood T cells was demonstrated after stimulation in vitro for 7 d with irradiated hTPO-transfected or untransfected autologous EBV-LCL. Anti-hTPO-reactive T cells were subsequently cloned in the presence of irradiated, autologous hTPO-transfected EBV-LCL and IL-2.10 T cell-cloned lines exhibited specific hTPO-induced proliferation (stimulation indices of 2.1-7.9) towards autologous hTPO-transfected EBV-LCL, and were subjected to human T cell receptor (hTCR) V gene analysis, using the PCR for the detection of V alpha and V beta hTcR gene families. The results indicated a preferential use of hTCR V alpha 1 and/or V alpha 3 in 9 of the 10 lines. In contrast, hTCR V beta gene family use was more variable. These data demonstrate a model for the endogenous presentation of human thyroid peroxidase in the absence of other thyroid specific antigens. The high frequency of antigen-specific T cells obtained from PBMC using this technique will facilitate further studies at both the functional and hTCR V gene level. Images PMID:7682574

  10. The human T-cell leukemia virus type 1 Rex regulatory protein exhibits an impaired functionality in human lymphoblastoid Jurkat T cells.

    PubMed Central

    Hamaia, S; Cassé, H; Gazzolo, L; Duc Dodon, M

    1997-01-01

    The Rex protein of human T-cell leukemia virus type 1 (HTLV-1) intervenes in the posttranscriptional regulation of proviral gene expression. Its binding to the Rex response element (XRE) present in the 3' long terminal repeat ensures the coordinate cytoplasmic accumulation of spliced and unspliced forms of viral messengers. Consequently, synthesis of viral structural and enzymatic proteins is strictly dependent on the Rex posttranscriptional activity. Here we report that synthesis of HTLV-1 envelope glycoproteins by Jurkat T cells could be detected only when they were regulated in a Rex-independent manner. Indeed, Jurkat T cells transfected with a Rex-dependent env expression vector (encompassing both the env and pX open reading frames) do not produce significant levels of envelope glycoproteins despite the production of significant amounts of Rex protein. The analysis of levels and distribution patterns of the unspliced env and of the singly spliced tax/rex transcripts suggests that the failure in envelope glycoprotein synthesis may be ascribed to a deficiency of Rex in mediating the nucleocytoplasmic transport of unspliced env RNAs in these cells. Furthermore, despite the synthesis of regulatory proteins, HTLV-1 structural proteins were not detected in Jurkat T cells transfected with an HTLV-1 infectious provirus. Conversely, and as expected, structural proteins were produced by Jurkat cells transfected by a human immunodeficiency virus type 1 (HIV-1) infectious provirus. This phenotype appeared to be linked to a specific dysfunction of Rex, since the functionally equivalent Rev protein of HIV-1 was shown to be fully efficient in promoting the synthesis of HTLV-1 envelope glycoproteins in Jurkat cells. Therefore, it seems likely that the block to Rex function in these lymphoblastoid T cells is determined by inefficient Rex-XRE interactions. These observations suggest that the acquisition of this Rex-deficient phenotype by in vivo-infected HTLV-1 T cells may

  11. Comparative investigations of sodium arsenite, arsenic trioxide and cadmium sulphate in combination with gamma-radiation on apoptosis, micronuclei induction and DNA damage in a human lymphoblastoid cell line.

    PubMed

    Hornhardt, Sabine; Gomolka, Maria; Walsh, Linda; Jung, Thomas

    2006-08-30

    In the field of radiation protection the combined exposure to radiation and other toxic agents is recognised as an important research area. To elucidate the basic mechanisms of simultaneous exposure, the interaction of the carcinogens and environmental toxicants cadmium and two arsenic compounds, arsenite and arsenic trioxide, in combination with gamma-radiation in human lymphoblastoid cells (TK6) were investigated. Gamma-radiation induced significant genotoxic effects such as micronuclei formation, DNA damage and apoptosis, whereas arsenic and cadmium had no significant effect on these indicators of cellular damage at non-toxic concentrations. However, in combination with gamma-radiation arsenic trioxide induced a more than additive apoptotic rate compared to the sum of the single effects. Here, the level of apoptotic cells was increased, in a dose-dependent way, up to two-fold compared to the irradiated control cells. Arsenite did not induce a significant additive effect at any of the concentrations or radiation doses tested. On the other hand, arsenic trioxide was less effective than arsenite in the induction of DNA protein cross-links. These data indicate that the two arsenic compounds interact through different pathways in the cell. Cadmium sulphate, like arsenite, had no significant effect on apoptosis in combination with gamma-radiation at low concentrations and, at high concentrations, even reduced the radiation-induced apoptosis. An additive effect on micronuclei induction was observed with 1muM cadmium sulphate with an increase of up to 80% compared to the irradiated control cells. Toxic concentrations of cadmium and arsenic trioxide seemed to reduce micronuclei induction. The results presented here indicate that relatively low concentrations of arsenic and cadmium, close to those occuring in nature, may interfere with radiation effects. Differences in action of the two arsenic compounds were identified.

  12. Different DNA damage response of cis and trans isomers of commonly used UV filter after the exposure on adult human liver stem cells and human lymphoblastoid cells.

    PubMed

    Sharma, Anežka; Bányiová, Katarína; Babica, Pavel; El Yamani, Naouale; Collins, Andrew Richard; Čupr, Pavel

    2017-09-01

    2-ethylhexyl 4-methoxycinnamate (EHMC), used in many categories of personal care products (PCPs), is one of the most discussed ultraviolet filters because of its endocrine-disrupting effects. EHMC is unstable in sunlight and can be transformed from trans-EHMC to emergent cis-EHMC. Toxicological studies are focusing only on trans-EHMC; thus the toxicological data for cis-EHMC are missing. In this study, the in vitro genotoxic effects of trans- and cis-EHMC on adult human liver stem cells HL1-hT1 and human-derived lymphoblastoid cells TK-6 using a high-throughput comet assay were studied. TK-6 cells treated with cis-EHMC showed a high level of DNA damage when compared to untreated cells in concentrations 1.56 to 25μgmL(-1). trans-EHMC showed genotoxicity after exposure to the two highest concentrations 12.5 and 25μgmL(-1). The increase in DNA damage on HL1-hT1 cells induced by cis-EHMC and trans-EHMC was detected at the concentration 25μgmL(-1). The No observed adverse effect level (NOAEL, mg kg(-1)bwday(-1)) was determined using a Quantitative in vitro to in vivo extrapolation (QIVIVE) approach: NOAELtrans-EHMC=3.07, NOAELcis-EHMC=0.30 for TK-6 and NOAELtrans-EHMC=26.46, NOAELcis-EHMC=20.36 for HL1-hT1. The hazard index (HI) was evaluated by comparing the reference dose (RfD, mgkg(-1)bwday(-1)) obtained from our experimental data with the chronic daily intake (CDI) of the female population. Using comet assay experimental data with the more sensitive TK-6 cells, HIcis-EHMC was 7 times higher than HItrans-EHMC. In terms of CDI, relative contributions were; dermal exposure route>oral>inhalation. According to our results we recommend the RfDtrans-EHMC=0.20 and RfDcis-EHMC=0.02 for trans-EHMC and cis-EHMC, respectively, to use for human health risk assessment. The significant difference in trans-EHMC and cis-EHMC response points to the need for toxicological reevaluation and application reassessment of both isomers in PCPs. Copyright © 2017 Elsevier B.V. All rights

  13. Fast neutrons-induced apoptosis is Fas-independent in lymphoblastoid cells

    SciTech Connect

    Fischer, Barbara; Benzina, Sami; Jeannequin, Pierre; Dufour, Patrick; Bergerat, Jean-Pierre; Denis, Jean-Marc; Gueulette, John; Bischoff, Pierre L. . E-mail: Pierre.Bischoff@ircad.u-strasbg.fr

    2005-08-26

    We have previously shown that ionizing radiation-induced apoptosis in human lymphoblastoid cells differs according to their p53 status, and that caspase 8-mediated cleavage of BID is involved in the p53-dependent pathway. In the present study, we investigated the role of Fas signaling in caspase 8 activation induced by fast neutrons irradiation in these cells. Fas and FasL expression was assessed by flow cytometry and by immunoblot. We also measured Fas aggregation after irradiation by fluorescence microscopy. We found a decrease of Fas expression after irradiation, but no change in Fas ligand expression. We also showed that, in contrast to the stimulation of Fas by an agonistic antibody, Fas aggregation did not occur after irradiation. Altogether, our data strongly suggest that fast neutrons induced-apoptosis is Fas-independent, even in p53-dependent apoptosis.

  14. Studying the protein expression in human B lymphoblastoid cells exposed to 1.8-GHz (GSM) radiofrequency radiation (RFR) with protein microarray

    SciTech Connect

    Zhijian, Chen; Xiaoxue, Li; Wei, Zheng; Yezhen, Lu; Jianlin, Lou; Deqiang, Lu; Shijie, Chen; Lifen, Jin; Jiliang, He

    2013-03-29

    Highlights: ► Protein microarray shows the differential expression of 27 proteins induced by RFR. ► RPA32 related to DNA repair is down-regulated in Western blot. ► p73 related to cell genome stability and apoptosis is up-regulated in Western blot. -- Abstract: In the present study, the protein microarray was used to investigate the protein expression in human B-cell lymphoblastoid cells intermittently exposed to 1.8-GHz GSM radiofrequency radiation (RFR) at the specific absorption rate (SAR) of 2.0 W/kg for 24 h. The differential expression of 27 proteins was found, which were related to DNA damage repair, apoptosis, oncogenesis, cell cycle and proliferation (ratio >1.5-fold, P < 0.05). The results validated with Western blot assay indicated that the expression of RPA32 was significantly down-regulated (P < 0.05) while the expression of p73 was significantly up-regulated in RFR exposure group (P < 0.05). Because of the crucial roles of those proteins in DNA repair and cell apoptosis, the results of present investigation may explain the biological effects of RFR on DNA damage/repair and cell apoptosis.

  15. Biased dA/dT somatic hypermutation as regulated by the heavy chain intronic iEmu enhancer and 3'Ealpha enhancers in human lymphoblastoid B cells.

    PubMed

    Komori, Atsumasa; Xu, Zhenming; Wu, Xiaoping; Zan, Hong; Casali, Paolo

    2006-04-01

    Somatic hypermutation (SHM) in immunoglobulin gene (Ig) variable (V) regions is critical for the maturation of the antibody response. It is dependent on the expression of activation-induced cytidine deaminase (AID) and translesion DNA polymerases in germinal center B cells as well as Ig V transcription, as regulated by the Ig heavy chain (H) intronic enhancer (iEmu) and the 3' enhancer (3'Ealpha) region. We analyzed the role of these cis elements in SHM by stably transfecting Ramos human lymphoblastoid B cells with a rearranged human IgH chain VD (diversity) J (joining) DNA construct containing a V(H) promoter at the 5' end and C(H)1 and C(H)2 exons of Cgamma1 at the 3' end. In this construct, mutations preferentially targeted dA/dT basepairs in the RGYW/WRCY hotspot. Most of the dA/dT mutations and accompanying dC/dG mutations were transitions. Deletion of iEmu resulted in decreased SHM which could be partially restored by insertion of the IgH hs1,2 enhancer. Other two 3'Ealpha enhancers, hs3-hs4, did not significantly increase the mutation frequency, but further strengthened the dA/dT bias. The frequency and spectrum of the mutations were independent of the genomic integration of the transgene or V gene transcription level. Thus, we have established a novel in vitro system to analyze SHM and identify the role of multiple cis-regulatory elements in regulating dA/dT biased SHM. This model system will be useful to further address the role of other cis-regulating elements and recruited trans-acting factors in expressing the modalities of SHM.

  16. In vitro evaluation of human hybrid cell lines generated by fusion of B-lymphoblastoid cells and ex vivo tumour cells as candidate vaccines for haematological malignancies.

    PubMed

    Mohamed, Yehia S; Dunnion, Debbie; Teobald, Iryna; Walewska, Renata; Browning, Michael J

    2012-10-12

    Fusions of dendritic cells (DCs) and tumour cells have been shown to induce protective immunity to tumour challenge in animal models, and to represent a promising approach to cancer immunotherapy. The broader clinical application of this approach, however, is potentially constrained by the lack of replicative capacity and limited standardisation of fusion cell preparations. We show here that fusion of ex vivo tumour cells isolated from patients with a range of haematological malignancies with the human B-lymphoblastoid cell line (LCL), HMy2, followed by chemical selection of the hybridomas, generated stable, self-replicating human hybrid cell lines that grew continuously in tissue culture, and survived freeze/thawing cycles. The hybrid cell lines expressed HLA class I and class II molecules, and the major T-cell costimulatory molecules, CD80 and CD86. All but two of 14 hybrid cell lines generated expressed tumour-associated antigens that were not expressed by HMy2 cells, and were therefore derived from the parent tumour cells. The hybrid cell lines stimulated allogeneic T-cell proliferative responses and interferon-gamma release in vitro to a considerably greater degree than their respective parent tumour cells. The enhanced T-cell stimulation was inhibited by CTLA4-Ig fusion protein, and by blocking antibodies to MHC class I and class II molecules. Finally, all of five LCL/tumour hybrid cell lines tested induced tumour antigen-specific cytotoxic T-cell responses in vitro in PBL from healthy, HLA-A2+ individuals, as detected by HLA-A2-peptide pentamer staining and cellular cytotoxicity. These data show that stable hybrid cell lines, with enhanced immunostimulatory properties and potential for therapeutic vaccination, can be generated by in vitro fusion and chemical selection of B-LCL and ex vivo haematological tumour cells. Copyright © 2012 Elsevier Ltd. All rights reserved.

  17. A Study of Alterations in DNA Epigenetic Modifications (5mC and 5hmC) and Gene Expression Influenced by Simulated Microgravity in Human Lymphoblastoid Cells

    PubMed Central

    Wang, Zhiping; Liu, Yunlong; Lossie, Amy C.; Thimmapuram, Jyothi; Irudayaraj, Joseph

    2016-01-01

    Cells alter their gene expression in response to exposure to various environmental changes. Epigenetic mechanisms such as DNA methylation are believed to regulate the alterations in gene expression patterns. In vitro and in vivo studies have documented changes in cellular proliferation, cytoskeletal remodeling, signal transduction, bone mineralization and immune deficiency under the influence of microgravity conditions experienced in space. However microgravity induced changes in the epigenome have not been well characterized. In this study we have used Next-generation Sequencing (NGS) to profile ground-based “simulated” microgravity induced changes on DNA methylation (5-methylcytosine or 5mC), hydroxymethylation (5-hydroxymethylcytosine or 5hmC), and simultaneous gene expression in cultured human lymphoblastoid cells. Our results indicate that simulated microgravity induced alterations in the methylome (~60% of the differentially methylated regions or DMRs are hypomethylated and ~92% of the differentially hydroxymethylated regions or DHMRs are hyperhydroxymethylated). Simulated microgravity also induced differential expression in 370 transcripts that were associated with crucial biological processes such as oxidative stress response, carbohydrate metabolism and regulation of transcription. While we were not able to obtain any global trend correlating the changes of methylation/ hydroxylation with gene expression, we have been able to profile the simulated microgravity induced changes of 5mC over some of the differentially expressed genes that includes five genes undergoing differential methylation over their promoters and twenty five genes undergoing differential methylation over their gene-bodies. To the best of our knowledge, this is the first NGS-based study to profile epigenomic patterns induced by short time exposure of simulated microgravity and we believe that our findings can be a valuable resource for future explorations. PMID:26820575

  18. Mutagenicity and clastogenicity of extracts of Helicobacter pylori detected by the Ames test and in the micronucleus test using human lymphoblastoid cells.

    PubMed

    Arimoto-Kobayashi, Sakae; Ohta, Kaori; Yuhara, Yuta; Ayabe, Yuka; Negishi, Tomoe; Okamoto, Keinosuke; Nakajima, Yoshihiro; Ishikawa, Takeshi; Oguma, Keiji; Otsuka, Takanao

    2015-07-01

    Epidemiological studies have demonstrated a close association between infection with Helicobacter pylori (H.pylori) and the development of gastric carcinoma. Chronic H.pylori infection increases the frequency of mutation in gastric epithelial cells. However, the mechanism by which infection of H.pylori leads to mutation in gastric epithelial cells is unclear. We suspected that components in H.pylori may be related to the mutagenic response associated with DNA alkylation, and could be detected with the Ames test using a more sensitive strain for alkylating agents. Our investigation revealed that an extract of H.pylori was mutagenic in the Ames test with Salmonella typhimurium YG7108, which is deficient in the DNA repair of O(6)-methylguanine. The extract of H.pylori may contain methylating or alkylating agents, which might induce O (6)-alkylguanine in DNA. Mutagenicity of the alkylating agents N-methyl-N-nitrosourea (MNU) and N-methyl-N'-nitro-N-nitrosoguanidine in the Ames test with S.typhimurium TA1535 was enhanced significantly in the presence of the extract of H.pylori. The tested extracts of H.pylori resulted in a significant induction of micronuclei in human-derived lymphoblastoid cells. Heat instability and dialysis resistance of the extracts of H.pylori suggest that the mutagenic component in the extracts of H.pylori is a heat-unstable large molecule or a heat-labile small molecule strongly attached or adsorbed to a large molecule. Proteins in the extracts of H.pylori were subsequently fractionated using ammonium sulphate precipitation. However, all fractions expressed enhancing effects toward MNU mutagenicity. These results suggest the mutagenic component is a small molecule that is absorbed into proteins in the extract of H.pylori, which resist dialysis. Continuous and chronic exposure of gastric epithelial cells to the alkylative mutagenic component from H.pylori chronically infected in the stomach might be a causal factor in the gastric carcinogenesis

  19. Growth of diploid, Epstein-Barr virus-carrying human lymphoblastoid cell lines heterotransplanted into nude mice under immunologically privileged conditions.

    PubMed

    Giovanella, B; Nilsson, K; Zech, L; Yim, O; Klein, G; Stehlin, J S

    1979-07-15

    Human Epstein-Barr virus-carrying lymphoid cell lines which have been classified on the basis of studies on clonality and morphological, chromosomal and functional parameters as lymphoblastoid cell lines (LCL) of presumed non-neoplastic origin were inoculated intracerebrally into nude mice. All eighteen of them grew, killing the host mice within 7 to 25 days, except for 2 which grew more slowly. At autopsy, the brain of the nudes was found to be invaded by infiltrating lymphomas. Sixteen of these lymphomas, when recultured in vitro, gave rise to cell lines with growth properties and morphology indistinguishable from those of the inoculated LCL. Chromosomal examinations showed that 3/7 cell lines injected, which grew as lymphomas in the brain, were still normal diploid on reexplantation whereas the remaining four had become aneuploid. Four lines derived from intracerebral lymphomas (2 diploid, 1 aneuploid and 1 untested) were inoculated subcutaneously into adult nude mice. None of them grew. When the corresponding four original LCL lines were inoculated subcutaneously into newborn nude mice, they grew rapidly, but failed to do so in newborn normal mice or intracerebrally in adult normal mice. One such line, U-1450, was treated with anti-lymphocyte serum (ALS). Small nodules developed at the site of inoculation. From one nodule a cell line was cultured, 1450 ALSAD. It was morphologically indistinguishable from the line of origin. The lines obtained from nude mice inoculated with polyclonal LCL seem to have a restricted clonal representation, but were not monoclonal, as evidenced by analyses of their pattern of immunoglobulin synthesis.

  20. Epitope analysis of peanut allergen Ara h1 with oligoclonal IgM antibody from human B-lymphoblastoid cells.

    USDA-ARS?s Scientific Manuscript database

    To analyze epitopes of peanut allergen Ara h1, Epstein-Barr virus-transformed human peripheral oligoclonal B-cells were cultured to obtain antibodies to Ara h1. The combined reaction pattern with six oligoclonal antibodies showed there were six antibody binding areas named a to f in Ara h1. We found...

  1. Interferon-dependent induction of mRNA for the major histocompatibility antigens in human fibroblasts and lymphoblastoid cells.

    PubMed Central

    Fellous, M; Nir, U; Wallach, D; Merlin, G; Rubinstein, M; Revel, M

    1982-01-01

    In human cells treated with interferons, there is an increase in the amount of HLA-A,B,C and beta 2-microglobulin exposed on the cell surface. We have used a cloned HLA-A,B,C cDNA probe to demonstrate by molecular hybridization that this effect of interferon is preceded by a large increase in the amount of HLA mRNA in the cell. This effect was found in five different human cell lines, with purified leukocyte and fibroblast interferons. The increase in HLA mRNA is comparable in its kinetics and dose-response to the induction of (2'-5') oligo(A) synthetase mRNA by interferons. Therefore, interferons seem to activate at least two cellular genes which have different biochemical functions. Images PMID:6179076

  2. Transfer and expression of three cloned human non-HLA-A,B,C class I major histocompatibility complex genes in mutant lymphoblastoid cells.

    PubMed Central

    Shimizu, Y; Geraghty, D E; Koller, B H; Orr, H T; DeMars, R

    1988-01-01

    The HLA-A, -B, and -C class I human histocompatibility antigens and the genes that encode them have been isolated and characterized. Apparently complete class I non-HLA-A, B, C genes have been identified on HindIII-generated 5.4-kilobase (kb), 6.0-kb, and 6.2-kb DNA fragments derived from lymphoblastoid cell line (LCL) 721. We studied the expressibility of these genes by subcloning them into the nonintegrating pHeBo vector and transferring the chimeric plasmids into mutant LCL 721.221. This mutant was derived from LCL 721 by means of immunoselections following gamma-ray mutagenesis that eliminated expressions of the HLA-A, -B, and -C alpha chains. The HLA-A, B, C-null phenotype of mutant 721.221 made it possible to monitor the expression of class I genes transferred into it by assaying cell surface binding of monoclonal antibodies BBM.1 and W6/32, which recognize beta 2-microglobulin and HLA class I alpha-chain epitopes, respectively. Increased binding of BBM.1 and W6/32 was clearly observed in transferents containing the class I gene of the 6.0-kb DNA fragment but not in transferents containing the class I genes of the 5.4- and 6.2-kb DNA fragments. However, one-dimensional gel electrophoresis of BBM.1 and W6/32 immunoprecipitates made with [35S]methionine-labeled cell lysates showed that transfer of each non-HLA-A, B, C class I gene into 721.221 resulted in the appearance of an alpha chain that coprecipitated with beta 2-microglobulin. The three previously unreported alpha chains differed from each other in size and were smaller than HLA-A, -B, and -C alpha chains. These observations clearly show that these three cloned, nonallelic, non-HLA-A, B, C class I genes encode alpha chains that can be expressed in human cells. Images PMID:3257565

  3. Induction of anti-EBNA-1 protein by 12-O-tetradecanoylphorbol-13-acetate treatment of human lymphoblastoid cells

    SciTech Connect

    Wen, Longthung; Tanaka, Akiko; Nonoyama, Meihan )

    1989-08-01

    Binding of the Epstein-Barr virus (EBV) nuclear antigen (EBNA-1) to BamHI-C DNA was studied by affinity column chromatography followed by immunoblotting with human serum specific for EBNA-1. Two species of EBNA-1 (68 and 70 kilodaltons) were identified in nuclear extracts of the EBV-positive Burkitt's lymphoma cell line Raji and not in nuclear extracts of the EBV-negative Burkitt's lymphoma cell line BJAB. Both EBNA-1s bound specifically to the region required for EBV plasmid DNA maintenance (oriP) located in the BamHI-C fragment. Upon treatment with 12-O-tetradecanoylphorbol-13-acetate, which activates latent EBV genome in Raji cells, the 68-kilodalton EBNA-1 was uncoupled from binding to EBV oriP. Nuclear extracts from 12-O-tetradecanoylphorbol-13-acetate-treated BJAB cells also uncoupled the binding of both EBNA-1s to oriP. DNA-cellulose column chromatography identified two protein species which competed for and uncoupled the binding of EBNA-1 to oriP. The two cellular competitors the authors called anti-EBNA-1 proteins had molecular masses of 60 and 40 kilodaltons, respectively. They were not found in nuclear extracts of BJAB cells not activated by 12-O-tetradecanoylphorbol-13-acetate.

  4. Effects of Vitamin K3 and K5 on Daunorubicin-resistant Human T Lymphoblastoid Leukemia Cells.

    PubMed

    Nakaoka, Eri; Tanaka, Sachiko; Onda, Kenji; Sugiyama, Kentaro; Hirano, Toshihiko

    2015-11-01

    Anticancer efficacy of vitamin K derivatives on multidrug-resistant cancer cells has been scarcely investigated. The effects of vitamins K3 and K5 on proliferation of human leukemia MOLT-4 cells and on daunorubicin-resistant MOLT-4/DNR cells were estimated by a WST assay. Apoptotic cells were detected by Annexin V and propidium iodide staining, followed by flow cytometry. Vitamins K3 and K5 significantly inhibited proliferation of leukemic cells at 10 and 100 μM (p<0.05), and these effects were almost equally observed in both MOLT-4 and MOLT/DNR drug-resistant cells. Vitamin K3 induced cell apoptosis at 10 and 100 μM in both MOLT-4 and MOLT-4/DNR cells (p<0.05). Vitamin K5 also increased apoptotic cells, while rather inducing necrotic cell death. Vitamins K3 and K5 suppress MOLT-4 and MOLT-4/DNR cell-proliferation partially through induction of apoptosis, and these vitamin derivatives can overcome drug resistance due to P-glycoprotein expression. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  5. Method for cloning lymphoblastoid cells

    SciTech Connect

    Hammerling, U.; Kosinski, S.

    1989-02-14

    A method is described for increasing cloning frequency of human lymphocyte or lumphoblastoid cells which have been transformed with Epstein Barr virus comprising growing the transformed cells in a semi-solid agarose medium. A lower and an upper layer of agarose are used, the lower layer comprising fibroblasts suspended in the agarose layer and the upper layer comprising irradiated fibroblasts and the transformed cells suspended in the agarose layer wherein the upper agarose layer is added after the lower layer has gelled.

  6. [Production of a dialysable transfer factor of cell mediated immunity by lymphoblastoid cells in continuous proliferation].

    PubMed

    Goust, J M; Viza, D; Moulias, R; Trejdosiewicz, L; Lesourd, B; Marescot, M R; Prévot, A

    1975-01-20

    Four lymphoblastoid cell lines tested in this work contain normally a dialysable moiety having by ultraviolet spectroscopy, column chromatography (Biogel P 10) and chemically the same properties than human dialysable Transfer Factor (TFd), but unable to transfer cell mediated immune response against common antigens. Two of them are able to do so after incubation with minimal amounts of TFd. Production of a molecule identical to human TFd is possible in some lymphoblastoid cell lines after induction with TFd.

  7. Apoptotic death induced by the cyclophosphamide analogue mafosfamide in human lymphoblastoid cells: Contribution of DNA replication, transcription inhibition and Chk/p53 signaling

    SciTech Connect

    Goldstein, Michael; Roos, Wynand P. Kaina, Bernd

    2008-05-15

    Cyclophosphamide is one of the most often used anticancer drugs. Although DNA interstrand cross-links are considered responsible for its cytotoxicity, the mechanism of initiation and execution of cell death is largely unknown. Using the cyclophosphamide analogue mafosfamide, which does not need metabolic activation, we show that mafosfamide induces apoptosis dose and time dependently in lymphoblastoid cells, with clearly more apoptosis in p53{sup wt} cells. We identified two upstream processes that initiate apoptosis, DNA replication blockage and transcriptional inhibition. In lymphoblastoid cells, wherein DNA replication can be switched off by tetracycline, proliferation is required for inducing apoptosis at low dose mafosfamide. At high dose, transcriptional inhibition also contributes to cell death. The RNA synthesis inhibitor {alpha}-amanitin induced similar to mafosfamide more apoptosis in p53{sup wt} than in p53{sup mt} cells. In combination with mafosfamide, however, {alpha}-amanitin had no additive effect. Mafosfamide caused p53 stabilization by phosphorylation of Ser15, 20 and 37, and activation of ATM/ATR and Chk1/Chk2. Inhibition of ATM/ATR, PI3-kinase and Chk1/Chk2 by CGK733, wortmannin and DBH, respectively, attenuated the apoptotic response in p53{sup wt} but not p53{sup mt} cells. Mafosfamide induced caspase dependent apoptosis and, for low dose treated cells, caspases were preferentially activated in the S-phase, whereas at high dose caspases were activated in all cell cycle stages. These data support the conclusion that at low dose level of mafosfamide, DNA replication blockage is the dominant apoptosis-inducing event, while at high dose, transcriptional inhibition comes into play. The data provide a mechanistic explanation of why cyclophosphamide applied at therapeutic doses preferentially kills replicating tumor cells.

  8. Proliferation rates and gene expression profiles in human lymphoblastoid cell lines from patients with depression characterized in response to antidepressant drug therapy.

    PubMed

    Breitfeld, J; Scholl, C; Steffens, M; Brandenburg, K; Probst-Schendzielorz, K; Efimkina, O; Gurwitz, D; Ising, M; Holsboer, F; Lucae, S; Stingl, J C

    2016-11-15

    The current therapy success of depressive disorders remains in need of improvement due to low response rates and a delay in symptomatic improvement. Reliable functional biomarkers would be necessary to predict the individual treatment outcome. On the basis of the neurotrophic hypothesis of antidepressant's action, effects of antidepressant drugs on proliferation may serve as tentative individual markers for treatment efficacy. We studied individual differences in antidepressant drug effects on cell proliferation and gene expression in lymphoblastoid cell lines (LCLs) derived from patients treated for depression with documented clinical treatment outcome. Cell proliferation was characterized by EdU (5-ethynyl-2'-deoxyuridine) incorporation assays following a 3-week incubation with therapeutic concentrations of fluoxetine. Genome-wide expression profiling was conducted by microarrays, and candidate genes such as betacellulin-a gene involved in neuronal stem cell regeneration-were validated by quantitative real-time PCR. Ex vivo assessment of proliferation revealed large differences in fluoxetine-induced proliferation inhibition between donor LCLs, but no association with clinical response was observed. Genome-wide expression analyses followed by pathway and gene ontology analyses identified genes with different expression before vs after 21-day incubation with fluoxetine. Significant correlations between proliferation and gene expression of WNT2B, FZD7, TCF7L2, SULT4A1 and ABCB1 (all involved in neurogenesis or brain protection) were also found. Basal gene expression of SULT4A1 (P=0.029), and gene expression fold changes of WNT2B by ex vivo fluoxetine (P=0.025) correlated with clinical response and clinical remission, respectively. Thus, we identified potential gene expression biomarkers eventually being useful as baseline predictors or as longitudinal targets in antidepressant therapy.

  9. Proliferation rates and gene expression profiles in human lymphoblastoid cell lines from patients with depression characterized in response to antidepressant drug therapy

    PubMed Central

    Breitfeld, J; Scholl, C; Steffens, M; Brandenburg, K; Probst-Schendzielorz, K; Efimkina, O; Gurwitz, D; Ising, M; Holsboer, F; Lucae, S; Stingl, J C

    2016-01-01

    The current therapy success of depressive disorders remains in need of improvement due to low response rates and a delay in symptomatic improvement. Reliable functional biomarkers would be necessary to predict the individual treatment outcome. On the basis of the neurotrophic hypothesis of antidepressant's action, effects of antidepressant drugs on proliferation may serve as tentative individual markers for treatment efficacy. We studied individual differences in antidepressant drug effects on cell proliferation and gene expression in lymphoblastoid cell lines (LCLs) derived from patients treated for depression with documented clinical treatment outcome. Cell proliferation was characterized by EdU (5-ethynyl-2'-deoxyuridine) incorporation assays following a 3-week incubation with therapeutic concentrations of fluoxetine. Genome-wide expression profiling was conducted by microarrays, and candidate genes such as betacellulin—a gene involved in neuronal stem cell regeneration—were validated by quantitative real-time PCR. Ex vivo assessment of proliferation revealed large differences in fluoxetine-induced proliferation inhibition between donor LCLs, but no association with clinical response was observed. Genome-wide expression analyses followed by pathway and gene ontology analyses identified genes with different expression before vs after 21-day incubation with fluoxetine. Significant correlations between proliferation and gene expression of WNT2B, FZD7, TCF7L2, SULT4A1 and ABCB1 (all involved in neurogenesis or brain protection) were also found. Basal gene expression of SULT4A1 (P=0.029), and gene expression fold changes of WNT2B by ex vivo fluoxetine (P=0.025) correlated with clinical response and clinical remission, respectively. Thus, we identified potential gene expression biomarkers eventually being useful as baseline predictors or as longitudinal targets in antidepressant therapy. PMID:27845776

  10. Diverse delayed effects in human lymphoblastoid cells surviving exposure to high-LET (56)Fe particles or low-LET (137)Cs gamma radiation

    NASA Technical Reports Server (NTRS)

    Evans, H. H.; Horng, M. F.; Ricanati, M.; Diaz-Insua, M.; Jordan, R.; Schwartz, J. L.

    2001-01-01

    To obtain information on the origin of radiation-induced genomic instability, we characterized a total of 166 clones that survived exposure to (56)Fe particles or (137)Cs gamma radiation, isolated approximately 36 generations after exposure, along with their respective control clones. Cytogenetic aberrations, growth alterations, responses to a second irradiation, and mutant frequencies at the Na(+)/K(+) ATPase and thymidine kinase loci were determined. A greater percentage of clones that survived exposure to (56)Fe particles exhibited instability (defined as clones showing one or more outlying characteristics) than in the case of those that survived gamma irradiation. The phenotypes of the unstable clones that survived exposure to (56)Fe particles were also qualitatively different from those of the clones that survived gamma irradiation. A greater percentage (20%) of the unstable clones that survived gamma irradiation than those that survived exposure to (56)Fe particles (4%) showed an altered response to the second irradiation, while an increase in the percentage of clones that had an outlying frequency of ouabain-resistant and thymidine kinase mutants was more evident in the clones exposed to (56)Fe particles than in those exposed to gamma rays. Growth alterations and increases in dicentric chromosomes were found only in clones with more than one alteration. These results underscore the complex nature of genomic instability and the likelihood that radiation-induced genomic instability arises from different original events.

  11. Diverse delayed effects in human lymphoblastoid cells surviving exposure to high-LET (56)Fe particles or low-LET (137)Cs gamma radiation

    NASA Technical Reports Server (NTRS)

    Evans, H. H.; Horng, M. F.; Ricanati, M.; Diaz-Insua, M.; Jordan, R.; Schwartz, J. L.

    2001-01-01

    To obtain information on the origin of radiation-induced genomic instability, we characterized a total of 166 clones that survived exposure to (56)Fe particles or (137)Cs gamma radiation, isolated approximately 36 generations after exposure, along with their respective control clones. Cytogenetic aberrations, growth alterations, responses to a second irradiation, and mutant frequencies at the Na(+)/K(+) ATPase and thymidine kinase loci were determined. A greater percentage of clones that survived exposure to (56)Fe particles exhibited instability (defined as clones showing one or more outlying characteristics) than in the case of those that survived gamma irradiation. The phenotypes of the unstable clones that survived exposure to (56)Fe particles were also qualitatively different from those of the clones that survived gamma irradiation. A greater percentage (20%) of the unstable clones that survived gamma irradiation than those that survived exposure to (56)Fe particles (4%) showed an altered response to the second irradiation, while an increase in the percentage of clones that had an outlying frequency of ouabain-resistant and thymidine kinase mutants was more evident in the clones exposed to (56)Fe particles than in those exposed to gamma rays. Growth alterations and increases in dicentric chromosomes were found only in clones with more than one alteration. These results underscore the complex nature of genomic instability and the likelihood that radiation-induced genomic instability arises from different original events.

  12. Cadmium chloride, benzo[a]pyrene and cyclophosphamide tested in the in vitro mammalian cell micronucleus test (MNvit) in the human lymphoblastoid cell line TK6 at Covance laboratories, Harrogate UK in support of OECD draft Test Guideline 487.

    PubMed

    Fowler, Paul; Whitwell, James; Jeffrey, Laura; Young, Jamie; Smith, Katie; Kirkland, David

    2010-10-29

    The following genotoxic chemicals were tested in the in vitro micronucleus assay, at Covance Laboratories, Harrogate, UK in the human lymphoblastoid cell line TK6. Cadmium chloride (an inorganic carcinogen), benzo[a]pyrene (a polycyclic aromatic hydrocarbon requiring metabolic activation) and cyclophosphamide (an alkylating agent requiring metabolic activation) were treated with and without cytokinesis block (by addition of cytochalasin B). This work formed part of a collaborative evaluation of the toxicity measures recommended in the draft OECD Test Guideline 487 for the in vitro micronucleus test. The toxicity measures used, capable of detecting both cytostasis and cell death, were relative population doubling, relative increase in cell counts and relative cell counts for treatments in the absence of cytokinesis block, and replication index or cytokinesis blocked proliferation index in the presence of cytokinesis block. All of the chemicals tested gave significant increases in the percentage of micronucleated cells with and without cytokinesis block at concentrations giving approximately 60% toxicity (cytostasis and cell death) or less by all of the toxicity measures used. The outcomes from this series of tests support the use of relative increase in cell counts and relative population doubling, as well as relative cell counts, as appropriate measures of cytotoxicity for the non-cytokinesis blocked in the in vitro micronucleus assay. Copyright © 2010 Elsevier B.V. All rights reserved.

  13. Mitomycin C, 5-fluoruracil, colchicine and etoposide tested in the in vitro mammalian cell micronucleus test (MNvit) in the human lymphoblastoid cell line TK6 at Novartis in support of OECD draft Test Guideline 487.

    PubMed

    Elhajouji, Azeddine

    2010-10-29

    The following reference genotoxic agents were tested in the in vitro micronucleus test, at Novartis, Basel, Switzerland. Mitomycin C, 5-fluoruracil, colchicine and etoposide were tested in the human lymphoblastoid cell line TK6, with and without cytokinesis block (in the presence of cytochalasin B). This was done in support of the toxicity measures recommended in the draft OECD Test Guideline on In Vitro Mammalian Cell Micronucleus Test (MNvit) and was part of an international collaborative work. As toxicity measures, detecting cytostasis and cell death, relative cell counts (RCC), relative increase in cell counts (RICC), and relative population doubling (RPD) were used for treatments in the absence of cytokinesis block, and replication index (RI) or cytokinesis-blocked proliferation in the presence of cytokinesis block. All four reference agents were positive in the assay with and without cytokinesis block at concentrations giving approximately 50% toxicity or less as assessed by all of the toxicity measures used. Accordingly, the results of this work support the use of relative population doubling and relative increase in cell counts, as well as relative cell counts, as appropriate measures of toxicity for the non-cytokinesis-blocked in vitro micronucleus assay.

  14. The effect of gamma irradiation on injectable human amnion collagen

    SciTech Connect

    Liu, B.C.; Harrell, R.; Davis, R.H.; Dresden, M.H.; Spira, M. )

    1989-08-01

    The effect of gamma irradiation on the physicochemical properties of injectable human amnion collagen was investigated. Pepsin-extracted human amnion collagen was purified, reconstituted, and irradiated with varying doses of gamma irradiation (0.25 Mrads to 2.5 Mrads). Gamma irradiation had a significant impact on the physical characteristics of the collagen. The neutral solubility of collagen in PBS at 45{degrees}C was decreased from 100% for the nonirradiated control sample to 16% for the 2.5 Mrads irradiated sample. SDS polyacrylamide gel electrophoresis also demonstrated the dose-dependent effect of gamma irradiation on collagen cross-links. Electron microscopic observation revealed that even at low irradiation dose (0.25 Mrads), collagen fibril diameter increased. The average diameter was 50 nm for nonirradiated control fibrils, while 4.4% of the irradiated collagen fibrils had a diameter greater than 100 nm. Irradiated collagen showed little evidence of damage. Well-preserved cross-striations were found in collagen fibrils at all doses of irradiation. Native amnion collagen irradiated with gamma rays demonstrated a slight increase in resistance to collagenase degradation compared with nonirradiated native collagen samples. Increased resistance to collagenase did not correlate with increasing irradiation dose. After 30 min of incubation at 37{degrees}C, both irradiated and nonirradiated collagen was completely digested by collagenase. However, gamma-irradiated collagen did become more sensitive to hydrolysis by trypsin. The higher the irradiation doses used, the greater sensitivity to trypsin was observed. At 0.25 Mrads irradiation only a slight increase was found. No marked differences in amino acid composition were noted among the high dose irradiated, low dose irradiated and control amnion collagen.

  15. Biased dA/dT somatic hypermutation as regulated by the heavy chain intronic iEµ enhancer and 3′Eα enhancers in human lymphoblastoid B cells

    PubMed Central

    Komori, Atsumasa; Xu, Zhenming; Wu, Xiaoping; Zan, Hong; Casali, Paolo

    2015-01-01

    Somatic hypermutation (SHM) in immunoglobulin gene (Ig) variable (V) regions is critical for the maturation of the antibody response. It is dependent on the expression of activation-induced cytidine deaminase (AID) and translesion DNA polymerases in germinal center B cells as well as Ig V transcription, as regulated by the Ig heavy chain (H) intronic enhancer (iEµ) and the 3′ enhancer (3′Eα) region. We analyzed the role of these cis elements in SHM by stably transfecting Ramos human lymphoblastoid B cells with a rearranged human IgH chain VD (diversity) J (joining) DNA construct containing a VH promoter at the 5′ end and CH1 and CH2 exons of Cγ1 at the 3′ end. In this construct, mutations preferentially targeted dA/dT basepairs in the RGYW/WRCY hotspot. Most of the dA/dT mutations and accompanying dC/dG mutations were transitions. Deletion of iEµ resulted in decreased SHM which could be partially restored by insertion of the IgH hs1,2 enhancer. Other two 3′Eα enhancers, hs3-hs4, did not significantly increase the mutation frequency, but further strengthened the dA/dT bias. The frequency and spectrum of the mutations were independent of the genomic integration of the transgene or V gene transcription level. Thus, we have established a novel in vitro system to analyze SHM and identify the role of multiple cis-regulatory elements in regulating dA/dT biased SHM. This model system will be useful to further address the role of other cis-regulating elements and recruited trans-acting factors in expressing the modalities of SHM. PMID:16412510

  16. Different toxic effects of YTX in tumor K-562 and lymphoblastoid cell lines.

    PubMed

    Fernández-Araujo, Andrea; Sánchez, Jon A; Alfonso, Amparo; Vieytes, Mercedes R; Botana, Luis M

    2015-01-01

    Yessotoxin (YTX) modulates cellular phosphodiesterases (PDEs). In this regard, opposite effects had been described in the tumor model K-562 cell line and fresh human lymphocytes in terms of cell viability, cyclic adenosine 3',5'-cyclic monophosphate (cAMP) production and protein expression after YTX treatment. Studies in depth of the pathways activated by YTX in K-562 cell line, have demonstrated the activation of two different cell death types, apoptosis, and autophagy after 24 and 48 h of treatment, respectively. Furthermore, the key role of type 4A PDE (PDE4A) in both pathways activated by YTX was demonstrated. Therefore, taking into account the differences between cellular lines and fresh cells, a study of cell death pathways activated by YTX in a non-tumor cell line with mitotic activity, was performed. The cellular model used was the lymphoblastoid cell line that represents a non-tumor model with normal apoptotic and mitotic machinery. In this context, cell viability and cell proliferation, expression of proteins involved in cell death activated by YTX and mitochondrial mass, were studied after the incubation with the toxin. Opposite to the tumor model, no cell death activation was observed in lymphoblastoid cell line in the presence of YTX. In this sense, variations in apoptosis hallmarks were not detected in the lymphoblastoid cell line after YTX incubation, whereas this type I of programmed cell death was observed in K-562 cells. On the other hand, autophagy cell death was triggered in this cellular line, while other autophagic process is suggested in lymphoblastoid cells. These YTX effects are related to PDE4A in both cellular lines. In addition, while cell death is triggered in K-562 cells after YTX treatment, in lymphoblastoid cells the toxin stops cellular proliferation. These results point to YTX as a specific toxic compound of tumor cells, since in the non-tumor lymphoblastoid cell line, no cell death hallmarks are observed.

  17. Different toxic effects of YTX in tumor K-562 and lymphoblastoid cell lines

    PubMed Central

    Fernández-Araujo, Andrea; Sánchez, Jon A.; Alfonso, Amparo; Vieytes, Mercedes R.; Botana, Luis M.

    2015-01-01

    Yessotoxin (YTX) modulates cellular phosphodiesterases (PDEs). In this regard, opposite effects had been described in the tumor model K-562 cell line and fresh human lymphocytes in terms of cell viability, cyclic adenosine 3',5'-cyclic monophosphate (cAMP) production and protein expression after YTX treatment. Studies in depth of the pathways activated by YTX in K-562 cell line, have demonstrated the activation of two different cell death types, apoptosis, and autophagy after 24 and 48 h of treatment, respectively. Furthermore, the key role of type 4A PDE (PDE4A) in both pathways activated by YTX was demonstrated. Therefore, taking into account the differences between cellular lines and fresh cells, a study of cell death pathways activated by YTX in a non-tumor cell line with mitotic activity, was performed. The cellular model used was the lymphoblastoid cell line that represents a non-tumor model with normal apoptotic and mitotic machinery. In this context, cell viability and cell proliferation, expression of proteins involved in cell death activated by YTX and mitochondrial mass, were studied after the incubation with the toxin. Opposite to the tumor model, no cell death activation was observed in lymphoblastoid cell line in the presence of YTX. In this sense, variations in apoptosis hallmarks were not detected in the lymphoblastoid cell line after YTX incubation, whereas this type I of programmed cell death was observed in K-562 cells. On the other hand, autophagy cell death was triggered in this cellular line, while other autophagic process is suggested in lymphoblastoid cells. These YTX effects are related to PDE4A in both cellular lines. In addition, while cell death is triggered in K-562 cells after YTX treatment, in lymphoblastoid cells the toxin stops cellular proliferation. These results point to YTX as a specific toxic compound of tumor cells, since in the non-tumor lymphoblastoid cell line, no cell death hallmarks are observed. PMID:26136685

  18. Poly(ADP-ribosyl)ation enhances H-RAS protein stability and causes abnormal cell cycle progression in human TK6 lymphoblastoid cells treated with hydroquinone.

    PubMed

    Liu, Linhua; Ling, Xiaoxuan; Tang, Huanwen; Chen, Jialong; Wen, Qiaosheng; Zou, Fei

    2015-08-05

    Hydroquinone (HQ), one of the most important benzene-derived metabolites, can induce aberrant cell cycle progression; however, the mechanism of this induction remains unclear. Poly(ADP-ribosyl)ation (PARylation), which is catalysed primarily by poly(ADP-ribose) polymerase-1 (PARP-1), participates in various biological processes, including cell cycle control. The results of the present study show an accumulation in G1 phase versus S phase of TK6 human lymphoblast cells treated with HQ for 48h compared with PBS-treated cells; after 72h of HQ treatment, the cells transitioned from G1 arrest to S phase arrest. We examined the expression of six genes related to the cell cycle or leukaemia to further explore the reason for this phenomenon. Among these genes, H-RAS was found to be associated with this phenomenon because its mRNA and protein expression decreased at 48h and increased at 72h. Experiments for PARP activity induction and inhibition revealed that the observed PARylation was positively associated with H-RAS expression. Moreover, in cells treated with HQ in conjunction with PARP-1 knockdown, expression of the H-RAS protein decreased and the number of cells in G1 phase increased. The degree of poly(ADP-ribosyl) modification of the H-RAS protein increased in cells treated with HQ for 72h, further supporting that changes in PARylation contributed to the rapid alteration of H-RAS protein expression, followed by abnormal progression of the cell cycle. Co-immunoprecipitation (co-IP) assays were employed to determine whether protein complexes were formed by PARP-1 and H-RAS proteins, and the direct interaction between these proteins indicated that PARylation regulated H-RAS expression. As detected by confocal microscopy, the H-RAS protein was found in the nucleus and cytoplasm. To our knowledge, this study is the first to reveal that H-RAS protein can be modified by PARylation.

  19. Irradiation strongly reduces tumorigenesis of human induced pluripotent stem cells.

    PubMed

    Inui, Shoki; Minami, Kazumasa; Ito, Emiko; Imaizumi, Hiromasa; Mori, Seiji; Koizumi, Masahiko; Fukushima, Satsuki; Miyagawa, Shigeru; Sawa, Yoshiki; Matsuura, Nariaki

    2017-03-03

    Induced pluripotent stem (iPS) cells have demonstrated they can undergo self-renewal, attain pluripotency, and differentiate into various types of functional cells. In clinical transplantation of iPS cells, however, a major problem is the prevention of tumorigenesis. We speculated that tumor formation could be inhibited by means of irradiation. Since the main purpose of this study was to explore the prevention of tumor formation in human iPS (hiPS) cells, we tested the effects of irradiation on tumor-associated factors such as radiosensitivity, pluripotency and cell death in hiPS cells. The irradiated hiPS cells showed much higher radiosensitivity, because the survival fraction of hiPS cells irradiated with 2 Gy was < 10%, and there was no change of pluripotency. Irradiation with 2 and 4 Gy caused substantial cell death, which was mostly the result of apoptosis. Irradiation with 2 Gy was detrimental enough to cause loss of proliferation capability and trigger substantial cell death in vitro. The hiPS cells irradiated with 2 Gy were injected into NOG mice (NOD/Shi-scid, IL-2 Rγnull) for the analysis of tumor formation. The group of mice into which hiPS cells irradiated with 2 Gy was transplanted showed significant suppression of tumor formation in comparison with that of the group into which non-irradiated hiPS cells were transplanted. It can be presumed that this diminished rate of tumor formation was due to loss of proliferation and cell death caused by irradiation. Our findings suggest that tumor formation following cell therapy or organ transplantation induced by hiPS cells may be prevented by irradiation.

  20. UVA system for human cornea irradiation

    NASA Astrophysics Data System (ADS)

    Pereira, Fernando R. A.; Stefani, Mario; Otoboni, José A.; Richter, Eduardo H.; Rossi, Giuliano; Mota, Alessandro D.; Ventura, Liliane

    2009-02-01

    According to recent studies, an increase in corneal stiffness is a promising alternative for avoiding ectasias and for stagnating keratoconus of grades 1 and 2. The clinical treatment consists essentially of instilling Riboflavin (vitamin B2), in the cornea and then irradiating the corneal tissue, with UVA (365nm) radiation at 3mW/cm2 for 30min. This procedure provides collagen cross-linking in the corneal surface, increasing its stiffness. This work presents a system for UVA irradiation of the corneas at a peak wavelength of 365nm with adjustable power up to 5mW. The system has closed loop electronics to control the emitted power with 20% precision from the sated power output. The system is a prototype for performing corneal cross-linking and has been clinically tested. The closed loop electronics is a differential from the equipments available on the market.

  1. Lymphoblastoid Cell Lines as a Tool to Study Inter-Individual Differences in the Response to Glucose

    PubMed Central

    Grassi, Michael A.; Rao, Vidhya R.; Chen, Siquan; Cao, Dingcai; Gao, Xiaoyu; Cleary, Patricia A.; Huang, R. Stephanie; Paterson, Andrew D.; Natarajan, Rama; Rehman, Jalees; Kern, Timothy S.

    2016-01-01

    Background White blood cells have been shown in animal studies to play a central role in the pathogenesis of diabetic retinopathy. Lymphoblastoid cells are immortalized EBV-transformed primary B-cell leukocytes that have been extensively used as a model for conditions in which white blood cells play a primary role. The purpose of this study was to investigate whether lymphoblastoid cell lines, by retaining many of the key features of primary leukocytes, can be induced with glucose to demonstrate relevant biological responses to those found in diabetic retinopathy. Methods Lymphoblastoid cell lines were obtained from twenty-three human subjects. Differences between high and standard glucose conditions were assessed for expression, endothelial adhesion, and reactive oxygen species. Results Collectively, stimulation of the lymphoblastoid cell lines with high glucose demonstrated corresponding changes on molecular, cellular and functional levels. Lymphoblastoid cell lines up-regulated expression of a panel of genes associated with the leukocyte-mediated inflammation found in diabetic retinopathy that include: a cytokine (IL-1B fold change = 2.11, p-value = 0.02), an enzyme (PKCB fold change = 2.30, p-value = 0.01), transcription factors (NFKB-p50 fold change = 2.05, p-value = 0.01), (NFKB-p65 fold change = 2.82, p-value = 0.003), and an adhesion molecule (CD18 fold change = 2.59, 0.02). Protein expression of CD18 was also increased (p-value = 2.14x10-5). The lymphoblastoid cell lines demonstrated increased adhesiveness to endothelial cells (p = 1.28x10-5). Reactive oxygen species were increased (p = 2.56x10-6). Significant inter-individual variation among the lymphoblastoid cell lines in these responses was evident (F = 18.70, p < 0.0001). Conclusions Exposure of lymphoblastoid cell lines derived from different human subjects to high glucose demonstrated differential and heterogeneous gene expression, adhesion, and cellular effects that recapitulated features found in

  2. Distribution of sensitivity to 4-nitroquinoline 1-oxide among Japanese lymphoblastoid cell lines

    SciTech Connect

    Kiyohara, Chikako; Hirohata, Tomio; Nagayama, Junya ); Kuratsune, Masanori Nakamura Junior Coll., Fukuoka )

    1991-01-01

    The processes through which the UV-mimic chemical carcinogen, 4-nitroquinoline 1-oxide (4NQO), leads to the DNA lesions are well characterized in E. coli, where the formation of stable 4NQO-purine adducts is critical. The DNA excision-repair mechanisms similar to those for E. coli occur in normal human cells. Xeroderma pigmentosum (XP) is an example of a rare recessive autosomal skin disorder which is characterized biochemically as a DNA repair-deficient disease. The fluorescein diacetate (FDA) method was recently used to determine the sensitivity of lymphoblastoid cell lines 4NQO. Viable cells take up, non-fluorescent chemical, FDA and convert it to, a fluorescent molecule, fluorescein by intracellular esterases. DNA damage produced by 4NQO could be evaluated on the basis of the cell lethality by this FDA method. In the present study the authors describe the distribution of sensitivity to 4NQO among lymphoblastoid cell lines established from Japanese.

  3. Effect of Low Dose Gamma Irradiation together with Lipid A on Human Leukocytes Activities In Vitro

    NASA Astrophysics Data System (ADS)

    Belyakova, E.; Dubnickova, M.; Boreyko, A.

    2010-01-01

    The influence of gamma irradiation and of Lipid A from Escherichia coli on phagocytosis, lyzosyme and peroxidase activities of human leukocytes, in vitro was investigated. Leukocytes samples were irradiated with 1 and 5 Gy, respectively. The number of irradiated leukocytes was decreased in the irradiated samples. Only samples with additive Lipid A were not damaged by irradiation. The Lipid A had positive influence on biological activities of the irradiated leukocytes.

  4. Human cytomegalovirus replicates in gamma-irradiated fibroblasts

    SciTech Connect

    Shanley, J.D.

    1986-12-01

    Because of the unique interdependence of human cytomegalovirus (HCMV) and the physiological state of the host cell, we evaluated the ability of human foreskin fibroblasts (HFF), exposed to gamma radiation, to support HCMV growth. Irradiation of HFF with 2,500 rADS prevented cellular proliferation and suppressed cellular DNA, but not RNA or protein synthesis. Treatment of HFF cells with 2,500 rADS 6 or 48 hours prior to infection did not alter the time course or virus yield during HCMV replication. Virus plaquing efficiency in irradiated cells was comparable to that of nonirradiated cells. As judged by thymidine incorporation and BUdR inhibition of virus replication, HCMV infection induced both thymidine kinase activity and host cell DNA synthesis in irradiated cells. In addition, virus could be recovered from HFF exposed to radiation 0-2 days after infection with HCMV. These studies indicate that the damage to cells by gamma irradiation does not alter the capacity of host cells to support HCMV replication.

  5. Influence of age, irradiation and humanization on NSG mouse phenotypes.

    PubMed

    Knibbe-Hollinger, Jaclyn S; Fields, Natasha R; Chaudoin, Tammy R; Epstein, Adrian A; Makarov, Edward; Akhter, Sidra P; Gorantla, Santhi; Bonasera, Stephen J; Gendelman, Howard E; Poluektova, Larisa Y

    2015-09-09

    Humanized mice are frequently utilized in bench to bedside therapeutic tests to combat human infectious, cancerous and degenerative diseases. For the fields of hematology-oncology, regenerative medicine, and infectious diseases, the immune deficient mice have been used commonly in basic research efforts. Obstacles in true translational efforts abound, as the relationship between mouse and human cells in disease pathogenesis and therapeutic studies requires lengthy investigations. The interplay between human immunity and mouse biology proves ever more complicated when aging, irradiation, and human immune reconstitution are considered. All can affect a range of biochemical and behavioral functions. To such ends, we show age- and irradiation-dependent influences for the development of macrocytic hyper chromic anemia, myelodysplasia, blood protein reductions and body composition changes. Humanization contributes to hematologic abnormalities. Home cage behavior revealed day and dark cycle locomotion also influenced by human cell reconstitutions. Significant age-related day-to-day variability in movement, feeding and drinking behaviors were observed. We posit that this data serves to enable researchers to better design translational studies in this rapidly emerging field of mouse humanization.

  6. Influence of age, irradiation and humanization on NSG mouse phenotypes

    PubMed Central

    Knibbe-Hollinger, Jaclyn S.; Fields, Natasha R.; Chaudoin, Tammy R; Epstein, Adrian A.; Makarov, Edward; Akhter, Sidra P.; Gorantla, Santhi; Bonasera, Stephen J.; Gendelman, Howard E.; Poluektova, Larisa Y.

    2015-01-01

    ABSTRACT Humanized mice are frequently utilized in bench to bedside therapeutic tests to combat human infectious, cancerous and degenerative diseases. For the fields of hematology-oncology, regenerative medicine, and infectious diseases, the immune deficient mice have been used commonly in basic research efforts. Obstacles in true translational efforts abound, as the relationship between mouse and human cells in disease pathogenesis and therapeutic studies requires lengthy investigations. The interplay between human immunity and mouse biology proves ever more complicated when aging, irradiation, and human immune reconstitution are considered. All can affect a range of biochemical and behavioral functions. To such ends, we show age- and irradiation-dependent influences for the development of macrocytic hyper chromic anemia, myelodysplasia, blood protein reductions and body composition changes. Humanization contributes to hematologic abnormalities. Home cage behavior revealed day and dark cycle locomotion also influenced by human cell reconstitutions. Significant age-related day-to-day variability in movement, feeding and drinking behaviors were observed. We posit that this data serves to enable researchers to better design translational studies in this rapidly emerging field of mouse humanization. PMID:26353862

  7. Assay for mutagenesis in heterozygous diploid human lymphoblasts

    DOEpatents

    Skopek, Thomas R.; Liber, Howard L.; Penman, Bruce W.; Thilly, William G.; Hoppe, IV, Henry

    1981-01-01

    An assay is disclosed for determining mutagenic damage caused by the administration of a known or suspected mutagen to diploid human lymphoblastoid cell lines. The gene locus employed for this assay is the gene for thymidine kinase, uridine kinase, or cytidine deaminase. Since human lymphoblastoid cells contain two genes for these enzymes, heterozygotes of human lymphoblastoid cells are used in this assay.

  8. Mitochondrial-targeted human catalase affords neuroprotection from proton irradiation.

    PubMed

    Liao, Alicia C; Craver, Brianna M; Tseng, Bertrand P; Tran, Katherine K; Parihar, Vipan K; Acharya, Munjal M; Limoli, Charles L

    2013-07-01

    Significant past work has linked radiation exposure of the CNS to elevated levels of oxidative stress and inflammation. These secondary reactive processes are both dynamic and persistent and are believed to compromise the functionality of the CNS, in part, by disrupting endogenous neurogenesis in the hippocampus. While evidence has shown neurogenesis to be sensitive to irradiation and redox state, the mechanistic basis underlying these effects is incompletely understood. To clarify the role of reactive oxygen species (ROS) in mediating radiation-induced changes in neurogenesis we have analyzed transgenic mice that overexpress human catalase localized to the mitochondria. With this model, we investigated the consequences of low dose and clinically relevant proton irradiation on neurogenesis, and how that process is modified in response to genetic disruption of mitochondrial ROS levels. In unirradiated animals, basal neurogenesis was improved significantly by reductions in mitochondrial ROS. In animals subjected to proton exposure, hippocampal progenitor cell proliferation was attenuated significantly by overexpression of human catalase in the mitochondria. Furthermore, expression of the MCAT transgene significantly improved neurogenesis in WT animals after low-dose proton exposure (0.5 Gy), with similar trends observed at higher dose (2 Gy). Our report documents for the first time the impact of proton irradiation on hippocampal neurogenesis, and the neuroprotective properties of reducing mitochondrial ROS through the targeted overexpression of catalase. © 2013 by Radiation Research Society

  9. Genetic association with overall survival of taxane-treated lung cancer patients - a genome-wide association study in human lymphoblastoid cell lines followed by a clinical association study

    PubMed Central

    2012-01-01

    Background Taxane is one of the first line treatments of lung cancer. In order to identify novel single nucleotide polymorphisms (SNPs) that might contribute to taxane response, we performed a genome-wide association study (GWAS) for two taxanes, paclitaxel and docetaxel, using 276 lymphoblastoid cell lines (LCLs), followed by genotyping of top candidate SNPs in 874 lung cancer patient samples treated with paclitaxel. Methods GWAS was performed using 1.3 million SNPs and taxane cytotoxicity IC50 values for 276 LCLs. The association of selected SNPs with overall survival in 76 small or 798 non-small cell lung cancer (SCLC, NSCLC) patients were analyzed by Cox regression model, followed by integrated SNP-microRNA-expression association analysis in LCLs and siRNA screening of candidate genes in SCLC (H196) and NSCLC (A549) cell lines. Results 147 and 180 SNPs were associated with paclitaxel or docetaxel IC50s with p-values <10-4 in the LCLs, respectively. Genotyping of 153 candidate SNPs in 874 lung cancer patient samples identified 8 SNPs (p-value < 0.05) associated with either SCLC or NSCLC patient overall survival. Knockdown of PIP4K2A, CCT5, CMBL, EXO1, KMO and OPN3, genes within 200 kb up-/downstream of the 3 SNPs that were associated with SCLC overall survival (rs1778335, rs2662411 and rs7519667), significantly desensitized H196 to paclitaxel. SNPs rs2662411 and rs1778335 were associated with mRNA expression of CMBL or PIP4K2A through microRNA (miRNA) hsa-miR-584 or hsa-miR-1468. Conclusions GWAS in an LCL model system, joined with clinical translational and functional studies, might help us identify genetic variations associated with overall survival of lung cancer patients treated paclitaxel. PMID:23006423

  10. Response of human fibroblasts to low dose rate gamma irradiation

    SciTech Connect

    Dritschilo, A.; Brennan, T.; Weichselbaum, R.R.; Mossman, K.L.

    1984-11-01

    Cells from 11 human strains, including fibroblasts from patients with the genetic diseases of ataxia telangiectasia (AT), xeroderma pigmentosum (XP), and Fanconi's anemia (FA), were exposed to ..gamma.. radiation at high (1.6-2.2 Gy/min) and at low (0.03-0.07 Gy/min) dose rates. Survival curves reveal an increase inthe terminal slope (D/sub 0/) when cells are irradiated at low dose rates compared to high dose rates. This was true for all cell lines tested, although the AT, FA, and XP cells are reported or postulated to have radiation repair deficiencies. From the response of these cells, it is apparent that radiation sensitivities differ; however, at low dose rate, all tested human cells are able to repair injury.

  11. Effects of Electron Beam and Microwave Irradiation on Human Blood Proteins

    NASA Astrophysics Data System (ADS)

    Martin, Diana I.; Stan, Dana E.; Radu, Roxana R.; Cinca, Sabin A.; Margaritescu, Irina D.; Chirita, Doru I.; Craciun, Gabriela D.; Manaila, Elena N.; Ighigeanu, Daniel I.; Iacob, Nicusor I.; Oproiu, Constantin V.

    2007-04-01

    The effects of separated and combined accelerated electron beam (EB) of 6.23 MeV and microwave (MW) of 2.45GHz irradiation on proteins in samples of human serum, human plasma and human integral blood are presented. Also, it was studied the effect of separate and combined EB and MW irradiation on proteins irradiated in samples of human integral blood, without and in the presence of a synthetic compound solution (S.C.S.) which is expected to exhibit various biological actions, such as to diminish or to increase the irradiation effects.

  12. Effects of Electron Beam and Microwave Irradiation on Human Blood Proteins

    SciTech Connect

    Martin, Diana I.; Craciun, Gabriela D.; Manaila, Elena N.; Ighigeanu, Daniel I.; Iacob, Nicusor I.; Oproiu, Constantin V.; Stan, Dana E.; Radu, Roxana R.; Margaritescu, Irina D.; Chirita, Doru I.

    2007-04-23

    The effects of separated and combined accelerated electron beam (EB) of 6.23 MeV and microwave (MW) of 2.45GHz irradiation on proteins in samples of human serum, human plasma and human integral blood are presented. Also, it was studied the effect of separate and combined EB and MW irradiation on proteins irradiated in samples of human integral blood, without and in the presence of a synthetic compound solution (S.C.S.) which is expected to exhibit various biological actions, such as to diminish or to increase the irradiation effects.

  13. The effect of irradiation at low doses on human embryos and fetuses

    SciTech Connect

    Romanova, L.K.; Zhorova, E.S.

    1994-05-01

    Data about the biological effect of irradiation at low dose on prenatal human development have been reviewed. The effect of irradiation is observed either immediately after it or in the progeny, as consequences of irradiation affecting the embryo or fetus. Human embryos and fetuses are most sensitive to ionizing irradiation during the peaks of proliferative activity and cell differentiation. The concept has been formulated that any dose of irradiation, however low, can inflict damage to the embryo or fetus. Problems and perspectives of studies in this field are discussed.

  14. Proteomic Analysis of Proton Beam Irradiated Human Melanoma Cells

    PubMed Central

    Kedracka-Krok, Sylwia; Jankowska, Urszula; Elas, Martyna; Sowa, Urszula; Swakon, Jan; Cierniak, Agnieszka; Olko, Pawel; Romanowska-Dixon, Bozena; Urbanska, Krystyna

    2014-01-01

    Proton beam irradiation is a form of advanced radiotherapy providing superior distributions of a low LET radiation dose relative to that of photon therapy for the treatment of cancer. Even though this clinical treatment has been developing for several decades, the proton radiobiology critical to the optimization of proton radiotherapy is far from being understood. Proteomic changes were analyzed in human melanoma cells treated with a sublethal dose (3 Gy) of proton beam irradiation. The results were compared with untreated cells. Two-dimensional electrophoresis was performed with mass spectrometry to identify the proteins. At the dose of 3 Gy a minimal slowdown in proliferation rate was seen, as well as some DNA damage. After allowing time for damage repair, the proteomic analysis was performed. In total 17 protein levels were found to significantly (more than 1.5 times) change: 4 downregulated and 13 upregulated. Functionally, they represent four categories: (i) DNA repair and RNA regulation (VCP, MVP, STRAP, FAB-2, Lamine A/C, GAPDH), (ii) cell survival and stress response (STRAP, MCM7, Annexin 7, MVP, Caprin-1, PDCD6, VCP, HSP70), (iii) cell metabolism (TIM, GAPDH, VCP), and (iv) cytoskeleton and motility (Moesin, Actinin 4, FAB-2, Vimentin, Annexin 7, Lamine A/C, Lamine B). A substantial decrease (2.3 x) was seen in the level of vimentin, a marker of epithelial to mesenchymal transition and the metastatic properties of melanoma. PMID:24392146

  15. Effects of Er:YAG laser irradiation on human cartilage

    NASA Astrophysics Data System (ADS)

    Glinkowski, Wojciech; Brzozowska, Malgorzata; Ciszek, Bogdan; Rowinski, Jan; Strek, Wieslaw

    1996-03-01

    Irradiation of the hyaline or fibrous cartilage excised from the body of a human cadaver with Er:YAG laser beam, single pulse with a dose of 1 J, produces a crater with a depth of approximately 500 micrometers and a diameter varying from 5 to 300 micrometers. Histological examination has revealed that the laser-made craters were surrounded by a thin rim (2-10 micrometer) of charred and coagulated tissue. No damage was observed in the cartilage surrounding the rim. The presence of sharp demarcation between the tissue areas ablated by laser energy and the undamaged areas argues for the potential usefulness of the Er:YAG laser in surgery of cartilages.

  16. Low-energy helium-neon laser irradiation increases the motility of cultured human keratinocytes

    SciTech Connect

    Haas, A.F.; Isseroff, R.R.; Wheeland, R.G.; Rood, P.A.; Graves, P.J. )

    1990-06-01

    Helium-neon (HeNe) laser irradiation is known to stimulate wound healing. We investigated whether the biostimulatory effects of HeNe irradiation result from enhancement of keratinocyte proliferation or motility. HeNe effects on keratinocyte motility were evaluated by irradiating a wounded culture with 0.8 J/cm2 3 times over a 20-h period. At 20 h post-irradiation, videocinemicroscopy and sequential quantitative measurements of the leading edge were taken over a 6-h period. There was a significant difference in migration of the leading edge in irradiated wounds compared to non-irradiated wounded controls (12.0 microns/h vs 4.0 microns/h, p less than 0.0001). To determine if the increase in migration observed in irradiated cultures resulted from a proliferative effect of HeNe irradiation, subconfluent human keratinocyte cultures were irradiated with single or multiple doses of different fluences of HeNe irradiation (0.4 to 7.2 J/cm2) and evaluated 72 h post-irradiation. Irradiated and non-irradiated keratinocyte cultures grown on a microporous membrane surface were co-cultured with irradiated and non-irradiated fibroblasts to determine if HeNe irradiation induced a paracrine effect on keratinocyte proliferation. No significant increase in keratinocyte proliferation was demonstrated in any of these treatments. The biostimulatory effects of HeNe irradiation may now be extended to include enhancement of keratinocyte motility in vitro; this may contribute to the efficacy of HeNe irradiation in wound healing.

  17. Transplantability of human lymphoid cell line, lymphoma, and leukemia in splenectomized and/or irradiated nude mice

    SciTech Connect

    Watanabe, S.; Shimosato, Y.; Kuroki, M.; Sato, Y.; Nakajima, T.

    1980-07-01

    The effects of splenectomy and/or whole-body irradiation of nude mice before xenotransplantation of lymphoid cell lines, lymphoma, and leukemia were studied. Transplantation after whole-body irradiation resulted in the increased ''take'' rate of three cultured cell lines (two of T-cell-derived acute lymphocytic leukemia and one of B-cell derived acute lymphocytic leukemia) and in the tumorous growth of Burkitt-derived Raji and spontaneously transformed lymphoblastoid cell lines. With splenectomy plus irradiation as a pretreatment, tumorous growth occurred in four other cell lines which were not transplantable after irradiation only (two cell lines of Epstein-Barr virus-transformed cord blood cells and one each of null acute lymphocytic leukemia and nodular lymphoma-derived cell lines). Direct transplantation of leukemia and lymphoma cells into the pretreated mice was successful in 7 of 24 cases (29%). B-cell-derived diffuse large lymphoid lymphoma was transplantable in three of seven cases (43%). However, lymphoma and leukemia of peripheral T-cell origin was difficult to transplant even with pretreatment, and only one pleomorphic T-cell lymphoma grew to a significant size (2 cm). One tumor each of B-cell-derived diffuse large lymphoid and T-cell diffuse lymphoblastic lymphoma became transplantable.

  18. Functional changes induced by chronic UVA irradiation to cultured human dermal fibroblasts.

    PubMed

    Naru, E; Suzuki, T; Moriyama, M; Inomata, K; Hayashi, A; Arakane, K; Kaji, K

    2005-12-01

    Ultraviolet (UV) irradiation induces damage of the skin, and in particular, photoageing is known to be the result of chronic UV irradiation. Many investigations have attempted to clarify the mechanisms of photoageing induced by chronic UVA irradiation, but consensus has not been achieved yet by in vivo experiments, mostly due to differences among UV sources and animals used for experiments. In vitro experiments have shown that a single exposure to UVA irradiation causes overexpression of matrix metalloproteinases and denaturation of collagen, but the mechanisms of the photoageing effects of chronic UVA irradiation are still unclear. To examine the effects of chronic UVA irradiation, we used an in vitro fibroblast cellular ageing system as a model of photoageing. Chronic UVA irradiation of normal human fibroblasts induced shortening of the cellular life span and an increase of cellular diameter, in parallel with expression of senescence-associated beta-galactosidase. Extracellular degradation enzyme, matrix metalloproteinase 1 (MMP-1) was overexpressed after repeated UVA irradiation, but tissue inhibitor of metalloproteinase 1 (TIMP-1) expression was hardly changed by chronic UVA irradiation. We conclude that chronic UVA irradiation of normal human fibroblasts induces cellular functional changes, leading to accelerated cellular ageing and MMP-1 overexpression.

  19. Human cell engraftment after busulfan or irradiation conditioning of NOD/SCID mice.

    PubMed

    Robert-Richard, Elodie; Ged, Cécile; Ortet, Jacqueline; Santarelli, Xavier; Lamrissi-Garcia, Isabelle; de Verneuil, Hubert; Mazurier, Frédéric

    2006-10-01

    Human hematopoietic stem cell (HSC) xenotransplantation in NOD/SCID mice requires recipient conditioning, classically achieved by sublethal irradiation. Pretreatment with immunosuppressive and alkylating agents has been reported, but has not been rigorously tested against standard irradiation protocols. Here, we report that treatment of mice with a single dose (35 mg/kg) of Busilvex, an injectable form of busulfan, enables equivalent engraftment compared to 3.5 Gy irradiation. Mice treated with two doses of 25 mg/kg to reduce busulfan toxicity showed increased chimerism. Busulfan conditioning and irradiation resulted in comparable sensitivity of HSC detection as evaluated by limiting dilution analysis.

  20. Reactive oxygen species production in mitochondria of human gingival fibroblast induced by blue light irradiation.

    PubMed

    Yoshida, Ayaka; Yoshino, Fumihiko; Makita, Tetsuya; Maehata, Yojiro; Higashi, Kazuyoshi; Miyamoto, Chihiro; Wada-Takahashi, Satoko; Takahashi, Shun-suke; Takahashi, Osamu; Lee, Masaichi Chang-il

    2013-12-05

    In recent years, it has become well known that the production of reactive oxygen species (ROS) induced by blue-light irradiation causes adverse effects of photo-aging, such as age-related macular degeneration of the retina. Thus, orange-tinted glasses are used to protect the retina during dental treatment involving blue-light irradiation (e.g., dental resin restorations or tooth bleaching treatments). However, there are few studies examining the effects of blue-light irradiation on oral tissue. For the first time, we report that blue-light irradiation by quartz tungsten halogen lamp (QTH) or light-emitting diode (LED) decreased cell proliferation activity of human gingival fibroblasts (HGFs) in a time-dependent manner (<5 min). Additionally, in a morphological study, the cytotoxic effect was observed in the cell organelles, especially the mitochondria. Furthermore, ROS generation induced by the blue-light irradiation was detected in mitochondria of HGFs using fluorimetry. In all analyses, the cytotoxicity was significantly higher after LED irradiation compared with cytotoxicity after QTH irradiation. These results suggest that blue light irradiation, especially by LED light sources used in dental aesthetic treatment, might have adverse effects on human gingival tissue. Hence, this necessitates the development of new dental aesthetic treatment methods and/or techniques to protect HGFs from blue light irradiation during dental therapy. Copyright © 2013 Elsevier B.V. All rights reserved.

  1. Irradiated human chondrocytes expressing bone morphogenetic protein 2 promote healing of osteoporotic bone fracture in rats.

    PubMed

    Yi, Youngsuk; Choi, Kyoung Baek; Lim, Chae-Lyul; Hyun, Jong-Pil; Lee, Hyeon-Youl; Lee, Kun Bok; Yun, Lillian; Ayverdi, Asli; Hwang, Sally; Yip, Vivian; Noh, Moon Jong; Lee, Kwan Hee

    2009-10-01

    Bone morphogenetic protein 2 (BMP2) was selected as a transgene to regenerate osteoporotic bone defects after several BMPs were tested using a bone formation study in nude mice. Human chondrocytes were transduced with a BMP2-containing retroviral vector, and single clones were selected. The cells were characterized over numerous passages for growth and BMP2 expression. The single clones were irradiated and tested for viability. BMP2 expression lasted for 3 weeks before dying off completely after approximately 1 month. Irradiated and non-irradiated transduced chondrocytes successfully healed fractures in osteoporotic rats induced by ovariectomy. The osteoinducing effect of irradiated cells was better than that of their non-irradiated counterparts or a chondrocytes-only control. This study showed that delivering BMP2 from the transduced and irradiated chondrocytes could be an effective and safe method of repairing osteoporotic bone fractures.

  2. Ultraviolet-C-induced apoptosis protected by 635-nm laser irradiation in human gingival fibroblasts.

    PubMed

    Lim, Wonbong; Ko, Mikyung; Lee, Sungga; Kim, Inae; Jung, Mina; Kim, Okjoon; Cho, Seonghoun; Yang, Kyuho; Choi, Namki; Kim, Sunmi; Choi, Hongran

    2008-06-01

    The purpose of this study was to examine the protection afforded by 635-nm irradiation against ultraviolet (UV)-C-induced apoptosis in primary human gingival fibroblasts (hGFs). UV irradiation is known to cause photoaging and cellular apoptosis of skin cells and is considered to be one of the leading causes of skin carcinogenesis. To induce apoptosis, UV-C (100 mJ/cm2) was used to irradiate hGFs. To protect them from apoptosis, pretreatment with 635-nm irradiation was performed for 1 h immediately after cell plating 36 or 48 h before UV-C irradiation. The light source used for irradiation was a continuous-wave 635-nm LED laser emitting at 1 mW/cm2. Experimental samples were selected 24 h after UV-C irradiation. To measure the numbers of apoptotic cells, MTT assay and flow cytometric analyses were performed. For histomorphologic findings, Diff-Quick staining was carried out. Also, the activities and mRNA expression of caspase-3, caspase-8, and caspase-9 were measured. In the present study, the number of apoptotic cells declined in the cells that were pretreated with 635-nm light irradiation in a time-dependent manner. In addition, the activities and mRNA expression of caspase-3, caspase-8, and caspase-9 were significantly recovered by pretreatment with 635-nm irradiation. These results suggest that 635-nm visible light irradiation may be used as a protective tool to prevent UV-C-induced apoptosis.

  3. Physical and Biological Characterization of the Gamma-Irradiated Human Cornea.

    PubMed

    Chae, J Jeremy; Choi, Joseph S; Lee, Justin D; Lu, Qiaozhi; Stark, Walter J; Kuo, Irene C; Elisseeff, Jennifer H

    2015-10-01

    To compare the physical and biological characteristics of commercial gamma-irradiated corneas with those of fresh human corneas and to determine suitability for transplantation. The physical properties of gamma-irradiated and fresh corneas were evaluated with respect to light transmittance, hydration (swelling ratio), elastic modulus (compressive modulus by the indentation method), matrix organization (differential scanning calorimetry), and morphology (light and transmission electron microscopy). The biological properties of the gamma-irradiated cornea, including residual cell content and cellular biocompatibility, were evaluated by quantifying DNA content and measuring the proliferation rate of human corneal epithelial cells, respectively. The hydration, light transmittance, elastic modulus, and proliferation rate of human corneal epithelial cells were not significantly different between fresh and gamma-irradiated corneas. However, differences were observed in tissue morphology, DNA content, and thermal properties. The density of collagen fibrils of the gamma-irradiated corneal sample (160.6 ± 33.2 fibrils/μm) was significantly lower than that of the fresh corneal sample (310.0 ± 44.7 fibrils/μm). Additionally, in the gamma-irradiated corneas, cell fragments-but not viable cells-were observed, supported by lower DNA content of the gamma-irradiated cornea (1.0 ± 0.1 μg/mg) than in fresh corneas (1.9 μg/mg). Moreover, the denaturation temperature of gamma-irradiated corneas (61.8 ± 1.1 °C) was significantly lower than that of fresh corneas (66.1 ± 1.9 °C). Despite structural changes due to irradiation, the physical and biological properties of the gamma-irradiated cornea remain similar to the fresh cornea. These factors, combined with a decreased risk of rejection and longer shelf life, make the gamma-irradiated tissue a viable and clinically desired option in various ophthalmic procedures.

  4. Effect of irradiated dib-cAMP on the tonic and phasic activity of human myometrium.

    PubMed

    Schachinger, L; Srivastava, A; Schippel, C; Klöter, H

    1983-06-01

    Cyclic AMP, used as dibutyryl derivative for better permeability, has a relaxing effect on smooth muscle preparations from human uterine tissue (surgical material). The observed decrease of tonus and frequency depends on the concentration applied, shown in the range between 50 and 300 microM. cAMP looses its physiological activity by irradiation in vitro; in addition an inhibitory action of the irradiation products on uterine tissue could be proved. From the data of the Lineweaver-Burk plots, showing the competition between non-irradiated and irradiated cAMP, a ten-to twentyfold higher affinity of the irradiation products to the receptor compared to that of this transmitter could be calculated. The results show that preparations of human origin with beta-receptors behave similarly to animal tissues with beta-receptors. They are further discussed with respect to a better understanding of dose-response curves for chemical and physiological inactivation.

  5. Sorting of chromosome 13 from lymphoblastoid cell lines derived from patients with Wilson disease

    SciTech Connect

    Nasedkina, T.V.; Polesskaya, A.N.; Surkov, S.A.; Poletaev, A.I. ); Aksenov, N.; Zenin, V.V. )

    1993-01-01

    Lymphoblastoid cell lines were established from patients with Wilson disease (WD) which maps to human chromosome 13 and served as a source of chromosomes. The authors used a modified isolation procedure to increase the yield of metaphase chromosomes and additional purification of the chromosome suspension on Percoll gradient to achieve more stable sorting conditions. Vibariate flow analysis using dual laser cell-sorter, ATC-3000, showed a sufficient resolution of the flow karyotype and a low level of debris. They sorted chromosome 13 at a speed of up to 5,000 chr/sec, providing about 2 million chromosomes per day. The purity of the sorted fraction was about 90%. The fractions will be further used to construct cosmid libraries to facilitate studies of the WD locus.

  6. Study of nuclear proteins in normal and xeroderma pigmentosum lymphoblastoid cells

    SciTech Connect

    Amari, N.M.B.

    1985-01-01

    Nuclear histone and nonhistone (NHP) proteins from normal human and xeroderma pigmentosum, complementation group A (XP-A) lymphoblastoid cells were compared both qualitatively, quantitatively and for binding affinity for DNA. Histones and four NHP fractions (NHP/sub 1-4/) were isolated from purified cell nuclei. Binding affinity to (/sup 3/H) melanoma DNA of histones and each NHP fraction was then determined using gradient dialysis followed by a filter assay. Histones and each NHP fraction were then sub-fractionated by polyacrylamide gel electrophoresis. Densitometric scans of the separation of these proteins on the gels were qualitatively, and quantitatively analyzed and compared between the two cell lines. No qualitative or quantitative differences were observed between histones from XP-A or normal cells.

  7. Controversial effects of low level laser irradiation on the proliferation of human osteoblasts

    NASA Astrophysics Data System (ADS)

    Bölükbaşı Ateş, Gamze; Ak, Ayşe.; Garipcan, Bora; Yüksel, Šahru; Gülsoy, Murat

    2015-03-01

    Low level laser irradiation (LLLI) is the application of red or near infrared lasers irradiating between 600-1100 nm with an output power of 1-500 mW. Several researches indicate that LLLI modulates cellular mechanisms and leads to enhance proliferation. Although the biological mechanisms are not fully understood, it is known that the effects depend on several parameters such as wavelength, irradiation duration, energy level, beam type and energy density. The aim of this study is to investigate the effect of low level laser irradiation at varying energy densities with two different wavelengths (635 nm and 809 nm) on the proliferation of human osteoblasts in vitro. The cells are seeded on 96 well plates (105cells/well) and after 24 h incubation cells are irradiated at energy densities 0.5 J/cm2, 1 J/cm2 and 2 J/cm2. Cell viability test is applied after 24 h, 48 h and 72 h in order to examine effects of laser irradiation on osteoblast proliferation. 635 nm light irradiation did not appear to have significant effect on the proliferation of osteoblasts as compared to the control. On the other hand, 809 nm laser irradiation caused significant (p ≤ 0.01) biostimulation effect on the osteoblast cell cultures at 48 h and 72 h. In conclusion, irradiation of both wavelengths did not cause any cytotoxic effects. 809 nm light irradiation can promote proliferation of human osteoblasts in vitro. On the other hand, 635 nm light irradiation has no positive effect on osteoblast proliferation. As a result, LLLI applied using different wavelengths on the same cell type may lead to different biological effects.

  8. Inactivation of a Human Norovirus Surrogate, Human Norovirus Virus-Like Particles, and Vesicular Stomatitis Virus by Gamma Irradiation

    PubMed Central

    Feng, Kurtis; Divers, Erin; Ma, Yuanmei; Li, Jianrong

    2011-01-01

    Gamma irradiation is a nonthermal processing technology that has been used for the preservation of a variety of food products. This technology has been shown to effectively inactivate bacterial pathogens. Currently, the FDA has approved doses of up to 4.0 kGy to control food-borne pathogens in fresh iceberg lettuce and spinach. However, whether this dose range effectively inactivates food-borne viruses is less understood. We have performed a systematic study on the inactivation of a human norovirus surrogate (murine norovirus 1 [MNV-1]), human norovirus virus-like particles (VLPs), and vesicular stomatitis virus (VSV) by gamma irradiation. We demonstrated that MNV-1 and human norovirus VLPs were resistant to gamma irradiation. For MNV-1, only a 1.7- to 2.4-log virus reduction in fresh produce at the dose of 5.6 kGy was observed. However, VSV was more susceptible to gamma irradiation, and a 3.3-log virus reduction at a dose of 5.6 kGy in Dulbecco's modified Eagle medium (DMEM) was achieved. We further demonstrated that gamma irradiation disrupted virion structure and degraded viral proteins and genomic RNA, which resulted in virus inactivation. Using human norovirus VLPs as a model, we provide the first evidence that the capsid of human norovirus has stability similar to that of MNV-1 after exposure to gamma irradiation. Overall, our results suggest that viruses are much more resistant to irradiation than bacterial pathogens. Although gamma irradiation used to eliminate the virus contaminants in fresh produce by the FDA-approved irradiation dose limits seems impractical, this technology may be practical to inactivate viruses for other purposes, such as sterilization of medical equipment. PMID:21441330

  9. Lymphoblastoid lines and skin fibroblasts from patients with tuberous sclerosis are abnormally sensitive to ionizing radiation and to a radiomimetic chemical

    SciTech Connect

    Scudiero, D.A.; Moshell, A.N.; Scarpinato, R.G.; Meyer, S.A.; Clatterbuck, B.E.; Tarone, R.E.; Robbins, J.H.

    1982-03-01

    Lymphoblastoid lines, derived by transforming peripheral blood lymphocytes with Epstein-Barr virus, and skin fibroblast lines were established from two patients with tuberous sclerosis. The number of viable lymphoblastoid cells was determined by their ability to exclude the vital dye trypan blue after their irradiation with x-rays or 254 nm ultraviolet light. The growth of fibroblasts was determined by their ability to form colonies after treatment with the radiomimetic, DNA-damaging chemical N-methyl-N'-nitro-N-nitrosoguanidine. The tuberous sclerosis lymphoblastoid lines were hypersensitive to x-rays but had normal sensitivity to the ultraviolet radiation. The tuberous sclerosis fibroblast lines were hypersensitive to the N-methyl-N'-nitro-N-nitrosoguanidine. The hypersensitivity of tuberous sclerosis cells to x-rays and to N-methyl-N'-nitro-N-nitrosoguanidine is believed to reflect defective repair of DNA damaged by these agents and may provide the basis for in vitro, including prenatal, diagnostic tests for tuberous sclerosis.

  10. Vanillin protects human keratinocyte stem cells against ultraviolet B irradiation.

    PubMed

    Lee, Jienny; Cho, Jae Youl; Lee, Sang Yeol; Lee, Kyung-Woo; Lee, Jongsung; Song, Jae-Young

    2014-01-01

    Ultraviolet-B (UVB) irradiation is one of major factors which induce cellular damages in the epidermis. We investigated protective effects and mechanisms of vanillin, a main constituent of vanilla beans, against UVB-induced cellular damages in keratinocyte stem cells (KSC). Here, vanillin significantly attenuated UVB irradiation-induced cytotoxicity. The vanillin effects were also demonstrated by the results of the senescence-associated β-galactosidase and alkaline comet assays. In addition, vanillin induced production of pro-inflammatory cytokines. Attempts to elucidate a possible mechanism underlying the vanillin-mediated effects revealed that vanillin significantly reduced UVB-induced phosphorylation of ataxia telangiectasia mutated (ATM), serine threonine kinase checkpoint kinase 2 (Chk2), tumor suppressor protein 53 (p53), p38/mitogen-activated protein kinase (p38), c-Jun N-terminal kinase/stress-activated protein kinase (JNK), S6 ribosomal protein (S6RP), and histone 2A family member X (H2A.X). UVB-induced activation of p53 luciferase reporter was also significantly inhibited by vanillin. In addition, while ATM inhibitor had no effect on the vanillin effects, mouse double minute 2 homolog (MDM2) inhibitor significantly attenuated suppressive effects of vanillin on UVB-induced activation of p53 reporter in KSC. Taken together, these findings suggest that vanillin protects KSC from UVB irradiation and its effects may occur through the suppression of downstream step of MDM2 in UVB irradiation-induced p53 activation.

  11. Effects of water-filtered infrared A irradiation on human fibroblasts.

    PubMed

    Jung, Tobias; Höhn, Annika; Piazena, Helmut; Grune, Tilman

    2010-01-01

    Infrared radiation is a substantial part of the solar energy output reaching the earth surface. Therefore, exposure of humans to infrared radiation is common. However, whether and how infrared (IR) or infrared A acts on human skin cells is still under debate. Recently the generation of reactive oxygen species by water-filtered infrared A (wIRA) irradiation was postulated. wIRA shows a spectral distribution similar to that of solar irradiation at the earth's surface. Thus, the need for protection of human skin from both solar- and artificially generated infrared A irradiation was concluded. Here we demonstrate that in human dermal fibroblasts this reactive oxygen species generation is dependent on heat formation by infrared A and can be reproduced by thermal exposure. On the other hand wIRA irradiation had no detectable effect if the temperature in the cells was kept constant, even if irradiance exceeded the extraterrestrial solar irradiance in the IR range by a factor of about 4 and the maximum at noontime in the tropics by a factor up to about 6. This could be demonstrated by the measurement of oxidant formation using H(2)DCFDA and the determination of protein carbonyls. In additional experiments we could show that during thermal exposure the mitochondria contribute significantly to oxidant production. Further experiments revealed that the major absorbance of infrared is due to absorption of the energy by cellular water. Copyright 2009 Elsevier Inc. All rights reserved.

  12. Impact of blue LED irradiation on proliferation and gene expression of cultured human keratinocytes

    NASA Astrophysics Data System (ADS)

    Becker, Anja; Sticht, Carsten; Dweep, Harsh; van Abeelen, Frank A.; Gretz, Norbert; Oversluizen, Gerrit

    2015-03-01

    Blue light is known for its anti-microbial, anti-proliferative and anti-inflammatory effects. Furthermore, it is already used for the treatment of neonatal jaundice and acne. However, little is known about the exact mechanisms of action on gene expression level. The aim of this study was to assess the impact of blue LED irradiation on the proliferation and gene expression in immortalized human keratinocytes (HaCaT) in vitro. Furthermore its safety was assessed. XTT-tests revealed a decrease in cell proliferation in blue light irradiated cells depending on the duration of light irradiation. Moreover, gene expression analysis demonstrated deregulated genes already 3 hours after blue light irradiation. 24 hours after blue light irradiation the effects seemed to be even more pronounced. The oxidative stress response was significantly increased, pointing to increased ROS production due to blue light, as well as steroid hormone biosynthesis. Downregulated pathways or biological processes were connected to anti-inflammatory response. Interestingly, also the melanoma pathway contained significantly downregulated genes 24 hours after blue light irradiation, which stands in accordance to literature that blue light can also inhibit proliferation in cancer cells. First tests with melanoma cells revealed a decrease in cell proliferation after blue light irradiation. In conclusion, blue light irradiation might open avenues to new therapeutic regimens; at least blue light seems to have no effect that induces cancer growth or formation.

  13. The use of gamma-irradiation and ultraviolet-irradiation in the preparation of human melanoma cells for use in autologous whole-cell vaccines.

    PubMed

    Deacon, Donna H; Hogan, Kevin T; Swanson, Erin M; Chianese-Bullock, Kimberly A; Denlinger, Chadrick E; Czarkowski, Andrea R; Schrecengost, Randy S; Patterson, James W; Teague, Mark W; Slingluff, Craig L

    2008-12-04

    Human cancer vaccines incorporating autologous tumor cells carry a risk of implantation and subsequent metastasis of viable tumor cells into the patient who is being treated. Despite the fact that the melanoma cell preparations used in a recent vaccine trial (Mel37) were gamma-irradiated (200 Gy), approximately 25% of the preparations failed quality control release criteria which required that the irradiated cells incorporate 3H-thymidine at no more than 5% the level seen in the non-irradiated cells. We have, therefore, investigated ultraviolet (UV)-irradiation as a possible adjunct to, or replacement for gamma-irradiation. Melanoma cells were gamma- and/or UV-irradiated. 3H-thymidine uptake was used to assess proliferation of the treated and untreated cells. Caspase-3 activity and DNA fragmentation were measured as indicators of apoptosis. Immunohistochemistry and Western blot analysis was used to assess antigen expression. UV-irradiation, either alone or in combination with gamma-irradiation, proved to be extremely effective in controlling the proliferation of melanoma cells. In contrast to gamma-irradiation, UV-irradiation was also capable of inducing significant levels of apoptosis. UV-irradiation, but not gamma-irradiation, was associated with the loss of tyrosinase expression. Neither form of radiation affected the expression of gp100, MART-1/MelanA, or S100. These results indicate that UV-irradiation may increase the safety of autologous melanoma vaccines, although it may do so at the expense of altering the antigenic profile of the irradiated tumor cells.

  14. Effect of gamma irradiation on the wear behaviour of human tooth enamel

    PubMed Central

    Qing, Ping; Huang, Shengbin; Gao, ShanShan; Qian, LinMao; Yu, HaiYang

    2015-01-01

    Radiotherapy is a frequently used treatment for oral cancer. Extensive research has been conducted to detect the mechanical properties of dental hard tissues after irradiation at the macroscale. However, little is known about the influence of irradiation on the tribological properties of enamel at the micro- or nanoscale. Therefore, this study aimed to investigate the effect of gamma irradiation on the wear behaviour of human tooth enamel in relation to prism orientation. Nanoscratch tests, surface profilometer and scanning electron microscope (SEM) analysis were used to evaluate the friction behaviour of enamel slabs before and after treatment with identical irradiation procedures. X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) were performed to analyse the changes in crystallography and chemical composition induced by irradiation. Surface microhardness (SMH) alteration was also evaluated. The results showed that irradiation resulted in different scratch morphologies, friction coefficients and remnant depth and width at different loads. An inferior nanoscratch resistance was observed independent of prism orientation. Moreover, the variation of wear behaviours was closely related to changes in the crystallography, chemical composition and SMH of the enamel. Together, these measures indicated that irradiation had a direct deleterious effect on the wear behaviour of human tooth enamel. PMID:26099692

  15. Effect of gamma irradiation on the wear behaviour of human tooth enamel.

    PubMed

    Qing, Ping; Huang, Shengbin; Gao, ShanShan; Qian, LinMao; Yu, HaiYang

    2015-06-23

    Radiotherapy is a frequently used treatment for oral cancer. Extensive research has been conducted to detect the mechanical properties of dental hard tissues after irradiation at the macroscale. However, little is known about the influence of irradiation on the tribological properties of enamel at the micro- or nanoscale. Therefore, this study aimed to investigate the effect of gamma irradiation on the wear behaviour of human tooth enamel in relation to prism orientation. Nanoscratch tests, surface profilometer and scanning electron microscope (SEM) analysis were used to evaluate the friction behaviour of enamel slabs before and after treatment with identical irradiation procedures. X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) were performed to analyse the changes in crystallography and chemical composition induced by irradiation. Surface microhardness (SMH) alteration was also evaluated. The results showed that irradiation resulted in different scratch morphologies, friction coefficients and remnant depth and width at different loads. An inferior nanoscratch resistance was observed independent of prism orientation. Moreover, the variation of wear behaviours was closely related to changes in the crystallography, chemical composition and SMH of the enamel. Together, these measures indicated that irradiation had a direct deleterious effect on the wear behaviour of human tooth enamel.

  16. Platelet-Rich Fibrin Lysate Can Ameliorate Dysfunction of Chronically UVA-Irradiated Human Dermal Fibroblasts.

    PubMed

    Wirohadidjojo, Yohanes Widodo; Budiyanto, Arief; Soebono, Hardyanto

    2016-09-01

    To determine whether platelet-rich fibrin lysate (PRF-L) could restore the function of chronically ultraviolet-A (UVA)-irradiated human dermal fibroblasts (HDFs), we isolated and sub-cultured HDFs from six different human foreskins. HDFs were divided into two groups: those that received chronic UVA irradiation (total dosages of 10 J cm⁻²) and those that were not irradiated. We compared the proliferation rates, collagen deposition, and migration rates between the groups and between chronically UVA-irradiated HDFs in control and PRF-L-treated media. Our experiment showed that chronic UVA irradiation significantly decreased (p<0.05) the proliferation rates, migration rates, and collagen deposition of HDFs, compared to controls. Compared to control media, chronically UVA-irradiated HDFs in 50% PRF-L had significantly increased proliferation rates, migration rates, and collagen deposition (p<0.05), and the migration rates and collagen deposition of chronically UVA-irradiated HDFs in 50% PRF-L were equal to those of normal fibroblasts. Based on this experiment, we concluded that PRF-L is a good candidate material for treating UVA-induced photoaging of skin, although the best method for its clinical application remains to be determined.

  17. Effect of gamma irradiation on the wear behaviour of human tooth enamel

    NASA Astrophysics Data System (ADS)

    Qing, Ping; Huang, Shengbin; Gao, Shanshan; Qian, Linmao; Yu, Haiyang

    2015-06-01

    Radiotherapy is a frequently used treatment for oral cancer. Extensive research has been conducted to detect the mechanical properties of dental hard tissues after irradiation at the macroscale. However, little is known about the influence of irradiation on the tribological properties of enamel at the micro- or nanoscale. Therefore, this study aimed to investigate the effect of gamma irradiation on the wear behaviour of human tooth enamel in relation to prism orientation. Nanoscratch tests, surface profilometer and scanning electron microscope (SEM) analysis were used to evaluate the friction behaviour of enamel slabs before and after treatment with identical irradiation procedures. X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) were performed to analyse the changes in crystallography and chemical composition induced by irradiation. Surface microhardness (SMH) alteration was also evaluated. The results showed that irradiation resulted in different scratch morphologies, friction coefficients and remnant depth and width at different loads. An inferior nanoscratch resistance was observed independent of prism orientation. Moreover, the variation of wear behaviours was closely related to changes in the crystallography, chemical composition and SMH of the enamel. Together, these measures indicated that irradiation had a direct deleterious effect on the wear behaviour of human tooth enamel.

  18. Platelet-Rich Fibrin Lysate Can Ameliorate Dysfunction of Chronically UVA-Irradiated Human Dermal Fibroblasts

    PubMed Central

    Budiyanto, Arief; Soebono, Hardyanto

    2016-01-01

    To determine whether platelet-rich fibrin lysate (PRF-L) could restore the function of chronically ultraviolet-A (UVA)-irradiated human dermal fibroblasts (HDFs), we isolated and sub-cultured HDFs from six different human foreskins. HDFs were divided into two groups: those that received chronic UVA irradiation (total dosages of 10 J cm-2) and those that were not irradiated. We compared the proliferation rates, collagen deposition, and migration rates between the groups and between chronically UVA-irradiated HDFs in control and PRF-L-treated media. Our experiment showed that chronic UVA irradiation significantly decreased (p<0.05) the proliferation rates, migration rates, and collagen deposition of HDFs, compared to controls. Compared to control media, chronically UVA-irradiated HDFs in 50% PRF-L had significantly increased proliferation rates, migration rates, and collagen deposition (p<0.05), and the migration rates and collagen deposition of chronically UVA-irradiated HDFs in 50% PRF-L were equal to those of normal fibroblasts. Based on this experiment, we concluded that PRF-L is a good candidate material for treating UVA-induced photoaging of skin, although the best method for its clinical application remains to be determined. PMID:27401663

  19. Influence of UV irradiation on the composition of human stratum corneum lipids

    SciTech Connect

    Wefers, H.; Melnik, B.C.; Fluer, M.B.; Bluhm, C.; Lehmann, P.; Plewig, G. )

    1991-06-01

    Irradiation with suberythemal doses of either UV-A or UV-B yielded an increase in the amount of stratum corneum lipids extracted from the lumbar skin area of 20 volunteers. These lipids were quantified after separation by high-performance thin-layer chromatography. Ten subfractions in the ceramide region were separated; two of them (fractions 7a and 7b) were only detectable after UV-A or UV-B irradiation. Improvement of barrier function after UV irradiation of human skin with suberythemal doses may be related to an increase in the stratum corneum ceramides.

  20. Characterization of a lymphoblastoid line deleted for lambda immunoglobulin genes

    SciTech Connect

    Hough, C.A., White, B.N., Holden, J.A.

    1995-04-01

    While characterizing the cat eye syndrome (CES) supernumerary chromosome for the presence of {lambda} immunoglobulin gene region sequences, a lymphoblastoid cell line from one CES patient was identified in which there was selection of cells deleted from some IGLC and IGLV genes. Two distinct deletions, one on each chromosome 22, were identified, presumably arising from independent somatic recombination events occurring during B-lymphocyte differentiation. The extent of the deleted regions was determined using probes from the various IGLV subgroups and they each covered at least 82 kilobases. The precise definition of the deletions was not possible because of conservation of some restriction sites in the IGLV region. The cell line was used to map putative IGLV genes within the recombinant phage {lambda}V{lambda}135 to the distal part of the IGLV gene region. 35 refs., 4 figs.

  1. Class I major histocompatibility complex-restricted cytotoxic T lymphocytes specific for Epstein-Barr virus (EBV)-transformed B lymphoblastoid cell lines against which they were raised.

    PubMed

    Hill, A B; Lee, S P; Haurum, J S; Murray, N; Yao, Q Y; Rowe, M; Signoret, N; Rickinson, A B; McMichael, A J

    1995-06-01

    We have raised CD8+ cytotoxic T lymphocytes (CTL) from three Epstein-Barr virus-seropositive donors by incubating peripheral blood lymphocytes with irradiated autologous B95.8-strain EBV-transformed B lymphoblastoid cells (LCL). However, to detect lysis in a standard 51Cr release assay of the LCL against which these CTL were raised, superinfection with recombinant vaccinia expressing the appropriate EBV protein or incubation with the peptide epitope was necessary. The untreated LCL were not lysed, even though Western blotting demonstrated that they expressed the EBV antigens containing the CTL epitopes. We have found CTL of this phenotype that are restricted by human leukocyte antigen-A2, -A3, -B7, or -B39, and which recognize the EBV latent proteins, EBV nuclear antigen (EBNA)-3A, EBNA-3C, or terminal protein. During these experiments, we identified a new human leukocyte antigen-A3-restricted EBNA-3A epitope, residues 603-611, RLRAEAGVK. We raised a spontaneous LCL, transformed by endogenous EBV, from one donor, but this was also not lysed. For at least one of the epitopes, CTL from another donor lysed the LCL without superinfection or addition of peptides. We conclude that the CTL were unable to achieve a high enough avidity of interaction with untreated LCL to trigger effector function, although the LCL were able to stimulate them to grow in vitro for up to 4 mo. To assess whether a small percentage of the LCL might possess a higher antigen density, we used an assay of tumor necrosis factor release from a CTL clone, which was able to detect antigen-bearing cells representing only 1% of a stimulating LCL population. Nevertheless, the untreated autologous LCL line failed to stimulate tumor necrosis factor release.

  2. Irradiation affects cellular properties and Eph receptor expression in human melanoma cells

    PubMed Central

    Mosch, Birgit; Pietzsch, Doreen; Pietzsch, Jens

    2012-01-01

    X-ray irradiation influences metastatic properties of tumor cells and, moreover, metastasis and cellular motility can be modified by members of the Eph receptor/ephrin family of receptor tyrosine kinases. We hypothesized that irradiation-induced changes in cellular properties relevant for metastasis in melanoma cells could be mediated by Eph receptor/ephrin signaling. In this pilot study, we analyzed one pre-metastatic (Mel-Juso) and three metastatic human melanoma (Mel-Juso-L3, A375, and A2058) cells lines and predominantly found anti-metastatic effects of X-ray irradiation with impaired cell growth, clonal growth and motility. Additionally, we observed an irradiation-induced increase in adhesion paralleled by a decrease in migration in Mel-Juso and Mel-Juso-L3 cells and, in part, also in A375 cells. We further demonstrate a decrease of EphA2 both in expression and activity at 7 d after irradiation paralleled by an upregulation of EphA3. Analyzing downstream signaling after irradiation, we detected decreased Src kinase phosphorylation, but unchanged focal adhesion kinase (FAK) phosphorylation, indicating, in part, irradiation-induced downregulation of signaling via the EphA2-Src-FAK axis in melanoma cells. However, to which extent this finding contributes to the modification of metastasis-relevant cellular properties remains to be elucidated. PMID:22568947

  3. Low Power Laser Irradiation Stimulates the Proliferation of Adult Human Retinal Pigment Epithelial Cells in Culture

    PubMed Central

    Song, Qing; Uygun, Basak; Banerjee, Ipsita; Nahmias, Yaakov; Zhang, Quan; Berthiaume, François; Latina, Mark; Yarmush, Martin L.

    2015-01-01

    We investigated the effects of low power laser irradiation on the proliferation of retinal pigment epithelial (RPE) cells. Adult human RPE cells were artificially pigmented by preincubation with sepia melanin, and exposed to a single sublethal laser pulse (590 nm, 1 µs, <200 mJ/cm2). DNA synthesis, cell number, and growth factor activity in irradiated RPE cells were subsequently monitored. The effect of sublethal laser irradiation on the “wound” healing response of an RPE monolayer in an in vitro scratch assay was also investigated. Single pulsed laser irradiation increased DNA synthesis in pigmented RPE cells measured 6 h post-treatment. In the scratch assay, laser irradiation increased the rates of cell proliferation and wound closure. Conditioned medium, collected 48 h following laser treatment, increased cell proliferation of unirradiated cells. Irradiation increased RPE cell secretion of platelet-derived growth factor (PDGF)-B chain, and increased mRNA levels of several growth factors and their receptors, including PDGF, transforming growth factor-β1, basic fibroblast growth factor, epidermal growth factor, insulin-like growth factor, as well as heat shock proteins. This demonstrates, for the first time, that low power single pulsed laser irradiation stimulates the proliferation of RPE cells, and upregulates growth factors that are mitogenic for RPE cells. PMID:26740823

  4. Effect of low-power laser irradiation on protein synthesis and ultrastructure of human gingival fibroblasts.

    PubMed

    Marques, Márcia M; Pereira, Aymann N; Fujihara, Neusa A; Nogueira, Fernando N; Eduardo, Carlos P

    2004-01-01

    Low-power lasers improve wound healing. Cell proliferation and protein secretion are important steps of this process. The aim of this study was to analyze both protein synthesis and ultrastructural morphology of human gingival fibroblasts irradiated by a low-power laser. The cell line FMM1 was grown in nutritional deficit. Laser irradiation was carried out with a gallium-aluminum-arsenate (Ga-Al-As) diode laser (904 nm, 120 mW, energy density of 3 J/cm(2)). The protein synthesis analysis and ultrastructural morphology of control (non-irradiated) and irradiated cultures were obtained. There were changes in the structure of cytoplasm organelles of treated cells. The procollagen was not altered by the laser irradiation; however, there were a significant reduction of the amount of protein in the DMEM conditioned by irradiated cells. Low-power laser irradiation causes ultrastructural changes in cultured fibroblasts. We suggest that these alterations may lead to disturbances in the collagen metabolism. Copyright 2004 Wiley-Liss, Inc.

  5. Measurements of solar ultraviolet irradiance with respect to the human body surface

    NASA Astrophysics Data System (ADS)

    Stick, Carsten; Harms, Volker; Pielke, Liane

    1994-07-01

    Solar UV irradiance is measured in Westerland, Germany (54.9 degree(s) N, 8.3 degree(s) E) in the immediate vicinity of the North Sea shoreline. Measurements have been done since July 1993, focussing on the biologically effective UV radiation and the human body geometry. A grid double monochromator radiometer (DM 150, Bentham Instruments Comp., Reading, England) is used to measure the spectral resolution of 1 nm. Weighting the spectral irradiance by the action spectrum for the erythema is more appropriate for determining the biological effectiveness than simply dividing the UV radiation into the UV-A and UV-B wavebands. The erythemal irradiance shows a close relation to the sun angle during the course of a day. The exposure times, calculated from the irradiance and the minimal erythemal doses, suggest that people might underestimate the risk of getting sunburnt before noon. Diffuse radiation scattered from the sky contribute about 70% of the erythemal irradiance at a 45 degree(s) sun angle. A receiver oriented directly to the sun, i.e. 45 degree(s) inclined, receives an additional 30% of the erythemal irradiance measured by a horizontally adjusted cosine response sensor. The relative irradiance of curved surfaces like the skin is determined by UV- B-sensitive paper placed around a cylinder. This device detected UV radiation reflected by the sea, which hardly is measured by horizontally adjusted receivers.

  6. Response of a human colon adenocarcinoma (DLD-1) to x irradiation and mitomycin C in vivo

    SciTech Connect

    Spremulli, E.N.; Leith, J.T.; Bliven, S.F.; Campbell, D.E.; Dexter, D.L.; Glicksman, A.S.; Calabresi, P.

    1983-08-01

    Mice hosting a heterogeneous human colon xenograft tumor produced by subcutaneous injection of the DLD-1 tumor cell line were treated either with x irradiation alone, with mitomycin C alone (4 mg/kg), or with x irradiation given two hours after intraperitoneal injection of mitomycin C (4 mg/kg). Radiation alone produced a dose dependent delay in the time needed for tumors to regrow to twice their size at the time of irradiation, and in the mice receiving mitomycin C plus x irradiation, an additional growth delay equivalent to that produced by 3 to 3.5 Gy of x rays was seen at all x ray dose levels. As the DLD-1 tumor xenografts do not appear to possess a significant hypoxic fraction, we conclude that the two agents are acting in a simple additive cytotoxic manner by the killing of oxic tumor cells.

  7. The selection of light emitting diode irradiation parameters for stimulation of human mesenchymal stem cells proliferation

    NASA Astrophysics Data System (ADS)

    Lewandowski, Rafał; Trafny, ElŻbieta A.; Stepińska, Małgorzata; Gietka, Andrzej; Kotowski, Paweł; Dobrzyńska, Monika; Łapiński, Mariusz P.

    2016-12-01

    Human mesenchymal stem cells (hMSCs) with their vast differentiation potential are very useful for cell-based regenerative medicine. To achieve sufficient numbers of cells for tissue engineering, many different methods have been used to reach the effective increase of cell proliferation. Low-energy red light provided by light emitting diodes (LEDs) have been recently introduced as a method that promoted biomodulation and proliferation of hMSCs in vitro. The purpose of this study was to find the optimum stimulatory dosimetric parameters of LED (630 nm) irradiation on the hMSCs proliferation. The energy density was 2, 3, 4, 10, 20 J/cm2 and the power density used was 7, 17 or 30 mW/cm2. Human MSCs were irradiated with single or triple exposures daily at room temperature and the cell proliferation rate was evaluated during nine days after irradiation. The results showed that after irradiation 4 J/cm2 and 17 mW/cm2 at a single dose the proliferation rate of hMSCs increased on day 5 and 9 (13% and 7%, respectively) when compared to nonirradiated cells. However, triple LED irradiation under the same parameters resulted in the decline in the cell proliferation rate on day 5, but the proliferation rate was at the same level on day 9, when compared with the cell proliferation after irradiation with a single dose. The effect of a single dose irradiation with 4 J/cm2 and 17 mW/cm2 on the proliferation of cells was the highest when the cells were irradiated in phosphate-buffered saline (PBS) instead of MSCGM culture medium.

  8. Long-term cognitive effects of human stem cell transplantation in the irradiated brain.

    PubMed

    Acharya, Munjal M; Martirosian, Vahan; Christie, Lori-Ann; Limoli, Charles L

    2014-09-01

    Radiotherapy remains a primary treatment modality for the majority of central nervous system tumors, but frequently leads to debilitating cognitive dysfunction. Given the absence of satisfactory solutions to this serious problem, we have used human stem cell therapies to ameliorate radiation-induced cognitive impairment. Here, past studies have been extended to determine whether engrafted cells provide even longer-term benefits to cognition. Athymic nude rats were cranially irradiated (10 Gy) and subjected to intrahippocampal transplantation surgery 2 days later. Human embryonic stem cells (hESC) or human neural stem cells (hNSC) were transplanted, and animals were subjected to cognitive testing on a novel place recognition task 8 months later. Grafting of hNSC was found to provide long lasting cognitive benefits over an 8-month post-irradiation interval. At this protracted time, hNSC grafting improved behavioral performance on a novel place recognition task compared to irradiated animals not receiving stem cells. Engrafted hESC previously shown to be beneficial following a similar task, 1 and 4 months after irradiation, were not found to provide cognitive benefits at 8 months. Our findings suggest that hNSC transplantation promotes the long-term recovery of the irradiated brain, where intrahippocampal stem cell grafting helps to preserve cognitive function.

  9. Total lymphatic irradiation and bone marrow in human heart transplantation

    SciTech Connect

    Kahn, D.R.; Hong, R.; Greenberg, A.J.; Gilbert, E.F.; Dacumos, G.C.; Dufek, J.H.

    1984-08-01

    Six patients, aged 36 to 59 years, had heart transplants for terminal myocardial disease using total lymphatic irradiation (TLI) and donor bone marrow in addition to conventional therapy. All patients were poor candidates for transplantation because of marked pulmonary hypertension, unacceptable tissue matching, or age. Two patients are living and well more than four years after the transplants. Two patients died of infection at six and seven weeks with normal hearts. One patient, whose preoperative pulmonary hypertension was too great for an orthotopic heart transplant, died at 10 days after such a procedure. The other patient died of chronic rejection seven months postoperatively. Donor-specific tolerance developed in 2 patients. TLI and donor bone marrow can produce specific tolerance to donor antigens and allow easy control of rejection, but infection is still a major problem. We describe a new technique of administering TLI with early reduction of prednisone that may help this problem.

  10. Effects of X-ray irradiation on human spermatogenesis

    NASA Technical Reports Server (NTRS)

    Thorslund, T. W.; Paulsen, C. A.

    1972-01-01

    Direct cell kill and inhibition of mitosis have been suggested as mechanisms to explain the occurrence of absolute sterility following the irradiation of the testes. In order to obtain information on the existence and dose dependency of the mechanisms for man, a controlled study was initiated. Sixty-four men received a single midorgan dose to both of their testes ranging from 7.5 to 400r (f = .95). It was deduced from resulting pre-sterile period and sterile period data that both cell kill and mitosis halting mechanisms were operating. The maximum observed sterile period was 501 days with eventual recovery observed in each individual where the follow-up was complete. Thus man appears to be highly radiosensitive in regard to temporary sterility but quite radioresistant in regard to permanent sterility.

  11. Ultraviolet-C Irradiation: A Novel Pasteurization Method for Donor Human Milk

    PubMed Central

    Christen, Lukas; Lai, Ching Tat; Hartmann, Ben; Hartmann, Peter E.; Geddes, Donna T.

    2013-01-01

    Background Holder pasteurization (milk held at 62.5°C for 30 minutes) is the standard treatment method for donor human milk. Although this method of pasteurization is able to inactivate most bacteria, it also inactivates important bioactive components. Therefore, the objective of this study was to investigate ultraviolet irradiation as an alternative treatment method for donor human milk. Methods Human milk samples were inoculated with five species of bacteria and then UV-C irradiated. Untreated and treated samples were analysed for bacterial content, bile salt stimulated lipase (BSSL) activity, alkaline phosphatase (ALP) activity, and fatty acid profile. Results All five species of bacteria reacted similarly to UV-C irradiation, with higher dosages being required with increasing concentrations of total solids in the human milk sample. The decimal reduction dosage was 289±17 and 945±164 J/l for total solids of 107 and 146 g/l, respectively. No significant changes in the fatty acid profile, BSSL activity or ALP activity were observed up to the dosage required for a 5-log10 reduction of the five species of bacteria. Conclusion UV-C irradiation is capable of reducing vegetative bacteria in human milk to the requirements of milk bank guidelines with no loss of BSSL and ALP activity and no change of FA. PMID:23840820

  12. Solar ultraviolet irradiation induces decorin degradation in human skin likely via neutrophil elastase.

    PubMed

    Li, Yong; Xia, Wei; Liu, Ying; Remmer, Henriette A; Voorhees, John; Fisher, Gary J

    2013-01-01

    Exposure of human skin to solar ultraviolet (UV) irradiation induces matrix metalloproteinase-1 (MMP-1) activity, which degrades type I collagen fibrils. Type I collagen is the most abundant protein in skin and constitutes the majority of skin connective tissue (dermis). Degradation of collagen fibrils impairs the structure and function of skin that characterize skin aging. Decorin is the predominant proteoglycan in human dermis. In model systems, decorin binds to and protects type I collagen fibrils from proteolytic degradation by enzymes such as MMP-1. Little is known regarding alterations of decorin in response to UV irradiation. We found that solar-simulated UV irradiation of human skin in vivo stimulated substantial decorin degradation, with kinetics similar to infiltration of polymorphonuclear (PMN) cells. Proteases that were released from isolated PMN cells degraded decorin in vitro. A highly selective inhibitor of neutrophil elastase blocked decorin breakdown by proteases released from PMN cells. Furthermore, purified neutrophil elastase cleaved decorin in vitro and generated fragments with similar molecular weights as those resulting from protease activity released from PMN cells, and as observed in UV-irradiated human skin. Cleavage of decorin by neutrophil elastase significantly augmented fragmentation of type I collagen fibrils by MMP-1. Taken together, these data indicate that PMN cell proteases, especially neutrophil elastase, degrade decorin, and this degradation renders collagen fibrils more susceptible to MMP-1 cleavage. These data identify decorin degradation and neutrophil elastase as potential therapeutic targets for mitigating sun exposure-induced collagen fibril degradation in human skin.

  13. Examining the influence of ultraviolet C irradiation on recombinant human γD-crystallin

    PubMed Central

    Wen, Wen-Sing

    2010-01-01

    Purpose Human γD crystallin is a principal protein component of the human eye lens and associated with the development of juvenile and mature-onset cataracts. Exposure to ultraviolet (UV) light is thought to perturb protein structure and eventually lead to aggregation. This work is aimed at exploring the effects of UV-C irradiation on recombinant human γD-crystallin (HGDC). Methods Recombinant HGDC proteins were expressed in E. coli strain BL21(DE3) harboring plasmid pEHisHGDC and purified using chromatographic methods. The proteins were then exposed to UV-C light (λmax=254 nm, 15 W) at the intensity of 420, 800, or 1850 μW/cm2. The UV-C-unexposed, supernatant fraction of UV-C-exposed, and re-dissolved precipitated fraction of UV-C exposed preparations were characterized by SDS–PAGE, turbidity measurement, CD spectroscopy, tryptophan fluorescence spectroscopy, acrylamide fluorescence quenching analysis, and sulfhydryl group measurements. Results The turbidity of the HGDC sample solution was found to be positively correlated with HGDC concentration, UV-C irradiation intensity, and UV-C irradiation duration. When exposed to UV-C, HGDC sample solutions became visibly turbid and a noticeable amount of larger protein particle, perceptible to the naked eye, was observed upon prolonged irradiation. The precipitated fraction of irradiated HGDC sample was found to be re-dissolved by guanidine hydrochloride. Electrophoresis, acrylamide fluorescence quenching, and spectroscopic analyses revealed differences in structures among the non-irradiated HGDC, the supernatant fraction of irradiated HGDC, and the re-dissolved precipitated fraction of irradiated HGDC. Through the use of L-cysteine, the measurements of sulfhydryl contents, and the reducing as well as non-reducing SDS–PAGE, our data further suggested that disulfide bond formation and/or cleavage probably play an important role in aggregation and/or precipitation of HGDC elicited by UV-C irradiation. Conclusions

  14. UV irradiance on the human skin: Effects of orientation and sky obstructions

    NASA Astrophysics Data System (ADS)

    Koepke, Peter; Hess, Michael; Bretl, Sebastian; Seefeldner, Meinhard

    2009-03-01

    Modification factors (MF) are presented that allow the transfer of the UV index (UVI) into actual values of the UV irradiance on the human skin. The UVI is the general information on solar UV irradiance and valid for a horizontal surface under a sky without obstructions. The human skin, however, may be tilted and present in an environment whereby the sun or sky is obstructed, such as within a street canyon, or under a sunshade or trees. These MFs are nearly independent of atmospheric conditions and thus can be used to determine the UV irradiances that are vital for sun burn, skin cancer, and vitamin D production, from the readily available actual UVI, which vary with the atmospheric conditions.

  15. Modeling the biological response of normal human cells, including repair processes, to fractionated carbon beam irradiation

    PubMed Central

    Wada, Mami; Suzuki, Masao; Liu, Cuihua; Kaneko, Yumiko; Fukuda, Shigekazu; Ando, Koichi; Matsufuji, Naruhiro

    2013-01-01

    To understand the biological response of normal cells to fractionated carbon beam irradiation, the effects of potentially lethal damage repair (PLDR) and sublethal damage repair (SLDR) were both taken into account in a linear-quadratic (LQ) model. The model was verified by the results of a fractionated cell survival experiment with normal human fibroblast cells. Cells were irradiated with 200-kV X-rays and monoenergetic carbon ion beams (290 MeV/u) at two irradiation depths, corresponding to linear energy transfers (LETs) of approximately 13 keV/μm and 75 keV/μm, respectively, at the Heavy Ion Medical Accelerator in Chiba of the National Institute of Radiological Sciences. When we only took into account the repair factor of PLDR, γ, which was derived from the delayed assay, the cell survival response to fractionated carbon ion irradiation was not fully explained in some cases. When both the effects of SLDR and PLDR were taken into account in the LQ model, the cell survival response was well reproduced. The model analysis suggested that PLDR occurs in any type of radiation. The γ factors ranged from 0.36–0.93. In addition, SLD was perfectly repaired during the fraction interval for the lower LET irradiations but remained at about 30% for the high-LET irradiation. PMID:23449640

  16. Whole-body irradiation transiently diminishes the adrenocorticotropin response to recombinant human interleukin-1{alpha}

    SciTech Connect

    Perlstein, R.S.; Mehta, N.R.; Neta, R.; Whitnall, M.H.; Mougey, E.H.

    1995-03-01

    Recombinant human interleukin-1{alpha} (rhIL-1{alpha}) has significant potential as a radioprotector and/or treatment for radiation-induced hematopoietic injury. Both IL-1 and whole-body ionizing irradiation acutely stimulate the hypothalamic-pituitary-adrenal axis. We therefore assessed the interaction of whole-body irradiation and rhIL-1{alpha} in altering the functioning of the axis in mice. Specifically, we determined the adrenocorticotropin (ACTH) and corticosterone responses to rhIL-1{alpha} administered just before and hours to days after whole-body or sham irradiation. Our results indicate that whole-body irradiation does not potentiate the rhIL-1{alpha}-induced increase in ACTH levels at the doses used. In fact, the rhIL-1{alpha}-induced increase in plasma ACTH is transiently impaired when the cytokine is administered 5 h after, but not 1 h before, exposure to whole-body irradiation. The ACTH response may be inhibited by elevated corticosterone levels after whole-body irradiation, or by other radiation-induced effects on the pituitary gland and hypothalamus. 36 refs., 3 figs.

  17. Gelam honey protects against gamma-irradiation damage to antioxidant enzymes in human diploid fibroblasts.

    PubMed

    Ahmad, Tengku Ahbrizal Farizal Tengku; Jubri, Zakiah; Rajab, Nor Fadilah; Rahim, Khairuddin Abdul; Yusof, Yasmin Anum Mohd; Makpol, Suzana

    2013-02-11

    The present study was designed to determine the radioprotective effects of Malaysian Gelam honey on gene expression and enzyme activity of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) of human diploid fibroblasts (HDFs) subjected to gamma-irradiation. Six groups of HDFs were studied: untreated control, irradiated HDFs, Gelam honey-treated HDFs and HDF treated with Gelam honey pre-, during- and post-irradiation. HDFs were treated with 6 mg/mL of sterilized Gelam honey (w/v) for 24 h and exposed to 1 Gray (Gy) of gamma rays at the dose rate of 0.25 Gy/min. Gamma-irradiation was shown to down-regulate SOD1, SOD2, CAT and GPx1 gene expressions (p < 0.05). Conversely, HDFs treated with Gelam honey alone showed up-regulation of all genes studied. Similarly, SOD, CAT and GPx enzyme activities in HDFs decreased with gamma-irradiation and increased when cells were treated with Gelam honey (p < 0.05). Furthermore, of the three different stages of study treatment, pre-treatment with Gelam honey caused up-regulation of SOD1, SOD2 and CAT genes expression and increased the activity of SOD and CAT. As a conclusion, Gelam honey modulates the expression of antioxidant enzymes at gene and protein levels in irradiated HDFs indicating its potential as a radioprotectant agent.

  18. Epidermal changes in human skin following irradiation with either UVB or UVA

    SciTech Connect

    Pearse, A.D.; Gaskell, S.A.; Marks, R.

    1987-01-01

    We have demonstrated previously that following UVB irradiation to normal volunteers there is an increase in epidermal and stratum corneum thickness and an increase in the thymidine autoradiographic labeling index. These changes are coupled with alterations in epidermal glucose-6-phosphate dehydrogenase and succinic dehydrogenase activities, despite the absence of erythema clinically. The use of a sunscreen did not completely prevent these changes. In this study, we have examined the effects of repeated irradiation of human skin with either UVB or UVA alone in order to compare the changes produced in the epidermis and to ascertain whether UVA irradiation could cause these. Irradiation with either UVB or UVA alone was found to increase the mean epidermal thickness, the mean stratum corneum thickness, and mean keratinocyte height significantly. Glucose-6-phosphate dehydrogenase activity was significantly increased throughout the epidermis, and succinic dehydrogenase activity was significantly decreased. The autoradiographic labeling index was significantly increased following UVB irradiation but not following UVA irradiation. These results demonstrate that UVA alone can have a direct effect on epidermal morphology and metabolism, suggesting that protection of skin from UV radiation should include adequate protection from UVA.

  19. Overview of Radiosensitivity of Human Tumor Cells to Low-Dose-Rate Irradiation

    SciTech Connect

    Williams, Jerry R. Zhang Yonggang; Zhou Haoming; Gridley, Daila S.; Koch, Cameron J.; Slater, James M.; Little, John B.

    2008-11-01

    Purpose: We compared clonogenic survival in 27 human tumor cell lines that vary in genotype after low-dose-rate (LDR) or high-dose rate (HDR) irradiation. We measured susceptibility to LDR-induced redistribution in the cell cycle in eight of these cell lines. Methods and Materials: We measured clonogenic survival after up to 96 hours of LDR (0.25 Gy/h) irradiation. We compared these with clonogenic survival after HDR irradiation (50 Gy/h). Using flow cytometry, we measured LDR-induced redistribution as a function of time during LDR irradiation in eight of these cell lines. Results: Coefficients that describe clonogenic survival after both LDR and HDR irradiation segregate into four radiosensitivity groups that associate with cell genotype: mutant (mut)ATM, wild-type TP53, mutTP53, and an unidentified gene in radioresistant glioma cells. The LDR and HDR radiosensitivity correlates at lower doses ({approx}2 Gy HDR, {approx}6 Gy LDR), but not at higher doses (HDR > 4 Gy; LDR > 6 Gy). The rate of LDR-induced loss of clonogenic survival changes at approximately 24 hours; wild-type TP53 cells become more resistant and mutTP53 cells become more sensitive. Redistribution induced by LDR irradiation also changes at approximately 24 hours. Conclusions: Radiosensitivity of human tumor cells to both LDR and HDR irradiation is genotype dependent. Analysis of coefficients that describe cellular radiosensitivity segregates 27 cell lines into four statistically distinct groups, each associating with specific genotypes. Changes in cellular radiosensitivity and redistribution in the cell cycle are strongly time dependent. Our data establish a genotype-dependent time-dependent model that predicts clonogenic survival, explains the inverse dose-rate effect, and suggests possible clinical applications.

  20. Dose Calculation Evolution for Internal Organ Irradiation in Humans

    SciTech Connect

    Jimenez V, Reina A.

    2007-10-26

    The International Commission of Radiation Units (ICRU) has established through the years, a discrimination system regarding the security levels on the prescription and administration of doses in radiation treatments (Radiotherapy, Brach therapy, Nuclear Medicine). The first level is concerned with the prescription and posterior assurance of dose administration to a point of interest (POI), commonly located at the geometrical center of the region to be treated. In this, the effects of radiation around that POI, is not a priority. The second level refers to the dose specifications in a particular plane inside the patient, mostly the middle plane of the lesion. The dose is calculated to all the structures in that plane regardless if they are tumor or healthy tissue. In this case, the dose is not represented by a point value, but by level curves called 'isodoses' as in a topographic map, so you can assure the level of doses to this particular plane, but it also leave with no information about how this values go thru adjacent planes. This is why the third level is referred to the volumetrical description of doses so these isodoses construct now a volume (named 'cloud') that give us better assurance about tissue irradiation around the volume of the lesion and its margin (sub clinical spread or microscopic illness). This work shows how this evolution has resulted, not only in healthy tissue protection improvement but in a rise of tumor control, quality of life, better treatment tolerance and minimum permanent secuelae.

  1. Dose-Dependent Metabolic Alterations in Human Cells Exposed to Gamma Irradiation

    PubMed Central

    Kwon, Yong-Kook; Ha, In Jin; Bae, Hyun-Whee; Jang, Won Gyo; Yun, Hyun Jin; Kim, So Ra; Lee, Eun Kyeong; Kang, Chang-Mo; Hwang, Geum-Sook

    2014-01-01

    Radiation exposure is a threat to public health because it causes many diseases, such as cancers and birth defects, due to genetic modification of cells. Compared with the past, a greater number of people are more frequently exposed to higher levels of radioactivity today, not least due to the increased use of diagnostic and therapeutic radiation-emitting devices. In this study, ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS)-based metabolic profiling was used to investigate radiation- induced metabolic changes in human fibroblasts. After exposure to 1 and 5 Gy of γ-radiation, the irradiated fibroblasts were harvested at 24, 48, and 72 h and subjected to global metabolite profiling analysis. Mass spectral peaks of cell extracts were analyzed by pattern recognition using principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA). The results showed that the cells irradiated with 1 Gy returned to control levels at 72 h post radiation, whereas cells irradiated with 5 Gy were quite unlike the controls; therefore, cells irradiated with 1 Gy had recovered, whereas those irradiated with 5 Gy had not. Lipid and amino acid levels increased after the higher-level radiation, indicating degradation of membranes and proteins. These results suggest that MS-based metabolite profiling of γ-radiation-exposed human cells provides insight into the global metabolic alterations in these cells. PMID:25419661

  2. Anti-angiogenic activity in metastasis of human breast cancer cells irradiated by a proton beam

    NASA Astrophysics Data System (ADS)

    Lee, Kyu-Shik; Shin, Jin-Sun; Nam, Kyung-Soo; Shon, Yun-Hee

    2012-07-01

    Angiogenesis is an essential process of metastasis in human breast cancer. We investigated the effects of proton beam irradiation on angiogenic enzyme activities and their expressions in MCF-7 human breast cancer cells. The regulation of angiogenic regulating factors, of transforming growth factor- β (TGF- β) and of vesicular endothelial growth factor (VEGF) expression in breast cancer cells irradiated with a proton beam was studied. Aromatase activity and mRNA expression, which is correlated with metastasis, were significantly decreased by irradiation with a proton beam in a dose-dependent manner. TGF- β and VEGF transcriptions were also diminished by proton beam irradiation. In contrast, transcription of tissue inhibitors of matrix metalloproteinases (TIMPs), also known as biological inhibitors of matrix metalloproteinases (MMPs), was dose-dependently enhanced. Furthermore, an increase in the expression of TIMPs caused th MMP-9 activity to be diminished and the MMP-9 and the MMP-2 expressions to be decreased. These results suggest that inhibition of angiogenesis by proton beam irradiation in breast cancer cells is closely related to inhibitions of aromatase activity and transcription and to down-regulation of TGF- β and VEGF transcription.

  3. Factors influencing the removal of thymine glycol from DNA in gamma-irradiated human cells.

    PubMed

    Weinfeld, M; Xing, J Z; Lee, J; Leadon, S A; Cooper, P K; Le, X C

    2001-01-01

    The toxic and mutagenic effects of ionizing radiation are believed to be caused by damage to cellular DNA. We have made use of a novel immunoassay for thymine glycol to examine the removal of this lesion from the DNA of irradiated human cells. Because of the sensitivity of the assay, we have been able to keep the radiation doses at or below the standard clinical dose of 2 Gy. Our initial observations indicated that although removal of thymine glycol is > 80% complete by 4 h post-irradiation with 2 Gy, there is a lag of 30-60 min before repair commences. However, if cells are irradiated with 0.25 Gy 4 h prior to the 2-Gy dose, removal of the thymine glycols commences immediately after the second irradiation, suggesting that repair of thymine glycol is inducible. Our current studies are directed at two aspects of the repair process, (1) factors involved in the repair process leading up to and including glycosylase-mediated removal of thymine glycol and (2) the control of the inducible response. We have observed that mutation of the XPG gene drastically reduced the level and rate of global removal of thymine glycol (induced by 2-Gy irradiation), and there was no evidence for an inducible response. Similar results were seen with a Cockayne syndrome B (CSB) cell line. We have also examined repair in quiescent and phytohemagglutinin-stimulated human lymphocytes. Both show similar kinetics for the rate of removal of thymine glycol under induced and noninduced conditions.

  4. Recruitment of Phosphorylated Chromatin Assembly Factor 1 to Chromatin after UV Irradiation of Human Cells

    PubMed Central

    Martini, Emmanuelle; Roche, Danièle M.J.; Marheineke, Kathrin; Verreault, Alain; Almouzni, Geneviève

    1998-01-01

    The subcellular distribution and posttranslational modification of human chromatin assembly factor 1 (CAF-1) have been investigated after UV irradiation of HeLa cells. In an asynchronous cell population only a subfraction of the two large CAF-1 subunits, p150 and p60, were found to exist in a chromatin-associated fraction. This fraction is most abundant during S phase in nonirradiated cells and is much reduced in G2 cells. After UV irradiation, the chromatin-associated form of CAF-1 dramatically increased in all cells irrespective of their position in the cell cycle. Such chromatin recruitment resembles that seen for PCNA, a DNA replication and repair factor. The chromatin-associated fraction of p60 was predominantly hypophosphorylated in nonirradiated G2 cells. UV irradiation resulted in the rapid recruitment to chromatin of phosphorylated forms of the p60 subunit. Furthermore, the amount of the p60 and p150 subunits of CAF-1 associated with chromatin was a function of the dose of UV irradiation. Consistent with these in vivo observations, we found that the amount of CAF-1 required to stimulate nucleosome assembly during the repair of UV photoproducts in vitro depended upon both the number of lesions and the phosphorylation state of CAF-1. The recruitment of CAF-1 to chromatin in response to UV irradiation of human cells described here supports a physiological role for CAF-1 in linking chromatin assembly to DNA repair. PMID:9813080

  5. Effect of duration and intensity of ganciclovir exposure on lymphoblastoid cell toxicity.

    PubMed

    Janoly-Dumenil, Audrey; Rouvet, Isabelle; Bleyzac, Nathalie; Bertrand, Yves; Aulagner, Gilles; Zabot, Marie-Thérèse

    2009-01-01

    Human cytomegalovirus infection is still a major complication after pediatric bone marrow transplantation and could be fatal in some cases. The toxicity of the drug in dividing transplanted haematopoietic cells combined with the suppression of cell growth caused by the virus remains a major problem in managing human cytomegalovirus infection. The aim of the current in vitro study was to evaluate the effect of the intensity (1-20 mg/l) and duration (1, 2, 7 or 14 days) of ganciclovir exposure on toxicity in B lymphoblastoid cells (using cell counting and viability measurements). A correlation was found between the dose of ganciclovir exposure and a decrease in total cell number when duration exceeded 2 days (r(2)=0.92 and 0.93 after 7 and 14 days, respectively). High levels (20 mg/l) of ganciclovir were not more toxic than lowest levels (1 mg/l) for the shortest durations of ganciclovir exposure (1 and 2 days). Moreover, 50% cytotoxic concentrations markedly decreased with the duration of ganciclovir exposure (374-3 mg/l from 1 to 14 days respectively) after 14 days of culture. This in vitro study demonstrated for the first time that ganciclovir exhibited an in vitro duration-dependent toxicity on haematopoietic-derived cells when in vivo doses of the drug were used.

  6. Time-resolved fluorimetric probing of DNA structure in irradiated human lymphocytes

    NASA Astrophysics Data System (ADS)

    Maves, Shelley R.; Greenstock, Clive L.

    2005-02-01

    An in situ technique has been developed that detects genomic conformational changes in irradiated human cells. Cells are treated on ice with detergent, mild alkali and ethidium bromide (EB) and the resulting intact nuclei are examined using kinetic spectrofluorimetry. In the nuclei of unirradiated lymphocytes the fluorescence decay profile is tri-exponential with a long-lived component (˜23 ns) attributable to EB intercalated within double-stranded DNA, an intermediate life-time component (˜6 ns) indicative of a loosely bound DNA biomolecular-EB complex, and a short-lived component (˜2 ns) corresponding to unbound EB. Irradiated fresh human lymphocytes show three similar components but their relative contributions are changed. Results from a typical donor, show that after 1 Gy the intermediate component decreased with a concomitant increase in the long-lived component while the short-lived component remained essentially unchanged. Fresh whole blood from healthy donors was irradiated at doses of 0.1-1 Gy, and the samples analyzed with or without post-irradiation incubation at 37 °C for 24 h prior to lymphocyte extraction. For doses of 1.0 Gy in the absence of incubation there is good agreement between multiple samples of the same individual, or among the six donors, as compared with the results from irradiated isolated lymphocytes. Whole blood incubation was unreliable but results from one individual at 0.1 and 1.0 Gy were similar to those observed without incubation. Fluorescence lifetime analysis can detect DNA structural/topological damage in irradiated human lymphoid cells, and it may have potential application to in vivo bio-dosimetry and bio-monitoring.

  7. The effects of lipid A on gamma-irradiated human peripheral blood lymphocytes in vitro

    NASA Astrophysics Data System (ADS)

    Dubničková, M.; Kuzmina, E. A.; Chausov, V. N.; Ravnachka, I.; Boreyko, A. V.; Krasavin, E. A.

    2016-03-01

    The modulatory effects of lipid A (diphosphoryl lipid A (DLA) and monophosphoryl lipid A (MLA)) on apoptosis induction and DNA structure damage (single and double-strand breaks (SSBs and DSBs, respectively)) in peripheral human blood lymphocytes are studied for 60Co gamma-irradiation. It is shown that in the presence of these agents the amount of apoptotic cells increases compared with the irradiated control samples. The effect is most strongly pronounced for DLA. In its presence, a significant increase is observed in the number of radiation-induced DNA SSBs and DSBs. Possible mechanisms are discussed of the modifying influence of the used agents on radiation-induced cell reactions

  8. High fluence laser irradiation induces reactive oxygen species generation in human lung adenocarcinoma cells

    NASA Astrophysics Data System (ADS)

    Wang, Fang; Xing, Da; Chen, Tong-Sheng

    2006-09-01

    Low-power laser irradiation (LPLI) has been used for therapies such as curing spinal cord injury, healing wound et al. Yet, the mechanism of LPLI remains unclear. Our previous study showed that low fluences laser irradiation induces human lung adenocarcinoma cells (ASTC-a-1) proliferation, but high fluences induced apoptosis and caspase-3 activation. In order to study the mechanism of apoptosis induced by high fluences LPLI further, we have measured the dynamics of generation of reactive oxygen species (ROS) using H IIDCFDA fluorescence probes during this process. ASTC-a-1 cells apoptosis was induced by He-Ne laser irradiation at high fluence of 120J/cm2. A confocal laser scanning microscope was used to perform fluorescence imaging. The results demonstrated that high fluence LPLI induced the increase of mitochondria ROS. Our studies contribute to clarify the biological mechanism of high fluence LPLI-induced cell apoptosis.

  9. Transformation of human diploid cells by adenovirus type 4 irradiated with ultraviolet light.

    PubMed

    Hozoc, M; Nastac, E; Suru, M; Stoian, M; Bercovici, S; Cajal, N

    1983-01-01

    Inoculation of ultraviolet (UV)-irradiated adenovirus type 4 (Ad4) led to in vitro transformation of human diploid cells (HDC). Two transformed cell lines could be established: cell line H 1418, from HDC inoculated with the 10(-3) dilution of Ad4 UV-irradiated for 20 min at a distance of 20 cm, co-cultivated with uninfected HDC, and cell line H 1557, from HDC inoculated with the 10(-2) dilution of Ad4 irradiated at the same distance for 12 min. Both transformed cell lines were resistant to superinfection with homologous virus. Virus-specific antigen could be made evident by the indirect immunofluorescence technique in the nuclei of both H 1418 and H 1557 cells.

  10. Schisandrin B protects against solar irradiation-induced oxidative injury in BJ human fibroblasts.

    PubMed

    Chiu, Po Yee; Lam, Philip Y; Yan, Chung Wai; Ko, Kam Ming

    2011-06-01

    The effects of schisandrin B (Sch B) and its analogs on solar irradiation-induced oxidative injury were examined in BJ human fibroblasts. Sch B and schisandrin C (Sch C) increased cellular reduced glutathione (GSH) level and protected against solar irradiation-induced oxidative injury. The photoprotection was paralleled by decreases in the elastases-type protease activity and matrix-metalloproteinases-1 expression in solar-irradiated fibroblasts. The cytochrome P-450-mediated metabolism of Sch B or Sch C caused ROS production. The results suggest that by virtue of its pro-oxidant action and the subsequent glutathione antioxidant response, Sch B or Sch C may offer the prospect of preventing skin photo-aging.

  11. Inactivation of enveloped and non-enveloped viruses on seeded human tissues by gamma irradiation.

    PubMed

    Moore, Mark A

    2012-08-01

    Human tissue allografts are widely used in a variety of clinical applications with over 1.5 million implants annually in the US alone. Since the 1990s, most clinically available allografts have been disinfected to minimize risk of disease transmission. Additional safety assurance can be provided by terminal sterilization using low dose gamma irradiation. The impact of such irradiation processing at low temperatures on viruses was the subject of this study. In particular, both human tendon and cortical bone samples were seeded with a designed array of viruses and the ability of gamma irradiation to inactivate those viruses was tested. The irradiation exposures for the samples packed in dry ice were 11.6-12.9 kGy for tendon and 11.6-12.3 kGy for bone, respectively. The viruses, virus types, and log reductions on seeded tendon and bone tissue, respectively, were as follows: Human Immunodeficiency Virus (RNA, enveloped), >2.90 and >3.20; Porcine Parvovirus (DNA, non-enveloped), 1.90 and 1.58; Pseudorabies Virus (DNA, enveloped), 3.80 and 3.79; Bovine Viral Diarrhea Virus (RNA, enveloped), 2.57 and 4.56; and Hepatitis A Virus (RNA, non-enveloped), 2.54 and 2.49, respectively. While proper donor screening, aseptic technique, and current disinfection practices all help reduce the risk of viral transmission from human allograft tissues, data presented here indicate that terminal sterilization using a low temperature, low dose gamma irradiation process inactivates both enveloped and non-enveloped viruses containing either DNA or RNA, thus providing additional assurance of safety from viral transmission.

  12. Transplantation of human fetal-derived neural stem cells improves cognitive function following cranial irradiation

    PubMed Central

    Acharya, Munjal M.; Christie, Lori-Ann; Hazel, Thomas G.; Johe, Karl K.; Limoli, Charles L.

    2013-01-01

    Treatment of CNS malignancies typically involves radiotherapy to forestall tumor growth and recurrence following surgical resection. Despite the many benefits of cranial radiotherapy, survivors often suffer from a wide range of debilitating and progressive cognitive deficits. Thus, while patients afflicted with primary and secondary malignancies of the CNS now experience longer local regional control and progression free survival, there remains no clinical recourse for the unintended neurocognitive sequelae associated with their cancer treatments. Multiple mechanisms contribute to disrupted cognition following irradiation, including the depletion of radiosensitive populations of stem and progenitor cells in the hippocampus. We have explored the potential of using intrahippocampal transplantation of human stem cells to ameliorate radiation-induced cognitive dysfunction. Past studies demonstrated the capability of cranially transplanted human embryonic (hESCs) and neural (hNSCs) stem cells to functionally restore cognition in rats 1 and 4-months post-head-only irradiation. The present study employed an FDA-approved fetal-derived human neural stem cell line capable of large scale-up under good manufacturing practice (GMP). Animals receiving cranial transplantation of these cells 1-month following irradiation showed improved hippocampal spatial memory and contextual fear conditioning performance compared to irradiated, sham surgery controls. Significant newly born (doublecortin positive) neurons and a smaller fraction of glial subtypes were observed within and nearby the transplantation core. Engrafted cells migrated and differentiated into neuronal and glial subtypes throughout the CA1 and CA3 subfields of the host hippocampus. These studies expand our prior findings to demonstrate that transplantation of fetal-derived human neural stem cells improves cognitive deficits in irradiated animals, as assessed by two separate cognitive tasks. PMID:23866792

  13. SU-C-204-04: Irradiation of Human Cell Lines Using Various Ions

    SciTech Connect

    Lin, Y; McMahon, S; Kaminuma, T; Held, K; Tessa, C; Rusek, A

    2016-06-15

    Purpose: The purpose of this study is to investigate and quantify the biological effects of ion radiation using several human cell lines. We aim to answer the question of whether carbon ion the most ideal ion species for heavy ion radiotherapy. Methods: The cells were irradiated at different positions along the pristine Bragg peak of several ions with different atomic number. The biological effectiveness was evaluated using the clonogenic cell survival assay. Irradiation of three human lung cancer cell lines and a fibroblast cell line were undertaken using the charged particle beam at the NASA Space Radiation Laboratory at Brookhaven National Lab. Four mono-energetic ion beams (carbon, oxygen, helium and lithium) were used to irradiate the cells. Water or media-filled T25 flasks were lined up along the beam line so that the cell-containing surfaces of the flasks were placed at a specific depth along the pristine Bragg curve. Four depths along the curve, representing entrance point, rising peak, peak and distal fall off, were selected to determine biological effectiveness. Gaf-chromic films were placed between the flasks to monitor the irradiation as soon as it was finished. Results: For all ion radiations, the maximum cell killing effect occurs at either peak or distal fall off, depending on the cell lines. For instance, for the fibroblast cell line AGO1522, RBEs of 1.4, 1.2, 1.4 and 1.9 were observed at the Bragg peak for Helium, Lithium, Carbon and Oxygen ions. Comparing positions, RBEs of 0.9, 1.2, 1.4 and 1.8 were observed for carbon irradiation of AGO-1522 cells positions corresponding to entrance, rising peak, peak and distal fall off. Conclusion: RBE values differ with position in the Bragg peak, ion species and cell line. Ions other than carbon may prove more effective in certain irradiation conditions and may contribute to optimized heavy ion therapy.

  14. Monitoring PAI-1 and VEGF Levels in 6 Human Squamous Cell Carcinoma Xenografts During Fractionated Irradiation

    SciTech Connect

    Bayer, Christine; Kielow, Achim; Schilling, Daniela; Maftei, Constantin-Alin; Zips, Daniel; Yaromina, Ala; Baumann, Michael; Molls, Michael; Multhoff, Gabriele

    2012-11-01

    Purpose: Previous studies have shown that the plasminogen activator inhibitor type-1 (PAI-1) and vascular endothelial growth factor (VEGF) are regulated by hypoxia and irradiation and are involved in neoangiogenesis. The aim of this study was to determine in vivo whether changes in PAI-1 and VEGF during fractionated irradiation could predict for radiation resistance. Methods and Materials: Six xenografted tumor lines from human squamous cell carcinomas (HSCC) of the head and neck were irradiated with 0, 3, 5, 10, and 15 daily fractions of 2 Gy. The PAI-1 and VEGF antigen levels in tumor lysates were determined by enzyme-linked immunosorbent assay kits. The amounts of PAI-1 and VEGF were compared with the dose to cure 50% of tumors (TCD{sub 50}). Colocalization of PAI-1, pimonidazole (hypoxia), CD31 (endothelium), and Hoechst 33342 (perfusion) was examined by immunofluorescence. Results: Human PAI-1 and VEGF (hVEGF) expression levels were induced by fractionated irradiation in UT-SCC-15, UT-SCC-14, and UT-SCC-5 tumors, and mouse VEGF (msVEGF) was induced only in UT-SCC-5 tumors. High hVEGF levels were significantly associated with radiation sensitivity after 5 fractions (P=.021), and high msVEGF levels were significantly associated with radiation resistance after 10 fractions (P=.007). PAI-1 staining was observed in the extracellular matrix, the cytoplasm of fibroblast-like stroma cells, and individual tumor cells at all doses of irradiation. Colocalization studies showed PAI-1 staining close to microvessels. Conclusions: These results indicate that the concentration of tumor-specific and host-specific VEGF during fractionated irradiation could provide considerably divergent information for the outcome of radiation therapy.

  15. Comparing the functional consequences of human stem cell transplantation in the irradiated rat brain.

    PubMed

    Acharya, Munjal M; Christie, Lori-Ann; Lan, Mary L; Limoli, Charles L

    2013-01-01

    Radiotherapy is a frontline treatment for the clinical management of CNS tumors. Although effective in eradicating tumor cells, radiotherapy also depletes neural stem and progenitor cells in the hippocampus that are important for neurogenesis and cognitive function. Consequently, the use of radiation to control primary and metastatic brain tumors often leads to debilitating and progressive cognitive decrements in surviving patients, representing a serious medical condition that, to date, has no satisfactory, long-term solutions. As a result, we have explored the use of stem cells as therapeutic agents to improve cognition after radiotherapy. Our past work has demonstrated the capability of cranially transplanted human embryonic (hESCs) and neural (hNSCs) stem cells to functionally restore cognition in rats 1 and 4 months after head-only irradiation. We have now expanded our cognitive analyses with hESCs and quantified both survival and differentiated fates of engrafted cells at 1 and 4 months after irradiation. Our findings indicate the capability of hESC transplantation to ameliorate radiation-induced cognitive dysfunction 1 month following cranial irradiation, using a hippocampal-dependent novel place recognition task. Irradiated animals not engrafted with stem cells experienced prolonged and significant cognitive dysfunction. Stereological estimates indicated that 35% and 17% of the transplanted hESCs survived at 1 and 4 months postgrafting, respectively. One month after irradiation and grafting, phenotypic analyses revealed that 26% and 31% of the hESCs differentiated into neurons and astrocytes, while at the 4-month time, neuronal and astrocytic differentiation was 7% and 46%, respectively. Comparison between present and past data with hESCs and hNSCs demonstrates equivalent cognitive restoration and a preference of hNSCs to commit to neuronal versus astrocytic lineages over extended engraftment times. Our data demonstrate the functional utility of human stem

  16. Scavenging of hydroxyl radicals generated in human plasma following X-ray irradiation.

    PubMed

    Hosokawa, Yoichiro; Sano, Tomoaki

    2015-11-01

    There are various antioxidant materials that scavenge free radicals in human plasma. It is possible that the radical-scavenging function causes a radiation protective effect in humans. This study estimated the hydroxyl (OH) radical-scavenging activity induced by X-ray irradiation in human plasma. The test subjects included 111 volunteers (75 males and 36 females) ranging from 22 to 35 years old (average, 24.0). OH radicals generated in irradiated human plasma were measured by electron spin resonance (ESR). The relationships between the amount of the OH radical and chemical and biological parameters [total protein, total cholesterol, triglycerides and hepatitis B surface (HBs) antibodies] were estimated in the plasma of the 111 volunteers by a multivariate analysis. The presence of HBs antibodies had the greatest influence on OH radical-scavenging activity. One volunteer who did not have the HBs antibody was given an inoculation of the hepatitis B vaccine. There was a remarkable decrease in the amount of OH radical generated from plasma after the HBs antibody was produced. The results indicate that the HBs antibody is an important factor for the scavenging of OH radicals initiated by X-ray irradiation in the human body.

  17. Effects Of Continuous Argon Laser Irradiation On Canine And Autopsied Human Cardiac Tissue

    NASA Astrophysics Data System (ADS)

    Ben-Shachar, Giora; Sivakoff, Mark; Bernard, Steven L.; Dahms, Beverly B.; Riemenschneider, Thomas A.

    1984-10-01

    In eight human formalin preserved cardiac specimens, various cardiac and vascular obstructions were relieved by argon laser irradiation. Interatrial communication was also produced by a transar'rial approach in a live dog. In-vivo fresh canine cardiac tissues required power density of at feast 80, 90, and 110 watts/cm2 for vaporization of myocardial, vascular and valvular tissues respectively. The fiber tip to tissue distance (effective irradiation distance) for effective vaporization was less than I mm for vascular and valvular tissues and less than 4 mm for myocardium. Light microscopy showed four zones of histological damage common to all tissues - central crater surrounded by layers of charring, vacuolization and coagulation necorsis. Myocardium showed additionally a layer of normal appearing muscle cells (skip area) surrounded by a peripheral coagulation halo. Laser irradiation effects on valvular tissue showed the most lateral extension of coagulation necrosis. It is concluded that palliation and treatment of certain congenital heart defects by laser irradiation is anatomi-cally feasible and may be safe for in vivo application when low power output and short exposure time are used from a very short irradiation distance.

  18. Dose-responses of Stem Cells from Human Exfoliated Teeth to Infrared LED Irradiation.

    PubMed

    Turrioni, Ana Paula Silveira; Montoro, Liege Aldrovandi; Basso, Fernanda Gonçalves; de Almeida, Leopoldina de Fátima Dantas; Costa, Carlos Alberto de Souza; Hebling, Josimeri

    2015-01-01

    Despite several reports regarding tissue regeneration, including pulp repair induced by different light sources, only limited data have been reported concerning the effects of light-emitting diodes (LED) on stem cells from human exfoliated deciduous teeth (SHEDs). The aim of this study was to evaluate the effects of different energy densities of infrared LED on the cell viability, number of cells and mineralized tissue production by SHEDs. SHEDs were obtained from near-exfoliation primary teeth (n=3), seeded in plain DMEM (104 cells/cm2), and irradiated by a LED prototype (LEDTable 850 nm, 40 mW/cm2) delivering 0 (control), 2, 4, 8, 15 or 30 J/cm2 (n=9). Cell viability (MTT assay), cell proliferation (trypan blue assay), and mineralized nodule (MN) formation (alizarin red stain) were assessed 12 and 72 h post-irradiation. Data were subjected to Kruskal-Wallis and Mann-Whitney tests (α=0.05). Cells irradiated with 2 or 4 J/cm2 exhibited higher metabolism at 72 h, and all energy densities provided increase in cell proliferation after 12 h. Regarding MN formation, the best results were observed at 72 h after SHED irradiation with 8 and 15 J/cm2. It was concluded that the cell viability, cell number and MN formation by pulp cells are enhanced after exposure to infrared LED irradiation. Overall, the greatest SHED biostimulation was obtained with 4 and 8 J/cm2.

  19. [Double-strand DNA breaks induction and repair in human blood lymphocytes irradiated with adapting dose].

    PubMed

    Osipov, A N; Lizunova, E Iu; Vorob'eva, N Iu; Pelevina, I I

    2009-01-01

    Using a DNA-comet assay was shown that irradiation of human blood lymphocytes at G1 cell cycle with a low conditioning dose (5 cGy) induces an adaptive response (AR) manifested in reduction of the double-strand DNA (DSB) amount induced by challenging dose at 10 Gy. 24 h after conditioning irradiation (48 h after PHA addition) in cells irradiated at both conditioning and challenging doses a relative DBS amount was approximately 24% less in comparison to versus a control irradiated at challenging dose only. 48 h after adapting irradiation this index increased to approximately 35%, while 72 h after was decreased to approximately 29%. AR observed by us during 72 h after its induction did not accompanied by statistically significant changes in DBS repair enhancing. It is possible to assume that basic role in AR forming in lymphocytes under experimental conditions used by us playing the processes preventing radiation-induced DBS formation (antioxidant defense system activation, chromatin conformation changes ets).

  20. Diarylheptanoids from Alpinia officinarum Cause Distinct but Overlapping Effects on the Translatome of B Lymphoblastoid Cells

    PubMed Central

    Kakegawa, Tomohito; Takase, Saeko; Masubuchi, Eri; Yasukawa, Ken

    2014-01-01

    Diarylheptanoids (AO-0001, AO-0002, and AO-0003) isolated from Alpinia officinarum inhibit proinflammatory mediators and exhibit cytotoxic and antiviral activity. However, the precise mechanisms of action of these diarylheptanoids are unknown as are their effects on expression of specific genes. Here, we used a translatome analysis to investigate the mechanisms and modes of action of these three diarylheptanoids. Polysome-associated messenger RNAs (mRNAs) were prepared from diarylheptanoids-treated and control cells from a human B lymphoblastoid cell line; these mRNA samples were then used for microarray analysis. Microarray Data Analysis Tool version 3.2 was used to analyze the microarray data analysis; this software uses pathway information of the WikiPathways for gene ontology analysis. Each of the diarylheptanoids caused upregulation or downregulation of the same 37 and 286 genes, respectively. Among the 37 upregulated genes, 16 were related to mRNA processing based on the WikiPathways analysis. Our findings provided new insights into the mode of action of diarylheptanoids from A. officinarum. PMID:25254051

  1. Characterization of the microDNA through the response to chemotherapeutics in lymphoblastoid cell lines.

    PubMed

    Mehanna, Pamela; Gagné, Vincent; Lajoie, Mathieu; Spinella, Jean-François; St-Onge, Pascal; Sinnett, Daniel; Brukner, Ivan; Krajinovic, Maja

    2017-01-01

    Recently, a new class of extrachromosomal circular DNA, called microDNA, was identified. They are on average 100 to 400 bp long and are derived from unique non-repetitive genomic regions with high gene density. MicroDNAs are thought to arise from DNA breaks associated with RNA metabolism or replication slippage. Given the paucity of information on this entirely novel phenomenon, we aimed to get an additional insight into microDNA features by performing the microDNA analysis in 20 independent human lymphoblastoid cell lines (LCLs) prior and after treatment with chemotherapeutic drugs. The results showed non-random genesis of microDNA clusters from the active regions of the genome. The size periodicity of 190 bp was observed, which matches DNA fragmentation typical for apoptotic cells. The chemotherapeutic drug-induced apoptosis of LCLs increased both number and size of clusters further suggesting that part of microDNAs could result from the programmed cell death. Interestingly, proportion of identified microDNA sequences has common loci of origin when compared between cell line experiments. While compatible with the original observation that microDNAs originate from a normal physiological process, obtained results imply complementary source of its production. Furthermore, non-random genesis of microDNAs depicted by redundancy between samples makes these entities possible candidates for new biomarker generation.

  2. Characterization of the microDNA through the response to chemotherapeutics in lymphoblastoid cell lines

    PubMed Central

    Mehanna, Pamela; Gagné, Vincent; Lajoie, Mathieu; Spinella, Jean-François; St-Onge, Pascal; Sinnett, Daniel; Brukner, Ivan

    2017-01-01

    Recently, a new class of extrachromosomal circular DNA, called microDNA, was identified. They are on average 100 to 400 bp long and are derived from unique non-repetitive genomic regions with high gene density. MicroDNAs are thought to arise from DNA breaks associated with RNA metabolism or replication slippage. Given the paucity of information on this entirely novel phenomenon, we aimed to get an additional insight into microDNA features by performing the microDNA analysis in 20 independent human lymphoblastoid cell lines (LCLs) prior and after treatment with chemotherapeutic drugs. The results showed non-random genesis of microDNA clusters from the active regions of the genome. The size periodicity of 190 bp was observed, which matches DNA fragmentation typical for apoptotic cells. The chemotherapeutic drug-induced apoptosis of LCLs increased both number and size of clusters further suggesting that part of microDNAs could result from the programmed cell death. Interestingly, proportion of identified microDNA sequences has common loci of origin when compared between cell line experiments. While compatible with the original observation that microDNAs originate from a normal physiological process, obtained results imply complementary source of its production. Furthermore, non-random genesis of microDNAs depicted by redundancy between samples makes these entities possible candidates for new biomarker generation. PMID:28877255

  3. Molecular signatures in response to Isoliquiritigenin in lymphoblastoid cell lines

    SciTech Connect

    Lee, Jae-Eun; Hong, Eun-Jung; Nam, Hye-Young; Hwang, Meeyul; Kim, Ji-Hyun; Han, Bok-Ghee; Jeon, Jae-Pil

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer We identified the inhibitory effect of ISL on cell proliferation of LCLs. Black-Right-Pointing-Pointer We found ISL-induced genes and miRNAs through microarray approach. Black-Right-Pointing-Pointer ISL-treated LCLs represented gene expression changes in cell cycle and p53 pathway. Black-Right-Pointing-Pointer We revealed 12 putative mRNA-miRNA functional pairs associated with ISL effect. -- Abstract: Isoliquiritigenin (ISL) has been known to induce cell cycle arrest and apoptosis of various cancer cells. However, genetic factors regulating ISL effects remain unclear. The aim of this study was to identify the molecular signatures involved in ISL-induced cell death of EBV-transformed lymphoblastoid cell lines (LCLs) using microarray analyses. For gene expression and microRNA (miRNA) microarray experiments, each of 12 LCL strains was independently treated with ISL or DMSO as a vehicle control for a day prior to total RNA extraction. ISL treatment inhibited cell proliferation of LCLs in a dose-dependent manner. Microarray analysis showed that ISL-treated LCLs represented gene expression changes in cell cycle and p53 signaling pathway, having a potential as regulators in LCL survival and sensitivity to ISL-induced cytotoxicity. In addition, 36 miRNAs including five miRNAs with unknown functions were differentially expressed in ISL-treated LCLs. The integrative analysis of miRNA and gene expression profiles revealed 12 putative mRNA-miRNA functional pairs. Among them, miR-1207-5p and miR-575 were negatively correlated with p53 pathway- and cell cycle-associated genes, respectively. In conclusion, our study suggests that miRNAs play an important role in ISL-induced cytotoxicity in LCLs by targeting signaling pathways including p53 pathway and cell cycle.

  4. Recombinant human epidermal growth factor accelerates the proliferation of irradiated human fibroblasts and keratinocytes in vitro and in vivo.

    PubMed

    Ryu, Seung-Hee; Moon, Soo Young; Yang, Youn-Joo; Moon, Sun Rock; Hong, Joon Pio; Choi, Jene; Lee, Sang-Wook

    2009-11-01

    Irradiation causes the impaired proliferation of cells lining mucosal membranes. Epidermal growth factor (EGF) facilitates proliferation of various skin cells; however, the wound healing effects of EGF on radiation-damaged cells is less well known. To evaluate the effects of recombinant human EGF (rhEGF) on the proliferation of cells following irradiation, we tested two types of fibroblast cell lines and one keratinocyte cell line. The viable cell numbers were significantly increased by rhEGF treatment for 24 h immediately after 8 Gy of irradiation. The most effective dose of rhEGF was 10 nM in all cell lines used in this study. The percentage of BrdU-labeled cells was also significantly increased by rhEGF treatment. To evaluate the effects of rhEGF on radiation-induced oral mucosal damage in BALB/c mice, we systematically injected 1 mg/kg/day EGF for three days after 17 Gy of irradiation. Administered rhEGF ameliorated radiation-induced mucosal damage in vivo. rhEGF significantly increased the epithelial cell layer thickness and the proliferation of basal layer cells as detected by Ki-67 staining. Our results suggest that rhEGF can be a therapeutic treatment for radiation-induced wounds by stimulating the proliferation of fibroblasts and keratinocytes following irradiation.

  5. Analysis of gene-expression profiles after gamma irradiation of normal human fibroblasts

    SciTech Connect

    Tachiiri, Seiji . E-mail: tachiiri@kuhp.kyoto-u.ac.jp; Katagiri, Toyomasa; Tsunoda, Tatsuhiko; Oya, Natsuo; Hiraoka, Masahiro; Nakamura, Yusuke

    2006-01-01

    Purpose: To understand comprehensive transcriptional profile of normal human fibroblast in response to irradiation. Methods and Materials: To identify genes whose expression is influenced by {gamma} radiation, we used a cDNA microarray to analyze expression of 23,000 genes in normal human fibroblasts at 7 timepoints (1, 3, 6, 12, 24, 48, and 72 hours) after 5 different doses (0.5, 2, 5, 15, and 50 Gy) of exposure. Results: Among the genes that showed altered expression patterns, some were already known to be regulated by irradiation, for instance ODC, EGR1, FGF2, PCNA, PKC, and several p53-target genes, including p53DINP1, BTG2, GADD45, and MDM2. The time course of each dose showed that from 350 to 600 genes were affected as to their expression; induction profiles characteristic to each dose were demonstrated. Of the total identified, only 89 genes were up-regulated; the vast majority was down-regulated over the 72-hour time course. We identified 21 genes that were distinctly induced by irradiation; 11 of them were functionally known, and 6 of those were p53-target genes. Conclusions: The results underscored the complexity of the transcriptional responses to irradiation, and the data should serve as a basis for global characterization of radiation-regulated genes and pathways.

  6. Surface nanomorphology of human dental enamel irradiated with an Er:YAG laser

    NASA Astrophysics Data System (ADS)

    Ţălu, Ş.; Contreras–Bulnes, R.; Morozov, I. A.; Rodríguez-Vilchis, L. E.; Montoya-Ayala, G.

    2016-02-01

    To determine the effects of Er:YAG laser irradiation on the surface nanomorphology of human dental enamel. Materials and methods: five samples of human dental enamel were divided into five groups: (a) I and II were irradiated with Er:YAG & water irrigation (12.7 J cm-2 and 25.5 J cm-2, respectively); (b) III and IV were Er:YAG laser irradiated & no water irrigation (12.7 J cm-2 and 25.5 J cm-2, respectively); (c) V or control (no laser irradiation). Nanomorphological changes were observed on 1 μm  ×  1 μm areas by AFM (contact mode and air). The partition functions and multifractal spectra were calculated. The graphical results showed that the larger the spectrum width Δα (Δα  =  α max  -  α min) of the multifractal spectra f(α) the more non-uniform the surface nanomorphology. One way analysis of variance (ANOVA) was performed (P  <  0.05) to distinguish significant differences between the groups. All the investigated surfaces exhibited multifractal behavior. The computational algorithm indicated that the multifractal spectra differ significantly from each other for the different groups. AFM (atomic force microscopy), the statistical surface roughness parameters, and multifractal analysis provided useful information about the surface nanomorphology and optimal surface characteristics. This approach could be extended to other enamel surfaces in order to characterize its structural 3D microrelief.

  7. Helium-neon and nitrogen laser irradiation accelerates the phagocytic activity of human monocytes.

    PubMed

    Hemvani, Nanda; Chitnis, Dhananjay Sadashiv; Bhagwanani, Nijram Satramdas

    2005-12-01

    Intracellular survival of mycobacteria within monocytes is a crucial stage in the pathogenesis of tuberculosis. The aim was to check intracellular survival of Mycobacterium fortuitum within the human monocytes exposed to He-Ne and nitrogen laser irradiation. Tuberculosis remains one of the most important infectious diseases for developing countries. Low-level laser therapy (LLLT) has been tried to treat tubercular cavitory lung disease with encouraging results. The in vitro photobiological effect of low level laser radiation on the intracellular mycobacteria needs to be evaluated before we could go for large clinical trials. The aliquots of human monocytes from peripheral blood of healthy volunteers and tuberculosis cases were exposed to He-Ne or nitrogen laser beam. The non-irradiated monocytes from the same source served as controls. The monocytes were then challenged with M. fortuitum, and surviving mycobacteria within monocytes were subjected to viable counts. Enhanced killing of mycobacterial cells was seen among monocytes exposed to He-Ne and nitrogen laser irradiation. He-Ne and nitrogen laser irradiation activates the monocytes to increase intracellular killing of mycobacteria.

  8. DNA damage in wounded, hypoxic and acidotic human skin fibroblast cell cultures after low laser irradiation

    NASA Astrophysics Data System (ADS)

    Hawkins Evans, D.; Mbene, A.; Zungu, I.; Houreld, N.; Abrahamse, H.

    2009-02-01

    Phototherapy has become more popular and widely used in the treatment of a variety of medical conditions. To ensure sound results as evidence of its effectiveness, well designed experiments must be conducted when determining the effect of phototherapy. Cell culture models such as hypoxic, acidotic and wounded cell cultures simulating different disease conditions including ischemic heart disease, diabetes and wound healing were used to determine the effect of laser irradiation on the genetic integrity of the cell. Even though phototherapy has been found to be beneficial in a wide spectrum of conditions, it has been shown to induce DNA damage. However, this damage appears to be repairable. The risk lies in the fact that phototherapy may help the medical condition initially but damage DNA at the same time leaving undetected damage that may result in late onset, more severe, induced medical conditions including cancer. Human skin fibroblasts were cultured and used to induce a wound (by the central scratch model), hypoxic (by incubation in an anaerobic jar, 95% N2 and 5% O2) and acidotic (reducing the pH of the media to 6.7) conditions. Different models were irradiated using a Helium-Neon (632.8 nm) laser with a power density of 2.07 mW/cm2 and a fluence of 5 J/cm2 or 16 J/cm2. The effect of the irradiation was determined using the Comet assay 1 and 24 h after irradiation. In addition, the Comet assay was performed with the addition of formamidopyrimidine glycosylase (FPG) obviating strand brakes in oxidized bases at a high fluence of 16 J/cm2. A significant increase in DNA damage was seen in all three injured models at both 1 and 24 h post-irradiation when compared to the normal un-injured cells. However, when compared to non-irradiated controls the acidotic model showed a significant decrease in DNA damage 24 h after irradiation indicating the possible induction of cellular DNA repair mechanisms. When wounded cells were irradiated with higher fluences of 16 J/cm2

  9. Inflammatory cytokines are suppressed by light-emitting diode irradiation of P. gingivalis LPS-treated human gingival fibroblasts: inflammatory cytokine changes by LED irradiation.

    PubMed

    Choi, HongRan; Lim, WonBong; Kim, InAe; Kim, JiSun; Ko, YoungJong; Kwon, Hyukil; Kim, SangWoo; Kabir, K M Ahsan; Li, Xiaojie; Kim, Oksu; Lee, YoungJoon; Kim, SeoYune; Kim, OkJoon

    2012-03-01

    Human gingival fibroblasts (hGFs) play an important role in the inflammatory reaction to lipopolysaccharide (LPS) from P. gingivalis, which infects periodontal connective tissue. In addition, although light-emitting diode (LED) irradiation has been reported to have biostimulatory effects, including anti-inflammatory activity, the pathological mechanisms of these effects are unclear. This study examined the effects of 635-nm irradiation of P. gingivalis LPS-treated human gingival fibroblasts on inflammatory cytokine profiles and the mitogen-activated protein kinase (MAPK) pathway, which is involved in cytokine production. Gingival fibroblasts treated or not treated with P. gingivalis LPS were irradiated with 635-nm LED light, and cytokine profiles in the supernatant were assessed using a human inflammation antibody array. Expression of cyclooxyginase-2 (COX-2) protein and phosphorylation of extracellular signal-regulated kinase (ERK 1/2), p38, and c-Jun-N-terminal kinase (JNK) were assessed by Western-blot analysis to determine the effects on the MAPK pathway, and prostaglandin E(2) (PGE(2)) in the supernatant was measured using an enzyme-linked immunoassay. COX-2 protein expression and PGE(2) production were significantly increased in the LPS-treated group and decreased by LED irradiation. LPS treatment of gingival fibroblasts led to the increased release of the pro-inflammatory-related cytokines interleukin-6 (IL-6) and IL-8, whereas LED irradiation inhibited their release. Analysis of MAPK signal transduction revealed a considerable decrease in p38 phosphorylation in response to 635-nm radiation either in the presence or absence of LPS. In addition, 635-nm LED irradiation significantly promoted JNK phosphorylation in the presence of LPS. LED irradiation can inhibit activation of pro-inflammatory cytokines, mediate the MAPK signaling pathway, and may be clinically useful as an anti-inflammatory tool.

  10. Chromosomal Instability in the progeny of human irradiated cells

    NASA Astrophysics Data System (ADS)

    Testard, I.; Boissière, A.; Martins, L. M.; Sabatier, L.

    Manned space missions recently increased in number and duration, thus it became important to estimate the biological risks encountered by astronauts. They are exposed to cosmic and galactic rays, a complex mixture of different radiations. In addition to the measurements realized by physical dosimeters, it becomes essential to estimate real biologically effective doses and compare them to physical doses. Biological dosimetry of radiation exposures has been widely performed using cytogenetic analysis of chromosomes. This approach has been used for many years in order to estimate absorbed doses in accidental or chronic overexposures of humans. Recent studies show that some alterations can appear many cell generations after the initial radiation exposure as a delayed genomic instability. This delayed instability is characterized by the accumulation of cell alterations leading to cell transformation, delayed cell death and mutations. Chromosome instability was shown in vitro in different model systems (Sabatier et al., 1992; Marder and Morgan, 1993; Kadhim et al., 1994 and Holmberg et al., 1993, 1995). All types of radiation used induce chromosome instability; however, heavy ions cause the most damage. The period of chromosome instability followed by the formation of clones with unbalanced karyotypes seems to be shared by cancer cells. The shortening of telomere sequences leading to the formation of telomere fusions is an important factor in the appearance of this chromosome instability.

  11. Quiescence does not affect p53 and stress response by irradiation in human lung fibroblasts

    SciTech Connect

    Dai, Jiawen; Itahana, Koji; Baskar, Rajamanickam

    2015-02-27

    Cells in many organs exist in both proliferating and quiescent states. Proliferating cells are more radio-sensitive, DNA damage pathways including p53 pathway are activated to undergo either G{sub 1}/S or G{sub 2}/M arrest to avoid entering S and M phase with DNA damage. On the other hand, quiescent cells are already arrested in G{sub 0}, therefore there may be fundamental difference of irradiation response between proliferating and quiescent cells, and this difference may affect their radiosensitivity. To understand these differences, proliferating and quiescent human normal lung fibroblasts were exposed to 0.10–1 Gy of γ-radiation. The response of key proteins involved in the cell cycle, cell death, and metabolism as well as histone H2AX phosphorylation were examined. Interestingly, p53 and p53 phosphorylation (Ser-15), as well as the cyclin-dependent kinase inhibitors p21 and p27, were induced similarly in both proliferating and quiescent cells after irradiation. Furthermore, the p53 protein half-life, and expression of cyclin A, cyclin E, proliferating cell nuclear antigen (PCNA), Bax, or cytochrome c expression as well as histone H2AX phosphorylation were comparable after irradiation in both phases of cells. The effect of radioprotection by a glycogen synthase kinase 3β inhibitor on p53 pathway was also similar between proliferating and quiescent cells. Our results showed that quiescence does not affect irradiation response of key proteins involved in stress and DNA damage at least in normal fibroblasts, providing a better understanding of the radiation response in quiescent cells, which is crucial for tissue repair and regeneration. - Highlights: • p53 response by irradiation was similar between proliferating and quiescent cells. • Quiescent cells showed similar profiles of cell cycle proteins after irradiation. • Radioprotection of GSK-3β inhibitor caused similar effects between these cells. • Quiescence did not affect p53 response despite its

  12. Metabolic changes in humans following total body irradiation. Report for February 1960-October 1961

    SciTech Connect

    Not Available

    1988-11-29

    These studies are designed to obtain new information about the metabolic effects of total body and partial body irradiation so as to have a better understanding of the acute and subacute effects of irradiation in the human. The initial studies are pointed toward the elucidation of biological indicators of radiation effects in humans. The major parameters being investigated at present are urinary amino aciduria and alterations in immunological patterns. Certain other parameters such as creatine and creatinine excretion and hematological effects are also being followed. The long-term program envisions carrying out the various observations at dose levels of 100 rad and gradually increasing the dose to 150, 200, 250 and 300 rad. Eventually doses up to 600 rad are anticipated. Also comparison of effects of radiomimetic drugs with total body radiation will be studied.

  13. Continuous irradiation with a 633-nm light-emitting diode exerts an anti-aging effect on human skin cells.

    PubMed

    Kim, Hak Sun; Park, Won Sang; Baek, Jong-In; Lee, Bo-Sub; Yoo, Dae Sung; Park, Si Jun

    2015-02-01

    Accumulating evidence has indicated that the light source emitted from light‑emitting diode (LED) has a potential anti-aging effect on human skin. Studies using single and interval LED irradiation have documented such effects; however, to the best of our knowledge, the anti-aging effects of continuous LED irradiation have not yet been investigated. In the present study, we demonstrated that continuous irradiation with a 633±3-nm LED exerted anti-aging effects in both in vitro and ex vivo experiments. More specifically, irradiation with a 633-nm LED for 2 days increased the synthesis of type 1 procollagen and decreased the expression of matrix metalloproteinase (MMP)1 and MMP2 in skin fibroblasts. In addition, irradiation with a 633-nm LED decreased the expression levels of inflammatory genes, such has cyclooxygenase-2 (COX-2), and interleukin-1-α (IL-1α) in keratinocytes. Furthermore, a 14-day LED irradiation moderately increased keratinocyte proliferation. Using human skin explants, we confirmed the safety of this 633-nm LED irradiation, which resulted in unaltered morphology and allergy-free potential in human tissue. Overall, these data provide insight into the anti-aging effects of continuous LED irradiation on human skin.

  14. Expression profiling of human melanocytes in response to UV-B irradiation

    PubMed Central

    López, Saioa; Smith-Zubiaga, Isabel; Alonso, Santos

    2015-01-01

    A comprehensive gene expression analysis of human melanocytes was performed assessing the transcriptional profile of dark melanocytes (DM) and light melanocytes (LM) at basal conditions and after UV-B irradiation at different time points (6, 12 and 24 h), and in culture with different keratinocyte-conditioned media (KCM + and KCM −). The data, previously published in [1], have been deposited in NCBI's Gene Expression Omnibus (GEO accession number: GSE70280). PMID:26697372

  15. Transplantation of human fetal-derived neural stem cells improves cognitive function following cranial irradiation.

    PubMed

    Acharya, Munjal M; Christie, Lori-Ann; Hazel, Thomas G; Johe, Karl K; Limoli, Charles L

    2014-01-01

    Treatment of central nervous system (CNS) malignancies typically involves radiotherapy to forestall tumor growth and recurrence following surgical resection. Despite the many benefits of cranial radiotherapy, survivors often suffer from a wide range of debilitating and progressive cognitive deficits. Thus, while patients afflicted with primary and secondary malignancies of the CNS now experience longer local regional control and progression-free survival, there remains no clinical recourse for the unintended neurocognitive sequelae associated with their cancer treatments. Multiple mechanisms contribute to disrupted cognition following irradiation, including the depletion of radiosensitive populations of stem and progenitor cells in the hippocampus. We have explored the potential of using intrahippocampal transplantation of human stem cells to ameliorate radiation-induced cognitive dysfunction. Past studies demonstrated the capability of cranially transplanted human embryonic (hESCs) and neural (hNSCs) stem cells to functionally restore cognition in rats 1 and 4 months after cranial irradiation. The present study employed an FDA-approved fetal-derived hNSC line capable of large scale-up under good manufacturing practice (GMP). Animals receiving cranial transplantation of these cells 1 month following irradiation showed improved hippocampal spatial memory and contextual fear conditioning performance compared to irradiated, sham surgery controls. Significant newly born (doublecortin positive) neurons and a smaller fraction of glial subtypes were observed within and nearby the transplantation core. Engrafted cells migrated and differentiated into neuronal and glial subtypes throughout the CA1 and CA3 subfields of the host hippocampus. These studies expand our prior findings to demonstrate that transplantation of fetal-derived hNSCs improves cognitive deficits in irradiated animals, as assessed by two separate cognitive tasks.

  16. Use of lymphoblastoid cell lines to evaluate the hypersensitivity to ultraviolet radiation in Cockayne syndrome

    SciTech Connect

    Otsuka, F.; Tarone, R.E.; Cayeux, S.; Robbins, J.H.

    1984-05-01

    Cockayne syndrome (CS) is a rare autosomal recessive disease characterized by acute sun sensitivity, cachectic dwarfism, and neurologic and skeletal abnormalities. Cultured skin fibroblasts from patients with this disease are known to be hypersensitive to the lethal effects of 254-nm UV radiation. The authors have studied the sensitivity of 254-nm UV radiation of lymphoblastoid lines derived from 3 typical CS patients, 1 atypical CS patient who had a very late age of onset of clinical manifestations, 2 patients who had both xeroderma pigmentosum (XP) and typical CS, and 3 heterozygous parents of these patients. Post-UV survival was determined by the trypan-blue dye-exclusion method. The lymphoblastoid lines from the 3 typical CS patients, the atypical CS patient, and the 2 patients with both CS and XP had decreased post-UV viability in comparison with lines from normal donors. Lines from the heterozygous parents had normal post-UV viability. The post-UV viability of the typical CS lines was similar to that of a XP complementation group C line. The relative post-UV viability of lymphoblastoid lines from the typical CS patients was similar to the relative post-UV survival of their fibroblast lines. The lymphoblastoid line from the atypical CS patient had a post-UV viability similar to that of the typical CS patients. Thus, the relative hypersensitivity of CS patients cells in vitro does not reflect the severity or age of onset of the patients clinical manifestations. The lymphoblastoid lines from the 2 patients who had both CS and XP were significantly more sensitive to the UV radiation than those from patients with only CS. Our studies demonstrate that lymphoblastoid lines from patients with CS are appropriate and useful cell lines for the study of the inherited hypersensitivity to UV radiation.

  17. Removal of cyclobutane pyrimidine dimers from a UV-irradiated shuttle vector introduced into human cells

    SciTech Connect

    Ganesan, A.K.; Hanawalt, P.C. )

    1994-05-01

    A shuttle vector (pZH-1) carrying the E. Coli lacZ gene under control of the SV40 early promoter was irradiated with UV and introduced into repair-proficient or repair-deficient human cell lines. The expression of irradiated lacZ compared to unirradiated lacZ was greater in repair-proficient cells (HT-1080) than in repair-deficient cells (XP12RO-SV40) belonging to xeroderma pigmentosum complementation group A. To ascertain whether the expression of lacZ in the repair-proficient cells was correlated with the removal of cyclobutane pyrimidine dimers (CPDs), the authors purified DNA from the recipient cells and used the CPD-specific enzyme T4 endonuclease V to measure the frequency of CPDs remaining in the plasmid as a whole and in two restriction fragments derived from it. They found that removal of CPDs occurred in both fragments in the repair-proficient cells but not in the repair-deficient cells. The results provide the first direct evidence for the removal of CPDs from UV irradiated plasmids introduced into human cells and support the notion that expression of the UV-damaged lacZ gene in repair-proficient human cells reflects the removal of transcription blocking lesions from the gene.

  18. The stress caused by nitrite with titanium dioxide nanoparticles under UVA irradiation in human keratinocyte cell.

    PubMed

    Tu, Min; Huang, Yi; Li, Hai-Ling; Gao, Zhong-Hong

    2012-09-04

    Our previous work found that in the presence of nitrite, titanium dioxide nanoparticles can cause protein tyrosine nitration under UVA irradiation in vivo. In this paper, the human keratinocyte cells was used as a skin cell model to further study the photo-toxicity of titanium dioxide nanoparticles when nitrite was present. The results showed that nitrite increased the photo-toxicity of titanium dioxide in a dose-dependant manner, and generated protein tyrosine nitration in keratinocyte cells. Morphological study of keratinocyte cells suggested a specific apoptosis mediated by apoptosis inducing factor. It was also found the main target nitrated in cells was cystatin-A, which expressed abundantly in cytoplasm and functioned as a cysteine protease inhibitor. The stress induced by titanium dioxide with nitrite under UVA irradiation in human keratinocyte cells appeared to trigger the apoptosis inducing factor mediated cell death and lose the inhibition of active caspase by cystatin-A. We conclude that nitrite can bring new damage and stress to human keratinocyte cells with titanium dioxide nanoparticles under UVA irradiation.

  19. Microarray Analysis of Human Liver Cells irradiated by 80MeV/u Carbon Ions

    NASA Astrophysics Data System (ADS)

    Wang, Xiao; Tian, Xiaoling; Kong, Fuquan; Li, Qiang; Jin, Xiaodong; Dai, Zhongying; Zhang, Hong; Yang, Mingjian; Zhao, Kui

    Objective Biological effect of heavy ion beam has the important significance for cancer therapy and space exploring owing its high LET and RBE, low OER, especially forming Bragg spike at the end of the tracks of charged particles. More serious damage for cells are induced by heavy ions and difficult repair than other irradiation such as X-ray and ν-ray . To explore the molecular mechanism of biological effect caused by heavy ionizing radiation (HIR) and to construct the gene expression profile database of HIR-induced human liver cells L02 by microarray analysis. Methods In this study, L02 cells were irradiated by 80MeV/u carbon ions at 5 Gy delivered by HIRFL (Heavy Ion Research Facility in Lanzhou) at room temperature. Total RNAs of cells incubated 6 hours and 24hours after irradiation were extracted with Trizol. Unirradiated cells were used as a control. RNAs were transcripted into cDNA by reverse transcription and labelled with cy5-dCTP and cy3-dCTP respectively. A human genome oligonucleotide set consisting of 5 amino acid-modified 70-mer probes and representing 21,329 well-characterized Homo sapiens genes was selected for microarray analysis and printed on amino-silaned glass slides. Arrays were fabricated using an OmniGrid microarrayer. Only genes whose alteration tendency was consistent in both microarrays were selected as differentially expressed genes. The Affymetrix's short oligonucleotide (25-mer) HG U133A 2.0 array analyses were performed per the manufacturer's instructions. Results Of the 21,329 genes tested, 37 genes showed changes in expression level with ratio higher than 2.0 and lower than 0.5 at 6hrs after irradiation. There were 19 genes showing up-regulation in radiated L02 cells, whereas 18 genes showing down-regulation; At 24hrs after irradiation, 269 genes showed changes in expression level with ratio higher than 2.0 and lower than 0.5. There were 67 genes showing up-regulation in radiated L02 cells, whereas 202 genes showing down

  20. Dramatic increase in oxidative stress in carbon-irradiated normal human skin fibroblasts.

    PubMed

    Laurent, Carine; Leduc, Alexandre; Pottier, Ivannah; Prévost, Virginie; Sichel, François; Lefaix, Jean-Louis

    2013-01-01

    Skin complications were recently reported after carbon-ion (C-ion) radiation therapy. Oxidative stress is considered an important pathway in the appearance of late skin reactions. We evaluated oxidative stress in normal human skin fibroblasts after carbon-ion vs. X-ray irradiation. Survival curves and radiobiological parameters were calculated. DNA damage was quantified, as were lipid peroxidation (LPO), protein carbonylation and antioxidant enzyme activities. Reduced and oxidized glutathione ratios (GSH/GSSG) were determined. Proinflammatory cytokine secretion in culture supernatants was evaluated. The relative biological effectiveness (RBE) of C-ions vs. X-rays was 4.8 at D₀ (irradiation dose corresponding to a surviving fraction of 37%). Surviving fraction at 2 Gy (SF2) was 71.8% and 7.6% for X-rays and C-ions, respectively. Compared with X-rays, immediate DNA damage was increased less after C-ions, but a late increase was observed at D(10%) (irradiation dose corresponding to a surviving fraction of 10%). LPO products and protein carbonyls were only increased 24 hours after C-ions. After X-rays, superoxide dismutase (SOD) activity was strongly increased immediately and on day 14 at D(0%) (irradiation dose corresponding to a surviving fraction of around 0%), catalase activity was unchanged and glutathione peroxidase (GPx) activity was increased only on day 14. These activities were decreased after C-ions compared with X-rays. GSH/GSSG was unchanged after X-rays but was decreased immediately after C-ion irradiation before an increase from day 7. Secretion of IL-6 was increased at late times after X-ray irradiation. After C-ion irradiation, IL-6 concentration was increased on day 7 but was lower compared with X-rays at later times. C-ion effects on normal human skin fibroblasts seemed to be harmful in comparison with X-rays as they produce late DNA damage, LPO products and protein carbonyls, and as they decrease antioxidant defences. Mechanisms leading to this

  1. Dramatic Increase in Oxidative Stress in Carbon-Irradiated Normal Human Skin Fibroblasts

    PubMed Central

    Laurent, Carine; Leduc, Alexandre; Pottier, Ivannah; Prévost, Virginie; Sichel, François; Lefaix, Jean-Louis

    2013-01-01

    Skin complications were recently reported after carbon-ion (C-ion) radiation therapy. Oxidative stress is considered an important pathway in the appearance of late skin reactions. We evaluated oxidative stress in normal human skin fibroblasts after carbon-ion vs. X-ray irradiation. Survival curves and radiobiological parameters were calculated. DNA damage was quantified, as were lipid peroxidation (LPO), protein carbonylation and antioxidant enzyme activities. Reduced and oxidized glutathione ratios (GSH/GSSG) were determined. Proinflammatory cytokine secretion in culture supernatants was evaluated. The relative biological effectiveness (RBE) of C-ions vs. X-rays was 4.8 at D0 (irradiation dose corresponding to a surviving fraction of 37%). Surviving fraction at 2 Gy (SF2) was 71.8% and 7.6% for X-rays and C-ions, respectively. Compared with X-rays, immediate DNA damage was increased less after C-ions, but a late increase was observed at D10% (irradiation dose corresponding to a surviving fraction of 10%). LPO products and protein carbonyls were only increased 24 hours after C-ions. After X-rays, superoxide dismutase (SOD) activity was strongly increased immediately and on day 14 at D0% (irradiation dose corresponding to a surviving fraction of around 0%), catalase activity was unchanged and glutathione peroxidase (GPx) activity was increased only on day 14. These activities were decreased after C-ions compared with X-rays. GSH/GSSG was unchanged after X-rays but was decreased immediately after C-ion irradiation before an increase from day 7. Secretion of IL-6 was increased at late times after X-ray irradiation. After C-ion irradiation, IL-6 concentration was increased on day 7 but was lower compared with X-rays at later times. C-ion effects on normal human skin fibroblasts seemed to be harmful in comparison with X-rays as they produce late DNA damage, LPO products and protein carbonyls, and as they decrease antioxidant defences. Mechanisms leading to this

  2. In vitro analysis of low-level laser irradiation on human osteoblast-like cells proliferation

    NASA Astrophysics Data System (ADS)

    Bloise, Nora; Saino, Enrica; Bragheri, Francesca; Minzioni, Paolo; Cristiani, Ilaria; Imbriani, Marcello; Visai, Livia

    2011-07-01

    The objective of this study was to examine the in vitro effect of a single or a multiple doses of low-level laser irradiation (LLLI) on proliferation of the human osteosarcoma cell line, SAOS-2. SAOS-2 cells were divided in five groups and exposed to LLLI (659 nm diode laser; 11 mW power output): group I as a control (dark), group II exposed to a single laser dose of 1 J/cm2, group III irradiated with a single dose of 3 J/cm2, and group IV and V exposed for three consecutive days to 1 or 3 J/cm², respectively. Cellular proliferation was assessed daily up to 7 days of culturing. The obtained results showed an increase in proliferative capacity of SAOS-2 cells during the first 96 h of culturing time in once-irradiated cells, as compared to control cells. Furthermore, a significantly higher proliferation in the group IV and V was detected if compared to a single dose or to control group after 96 h and 7 days. In conclusion, the effect of the single dose on cell proliferation was transitory and repeated irradiations were necessary to observe a strong enhancement of SAOS-2 growth. As a future perspective, we would like to determine the potential of LLLI as a new approach for promoting bone regeneration onto biomaterials.

  3. Tensile Bond Strengths of Two Adhesives on Irradiated and Nonirradiated Human Dentin

    PubMed Central

    Bernard, Cécile; Abouelleil, Hazem; Gustin, Marie-Paule; Grosgogeat, Brigitte

    2015-01-01

    The aim of this study was to assess the effect of radiotherapy on bond efficiency of two different adhesive systems using tensile bond strength test. Twenty extracted teeth after radiotherapy and twenty nonirradiated extracted teeth were used. The irradiation was applied in vivo to a minimal dose of 50 Gy. The specimens of each group were randomly assigned to two subgroups to test two different adhesive systems. A three-step/etch-and-rinse adhesive system (Optibond FL) and a two-steps/self-etch adhesive system (Optibond XTR) were used. Composite buildups were performed with a nanohybrid composite (Herculite XTR). All specimens were submitted to thermocycling ageing (10000 cycles). The specimens were sectioned in 1 mm2 sticks. Microtensile bond strength tests were measured. Nonparametric statistical analyses were performed due to nonnormality of data. Optibond XTR on irradiated and nonirradiated teeth did not show any significant differences. However, Optibond FL bond strength was more effective on nonirradiated teeth than on irradiated teeth. Within the limitations of an in vitro study, it can be concluded that radiotherapy had a significant detrimental effect on bond strength to human dentin. However, it seems that adhesive choice could be adapted to the substrata. According to the present study, the two-steps/self-etch (Optibond XTR) adhesive system tested could be more effective on irradiated dentin compared to three-steps/etch-and-rinse adhesive system (Optibond FL). PMID:26783528

  4. Induction of proteins and mRNAs after uv irradiation of human epidermal keratinocytes

    SciTech Connect

    Kartasova, T.; Ponec, M.; van de Putte, P.

    1988-02-01

    uv sensitivity of cultured human epidermal keratinocytes was analyzed at different growth conditions and compared with the sensitivity of dermal fibroblasts derived from the same skin specimen. No significant differences in survival curves were found between these two cell types, although keratinocytes grown under standard conditions were slightly more resistant to uv irradiation than fibroblasts. The extracellular concentration of calcium appeared to be critical not only in the regulation of keratinocyte proliferation and differentiation, but also in the uv sensitivity of these cells: keratinocytes grown under conditions which favor cell proliferation (low calcium concentration) are more resistant to uv irradiation than those grown under conditions favoring differentiation (high calcium concentration). Two-dimensional protein gel electrophoresis was used to detect a possible effect of uv irradiation on the accumulation of specific mRNAs in the cytoplasm and/or on the synthesis of specific proteins. Proteins were pulse labeled in vivo with (/sup 35/S)methionine or synthesized in vitro in rabbit reticulocyte lysates on mRNA isolated from keratinocytes that were irradiated with different uv doses at different periods of time prior to isolation. Alterations in expression were demonstrated for several proteins in both in vivo and in vitro experiments.

  5. Antioxidant protection against curative and palliative doses of ionizing irradiation in human blood decreases with aging.

    PubMed

    Kasapović, Jelena; Stojiljković, Vesna; Gavrilović, Ljubica; Popović, Nataša; Milićević, Zorka

    2012-01-01

    Reactive oxygen species (ROS) are independently recognized to play a significant role in radiation-induced damage on healthy tissue and in aging process. However, an age-related alteration of antioxidant (AO) system in radiation response in humans is poorly investigated. The aim of this paper was to evaluate the irradiation effects on the activities and expression of AO system in the blood of healthy women during aging. Blood samples were irradiated with curative and palliative doses of 2 Gy or 9 Gy γ-rays. AO capacity for detoxification of O(2)•(-) and H(2)O(2) in response to 2 Gy γ-irradiation decreases in women above 58 years, while in response to 9 Gy shows signs of weakening after 45 years of age. Due to reduction of AO capacity during aging, cytotoxic effects of curative and palliative doses of irradiation, mediated by ROS, may significantly increase in older subjects, while removal of H(2)O(2) excess could reduce them.

  6. Mapping of UV photoproducts along the c-jun promoter in UV-irradiated human cells

    SciTech Connect

    Tornaletti, S.; Pfeifer, G.P.

    1994-12-31

    The formation of UV photoproducts is implicated in the induction of mutations and the development of skin cancer. Cyclobutane dimers and (6-4) photoproducts are the two major causes of mutagenic DNA photoproducts produced by UV irradiation. It is known that the distribution of these DNA adducts is not only influenced by DNA sequence but also by chromatin structure. To analyse possible effects of chromatin structure on the photoproduct spectrum, we have compared the distribution of cyclobutane pyrimidine dimers and pyrimidine (6-4) pyrimidine photoproducts, along the c-jun promoter in UV-irradiated HeLa cells, with that obtained from irradiated purified DNA. After UV irradiation, cyclobutane pyrimidine dimers and pyrimidine (6-4) pyrimidine photoproducts were converted into DNA strand breaks by treatment with hot pyrimidine or T4 endonuclease v/photolyase cleavage. Ligation-mediated PCR was then used to map both types of UV photoproducts at the DNA sequence level. Photoproduct frequency within transcription factor binding sties was suppressed or enhanced relative to naked DNA. Photofootprints were localized to an AP1 like sequence (nt. -71 to -64), a CCAAT box element (nt. -91 to -87), and SP1 sequence (nt. -123 to -118), a nuclear factor jun (NF-jun) site (nt. -140 to -132) and a second AP1 like sequence (nt. -190 to -183). These findings have possible implications on molecular mechanisms of mutagenesis in the human genome.

  7. Crystalline structure of human enamel irradiated with Er,Cr:YSGG laser

    NASA Astrophysics Data System (ADS)

    Bachmann, L.; Rosa, K.; da Ana, P. A.; Zezell, D. M.; Craievich, A. F.; Kellermann, G.

    2009-02-01

    The Er,Cr:YSGG system is commonly employed in tissue removal, but recently it has also been clinically evaluated for caries prevention. The present work explains the clinical and pre-clinical observations on the basis of the crystallographic changes that this laser can produce in the dental enamel. The analyzed samples were obtained from sound human third molar teeth. The laser irradiation was conducted with a Er,Cr:YSGG laser with 12.5 mJ/pulse, 0.25 W, and 2.8 J/cm2. The laser device operates at a wavelength of 2.79 μm, and the pulse width duration is 140 μs, with a repetition rate of 20 Hz of spot size of 750 μm. The crystalline structure of the samples was evaluated by X-ray diffraction at a synchrotron beamline The X-ray beam was configured at a grazing angle, to maximize the surface diffraction signal and to better detect the possible new crystallographic phase produced after the laser irradiation. It was observed that the crystallographic structure tetracalcium phosphate (TetCP, JCPDF 25-1137) exhibits several peaks that match more precisely with the new experimental peaks of the irradiated enamel. The present results suggesting the coexistence of tetracalcium phosphate with hydroxyapatite in enamel irradiated with Er,Cr:YSGG laser and can be the answer to the clinical and pre-clinical observations reported in the literature.

  8. Action of low-power laser irradiation on the proliferation of human gingival fibroblasts in vitro

    NASA Astrophysics Data System (ADS)

    Almeida-Lopes, Luciana; Jaeger, Marcia M. M.; Brugnera, Aldo, Jr.; Rigau, Josepa

    1998-04-01

    The low level power laser has been used in dental treatments aiming to improve tissue healing. An in vitro study was performed to analyze the laser influence on gingival fibroblast. A human gingival fibroblast culture (LMF) was produced in DME medium with 10% bovine fetal serum (BFS) cells (LMF) were allocated in Petri plates and cultured in different SFB concentrations (0%, 5% e 10%). After 48 hours the plates were divided in 9 groups: 3 control: 3 irradiated by 635 nm laser; and 3 irradiated by 780 nm laser. The cultured cells received 4 applications, in 12 hours intervals, with energy dosage of 2 joules for each plate, by means of a punctual technique. The growth curves showed that the growth levels were lower in low BFS concentrations. The irradiation with laser accelerated the growth rate in all groups. Additionally, the number of cells developed in low BFS concentration (5%) and irradiated was similar to the number of control cells developed in ideal conditions (10% BFS). There was no statistically significant differences between the effects of the two types of laser studied.

  9. Unstable Chromosome Aberrations Do Not Accumulate in Normal Human Fibroblast after Fractionated X-Irradiation

    PubMed Central

    Ojima, Mitsuaki; Ito, Maki; Suzuki, Keiji; Kai, Michiaki

    2015-01-01

    We determined the frequencies of dicentric chromosomes per cell in non-dividing confluent normal human fibroblasts (MRC-5) irradiated with a single 1 Gy dose or a fractionated 1 Gy dose (10X0.1 Gy, 5X0.2 Gy, and 2X0.5 Gy). The interval between fractions was between 1 min to 1440 min. After the completion of X-irradiation, the cells were incubated for 24 hours before re-plating at a low density. Then, demecolcine was administrated at 6 hours, and the first mitotic cells were collected for 42 hours. Our study demonstrated that frequencies of dicentric chromosomes in cells irradiated with a 1 Gy dose at different fractions were significantly reduced if the fraction interval was increased from 1 min to 5 min (p<0.05, χ2-test). Further increasing the fraction interval from 5 up to 1440 min did not significantly affect the frequency of dicentric chromosomes. Since misrejoining of two independent chromosome breaks introduced in close proximity gives rise to dicentric chromosome, our results indicated that such circumstances might be quite infrequent in cells exposed to fractionated X-irradiation with prolonged fraction intervals. Our findings should contribute to improve current estimation of cancer risk from chronic low-dose-rate exposure, or intermittent exposure of low-dose radiation by medical exposure. PMID:25723489

  10. Ionising irradiation alters the dynamics of human long interspersed nuclear elements 1 (LINE1) retrotransposon.

    PubMed

    Tanaka, Atsushi; Nakatani, Youko; Hamada, Nobuyuki; Jinno-Oue, Atsushi; Shimizu, Nobuaki; Wada, Seiichi; Funayama, Tomoo; Mori, Takahisa; Islam, Salequl; Hoque, Sheikh Ariful; Shinagawa, Masahiko; Ohtsuki, Takahiro; Kobayashi, Yasuhiko; Hoshino, Hiroo

    2012-09-01

    It is important to identify the mechanism by which ionising irradiation induces various genomic alterations in the progeny of surviving cells. Ionising irradiation activates mobile elements like retrotransposons, although the mechanism of its phenomena consisting of transcriptions and insertions of the products into new sites of the genome remains unclear. In this study, we analysed the effects of sparsely ionising X-rays and densely ionising carbon-ion beams on the activities of a family of active retrotransposons, long interspersed nuclear elements 1 (L1). We used the L1/reporter knock-in human glioma cell line, NP-2/L1RP-enhanced GFP (EGFP), that harbours full-length L1 tagged with EGFP retrotransposition detection cassette (L1RP-EGFP) in the chromosomal DNA. X-rays and carbon-ion beams similarly increased frequencies the transcription from L1RP-EGFP and its retrotransposition. Short-sized de novo L1RP-EGFP insertions with 5'-truncation were induced by X-rays, while full-length or long-sized insertions (>5 kb, containing ORF1 and ORF2) were found only in cell clones irradiated by the carbon-ion beams. These data suggest that X-rays and carbon-ion beams induce different length of de novo L1 insertions, respectively. Our findings thus highlight the necessity to investigate the mechanisms of mutations caused by transposable elements by ionising irradiation.

  11. Photothermal response of human and murine cancer cells to multiwalled carbon nanotubes after laser irradiation.

    PubMed

    Fisher, Jessica W; Sarkar, Saugata; Buchanan, Cara F; Szot, Christopher S; Whitney, Jon; Hatcher, Heather C; Torti, Suzy V; Rylander, Christopher G; Rylander, Marissa Nichole

    2010-12-01

    This study demonstrates the capability of multiwalled carbon nanotubes (MWNTs) coupled with laser irradiation to enhance treatment of cancer cells through enhanced and more controlled thermal deposition, increased tumor injury, and diminished heat shock protein (HSP) expression. We also explored the potential promise of MWNTs as drug delivery agents by observing the degree of intracellular uptake of these nanoparticles. To determine the heat generation capability of MWNTs, the absorption spectra and temperature rise during heating were measured. Higher optical absorption was observed for MWNTs in water compared with water alone. For identical laser parameters, MWNT-containing samples produced a significantly greater temperature elevation compared to samples treated with laser alone. Human prostate cancer (PC3) and murine renal carcinoma (RENCA) cells were irradiated with a 1,064-nm laser with an irradiance of 15.3 W/cm(2) for 2 heating durations (1.5 and 5 minutes) alone or in combination with MWNT inclusion. Cytotoxicity and HSP expression following laser heating was used to determine the efficacy of laser treatment alone or in combination with MWNTs. No toxicity was observed for MWNTs alone. Inclusion of MWNTs dramatically decreased cell viability and HSP expression when combined with laser irradiation. MWNT cell internalization was measured using fluorescence and transmission electron microscopy following incubation of MWNTs with cells. With increasing incubation duration, a greater number of MWNTs were observed in cellular vacuoles and nuclei. These findings offer an initial proof of concept for the application of MWNTs in cancer therapy.

  12. Photothermal Response of Human and Murine Cancer Cells to Multiwalled Carbon Nanotubes after Laser Irradiation

    PubMed Central

    Fisher, Jessica W.; Sarkar, Saugata; Buchanan, Cara F.; Szot, Christopher S.; Whitney, Jon; Hatcher, Heather C.; Torti, Suzy V.; Rylander, Christopher G.; Rylander, Marissa Nichole

    2013-01-01

    This study demonstrates the capability of multiwalled carbon nanotubes (MWNTs) coupled with laser irradiation to enhance treatment of cancer cells through enhanced and more controlled thermal deposition, increased tumor injury, and diminished heat shock protein (HSP) expression. We also explored the potential promise of MWNTs as drug delivery agents by observing the degree of intracellular uptake of these nanoparticles. To determine the heat generation capability of MWNTs, the absorption spectra and temperature rise during heating were measured. Higher optical absorption was observed for MWNTs in water compared with water alone. For identical laser parameters, MWNT-containing samples produced a significantly greater temperature elevation compared to samples treated with laser alone. Human prostate cancer (PC3) and murine renal carcinoma (RENCA) cells were irradiated with a 1,064-nm laser with an irradiance of 15.3 W/cm2 for 2 heating durations (1.5 and 5 minutes) alone or in combination with MWNT inclusion. Cytotoxicity and HSP expression following laser heating was used to determine the efficacy of laser treatment alone or in combination with MWNTs. No toxicity was observed for MWNTs alone. Inclusion of MWNTs dramatically decreased cell viability and HSP expression when combined with laser irradiation. MWNT cell internalization was measured using fluorescence and transmission electron microscopy following incubation of MWNTs with cells. With increasing incubation duration, a greater number of MWNTs were observed in cellular vacuoles and nuclei. These findings offer an initial proof of concept for the application of MWNTs in cancer therapy. PMID:21098701

  13. Differential gene expression in human fibroblasts after alpha-particle emitter (211)At compared with (60)Co irradiation.

    PubMed

    Danielsson, Anna; Claesson, Kristina; Parris, Toshima Z; Helou, Khalil; Nemes, Szilárd; Elmroth, Kecke; Elgqvist, Jörgen; Jensen, Holger; Hultborn, Ragnar

    2013-04-01

    The aim of this study was to identify gene expression profiles distinguishing alpha-particle (211)At and (60)Co irradiation. Gene expression microarray profiling was performed using total RNA from confluent human fibroblasts 5 hours after exposure to (211)At labeled trastuzumab monoclonal antibody (0.25, 0.5, and 1 Gy) and (60)Co (1, 2, and 3 Gy). We report gene expression profiles that distinguish the effect different radiation qualities and absorbed doses have on cellular functions in human fibroblasts. In addition, we identified commonly expressed transcripts between (211)At and (60)Co irradiation. A greater number of transcripts were modulated by (211)At than (60)Co irradiation. In addition, down-regulation was more prevalent than up-regulation following (211)At irradiation. Several biological processes were enriched for both irradiation qualities such as transcription, cell cycle regulation, and cell cycle arrest, whereas mitosis, spindle assembly checkpoint, and apoptotic chromosome condensation were uniquely enriched for alpha particle irradiation. LET-dependent transcriptional modulations were observed in human fibroblasts 5 hours after irradiation exposure. These findings suggest that in comparison with (60)Co, (211)At has the clearest influence on both tumor protein p53-activated and repressed genes, which impose a greater overall burden to the cell following alpha particle irradiation.

  14. [Protective effects of human bone marrow mesenchymal stem cells on hematopoietic organs of irradiated mice].

    PubMed

    Chen, Ling-Zhen; Yin, Song-Mei; Zhang, Xiao-Ling; Chen, Jia-Yu; Wei, Bo-Xiong; Zhan, Yu; Yu, Wei; Wu, Jin-Ming; Qu, Jia; Guo, Zi-Kuan

    2012-12-01

    The objective of this study was to explore the protective effects of human bone marrow mesenchymal stem cells (MSC) on hematopoietic organs of irradiated mice. Human bone marrow MSC were isolated, ex vivo expanded, and identified by cell biological tests. Female BALB/c mice were irradiated with (60)Co γ-ray at a single dose of 6 Gy, and received different doses of human MSC and MSC lysates or saline via tail veins. The survival of mice was record daily, and the femurs and spleens were harvested on day 9 and 16 for pathologic examination. The histological changes were observed and the cellularity was scored. The results showed that the estimated survival time of MSC- and MSC lysate-treated mice was comparable to that of controls. The hematopoiesis in the bone marrow of mice that received high-dose (5×10(6)) of MSC or MSC lysates was partially restored on day 9 and the capacity of hemopoietic tissue and cellularity scorings were significantly elevated as compared with that of controls (P < 0.05). Proliferative nudes were also obviously observed in the spleens of mice that received high-dose of MSC or MSC lysates on d 9 after irradiation. The histological structures of the spleen and bone marrow of the mice that received high-doses (5×10(6)) of MSC or MSC lysates were restored to normal, the cell proliferation displayed extraordinarily active. Further, the cellularity scores of the bone marrow were not significantly different between the high-dose MSC and MSC lysate-treated mice. It is concluded that the bone marrow MSC can promote the hematopoietic recovery of the irradiated mice, which probably is associated with the bioactive materials inherently existed in bone marrow cells.

  15. Lymphoblastoid interferon-alpha inhibits T cell proliferation and expression of eosinophil-activating cytokines.

    PubMed

    Krishnaswamy, G; Smith, J K; Srikanth, S; Chi, D S; Kalbfleisch, J H; Huang, S K

    1996-10-01

    T cell-derived cytokines, such as interleukin-5 (IL-5) and granulocyte-macrophage colony-stimulating factor (GM-CSF) activate eosinophils, whereas other cytokines, such as tumor necrosis factor (TNF)-alpha and IL-13, determine eosinophil recruitment. Interferon-alpha (IFN-alpha), a leukocyte-derived cytokine, has been shown to have beneficial effects in eosinophil-mediated disorders, such as the hypereosinophilic syndrome and a murine model of allergic asthma, where it inhibited eosinophil recruitment. We tested the hypothesis that IFN-alpha acted in eosinophil-mediated disorders by modulating T cell cytokine expression. Peripheral blood mononuclear cells (PBMC) or human ragweed-specific TH1 (2B8) and TH2 (2D2) T cell clones were cultured in the presence of 5 micrograms/ml of phytohemagglutinin (PHA) or 25 micrograms/ml of antigen Amb a 1 (short ragweed allergen), respectively, and lymphoblastoid IFN-alpha (varying from 0 to 10,000 U/ml). We assessed T cell proliferation by [3H]thymidine incorporation and production of IL-5 and GM-CSF by ELISA. Expression of cytokine transcripts was analyzed by the reverse transcription-polymerase chain reaction technique (RT-PCR). IFN-alpha induced a dose-dependent suppression of T cell proliferation of both PBMC (p < 0.001) and the T cell clones (p < 0.001). IFN-alpha inhibited gene expression of IL-5, GM-CSF, TNF-alpha, and IL-13 in PBMC. Furthermore, IFN-alpha significantly inhibited mitogen-induced and antigen-induced production of IL-5 and GM-CSF. IFN-alpha may benefit eosinophil-mediated disorders by inhibiting T cell function and production of cytokines active on human eosinophils.

  16. In vitro effects of helium-neon laser irradiation on human blood: blood viscosity and deformability of erythrocytes.

    PubMed

    Mi, Xian-Qiang; Chen, Ji-Yao; Liang, Zi-Jun; Zhou, Lu-Wei

    2004-12-01

    The purpose of this study was to investigate the in vitro effects of He-Ne laser irradiation on some rheological factors of human blood, such as blood viscosity, erythrocyte deformability, and sedimentation rate. The intravascular irradiation of low power laser has been applied in pre-clinical and clinical to treat various pathological processes. However, the mechanism is not fully understood so far. Especially the interaction and related mechanism between the laser and blood are unclear. In this work, by measuring the change of the main rheological factors after laser irradiation, the interaction and mechanism were explored. A30-mW He-Ne laser was used for irradiation with a 4-5-mm-diameter beam spot on blood samples, with a fluence rate of about 150 mW/cm.(2) The irradiation time was 60 min, so the total dose of irradiation was 540 J/cm.(2) The pathological samples of blood were obtained from patients (volunteers), and each sample was divided into two tubes for irradiation and control. The blood viscosity, erythrocyte deformability, and sedimentation rate were measured after laser irradiation and compared with un-irradiated control. The blood samples with poor erythrocyte deformability were prepared by adding Ca(2+) to the normal erythrocytes of a healthy person for investigating the laser effect on erythrocyte deformability further. Laser irradiation reduced the erythrocyte sedimentation rate of blood samples, which had a hyper-sedimentation rate originally. The blood viscosity of samples in hyper-values was lowered by laser irradiation in all shear rates measured (10-110 S(-1)), with a relative variation of approximately 10%. The deformability of erythrocytes from pathological samples and Ca(2+)-treated samples was improved after laser irradiation. The positive effects of laser irradiation on improving the rheological properties of blood were demonstrated in vitro.

  17. Irradiation With Carbon Ion Beams Induces Apoptosis, Autophagy, and Cellular Senescence in a Human Glioma-Derived Cell Line

    SciTech Connect

    Jinno-Oue, Atsushi; Shimizu, Nobuaki; Hamada, Nobuyuki; Wada, Seiichi; Tanaka, Atsushi; Shinagawa, Masahiko; Ohtsuki, Takahiro; Mori, Takahisa; Saha, Manujendra N.; Hoque, Ariful S.; Islam, Salequl; Kogure, Kimitaka; Funayama, Tomoo; Kobayashi, Yasuhiko

    2010-01-15

    Purpose: We examined biological responses of human glioma cells to irradiation with carbon ion beams (C-ions). Methods and Materials: A human glioma-derived cell line, NP-2, was irradiated with C-ions. Apoptotic cell nuclei were stained with Hoechst 33342. Induction of autophagy was examined either by staining cells with monodansylcadaverine (MDC) or by Western blotting to detect conversion of microtuble-associated protein light chain 3 (MAP-LC3) (LC3-I) to the membrane-bound form (LC3-II). Cellular senescence markers including induction of senescence-associated beta-galactosidase (SA-beta-gal) were examined. The mean telomere length of irradiated cells was determined by Southern blot hybridization. Expression of tumor suppressor p53 and cyclin/cyclin-dependent kinase inhibitor p21{sup WAF1/CIP1} in the irradiated cells was analyzed by Western blotting. Results: When NP-2 cells were irradiated with C-ions at 6 Gy, the major population of the cells died of apoptosis and autophagy. The residual fraction of attached cells (<1% of initially irradiated cells) could not form a colony: however, they showed a morphological phenotype consistent with cellular senescence, that is, enlarged and flattened appearance. The senescent nature of these attached cells was further indicated by staining for SA-beta-gal. The mean telomere length was not changed after irradiation with C-ions. Phosphorylation of p53 at serine 15 as well as the expression of p21{sup WAF1/CIP1} was induced in NP-2 cells after irradiation. Furthermore, we found that irradiation with C-ions induced cellular senescence in a human glioma cell line lacking functional p53. Conclusions: Irradiation with C-ions induced apoptosis, autophagy, and cellular senescence in human glioma cells.

  18. Apoptosis preferentially eliminates irradiated g0 human lymphocytes bearing dicentric chromosomes.

    PubMed

    Belloni, P; Meschini, R; Lewinska, D; Palitti, F

    2008-02-01

    G(0) human peripheral blood lymphocytes were X-irradiated to determine whether there is a direct relationship between radiation-induced dicentric chromosomes and the triggering of apoptosis. Immediately after X-ray exposure, control and irradiated lymphocytes were analyzed for viability, apoptosis and chromosome damage using the premature chromosome condensation technique. A batch of lymphocytes was kept in liquid holding for 48 h and then loaded on Ficoll-Paque medium to separate apoptotic (high-density) and normal (normal-density) cells. Then the same end points were analyzed in high-density and normal-density fractions of control and irradiated lymphocytes. After 48 h of liquid holding, the majority of apoptotic cells contained dicentric chromosomes. These results demonstrate that in human lymphocytes, the type of chromosome damage influences the induction of programmed cell death and provide direct evidence that cells bearing dicentrics are eliminated by apoptosis. G0 lymphocytes are the most common tissue used in biodosimetry studies, and the amount of chromosomal damage detected depends on the time between exposure and sampling. Since the radiation-induced apoptotic cells show the presence of dicentrics, radiation-induced damage can be underestimated. These results may have relevance in evaluations of the efficacy of radiotherapy based on the frequencies of chromosomal aberrations.

  19. A theoretical investigation of human skin thermal response to near-infrared laser irradiation

    NASA Astrophysics Data System (ADS)

    Dai, Tianhong; Pikkula, Brian M.; Wang, Lihong V.; Anvari, Bahman

    2004-07-01

    Near-infrared wavelengths are absorbed less by epidermal melanin mainly located at the basal layer of epidermis (dermo-epidermal junction), and penetrate deeper into human skin dermis and blood than visible wavelengths. Therefore, laser irradiation using near-infrared wavelength may improve the therapeutic outcome of cutaneous hyper-vascular malformations in moderately to heavily pigmented skin patients and those with large-sized blood vessels or blood vessels extending deeply into the skin. A mathematical model composed of a Monte Carlo algorithm to estimate the distribution of absorbed light followed by numerical solution of a bio-heat diffusion equation was utilized to investigate the thermal response of human skin to near-infrared laser irradiation, and compared it with that to visible laser irradiation. Additionally, the effect of skin surface cooling on epidermal protection was theoretically investigated. Simulation results indicated that 940 nm wavelength is superior to 810 and 1064 nm in terms of the ratio of light absorption by targeted blood vessel to the absorption by the basal layer of epidermis, and is more efficient than 595 nm wavelength for the treatment of patients with large-sized blood vessels and moderately to heavily pigmented skin. Dermal blood content has a considerable effect on the laser-induced peak temperature at the basal layer of epidermis, while the effect of blood vessel size is minimum.

  20. Cell and Molecular Biology of Ataxia Telangiectasia Heterozygous Human Mammary Epithelial Cells Irradiated in Culture

    NASA Technical Reports Server (NTRS)

    Richmond, Robert C.

    2001-01-01

    Autologous isolates of cell types from obligate heterozygotes with the autosomal disorder ataxia-telangiectasia (A-T)were used to begin a tissue culture model for assessing pathways of radiation-induced cancer formation in this target tissue. This was done by establishing cultures of stromal fibroblasts and long-term growth human mammary epithelial cells (HMEC) in standard 2-dimensional tissue culture in order to establish expression of markers detailing early steps of carcinogenesis. The presumptive breast cancer susceptibility of A-T heterozygotes as a sequel to damage caused by ionizing radiation provided reason to study expression of markers in irradiated HMEC. Findings from our study with HMEC have included determination of differences in specific protein expression amongst growth phase (e.g., log vs stationary) and growth progression (e.g., pass 7 vs pass 9), as well as differences in morphologic markers within populations of irradiated HMEC (e.g., development of multinucleated cells).

  1. Cell and Molecular Biology of Ataxia Telangiectasia Heterozygous Human Mammary Epithelial Cells Irradiated in Culture

    NASA Technical Reports Server (NTRS)

    Richmond, Robert C.

    2001-01-01

    Autologous isolates of cell types from obligate heterozygotes with the autosomal disorder ataxia-telangiectasia (A-T)were used to begin a tissue culture model for assessing pathways of radiation-induced cancer formation in this target tissue. This was done by establishing cultures of stromal fibroblasts and long-term growth human mammary epithelial cells (HMEC) in standard 2-dimensional tissue culture in order to establish expression of markers detailing early steps of carcinogenesis. The presumptive breast cancer susceptibility of A-T heterozygotes as a sequel to damage caused by ionizing radiation provided reason to study expression of markers in irradiated HMEC. Findings from our study with HMEC have included determination of differences in specific protein expression amongst growth phase (e.g., log vs stationary) and growth progression (e.g., pass 7 vs pass 9), as well as differences in morphologic markers within populations of irradiated HMEC (e.g., development of multinucleated cells).

  2. Protein degradation in a LAMP-2-deficient B-lymphoblastoid cell line from a patient with Danon disease.

    PubMed

    Sánchez-Lanzas, Raul; Alvarez-Castelao, Beatriz; Bermejo, Teresa; Ayuso, Teresa; Tuñón, Teresa; Castaño, José G

    2016-08-01

    Danon disease, a condition characterized by cardiomyopathy, myopathy, and intellectual disability, is caused by mutations in the LAMP-2 gene. Lamp-2A protein, generated by alternative splicing from the Lamp-2 pre-mRNA, is reported to be the lysosomal membrane receptor essential for the chaperone-mediated autophagic pathway (CMA) aimed to selective protein targeting and translocation into the lysosomal lumen for degradation. To study the relevance of Lamp-2 in protein degradation, a lymphoblastoid cell line was obtained by EBV transformation of B-cells from a Danon patient. The derived cell line showed no significant expression of Lamp-2 protein. The steady-state mRNA and protein levels of alpha-synuclein, IΚBα, Rcan1, and glyceraldehyde-3-phosphate dehydrogenase, four proteins reported to be selective substrates of the CMA pathway, were similar in control and Lamp-2-deficient cells. Inhibition of protein synthesis showed that the half-life of alpha-synuclein, IΚBα, and Rcan1 was similar in control and Lamp-2-deficient cells, and its degradation prevented by proteasome inhibitors. Both in control and Lamp-2-deficient cells, induction of CMA and macroautophagy by serum and aminoacid starvation of cells for 8h produced a similar decrease in IΚBα and Rcan1 protein levels and was prevented by the addition of lysosome and autophagy inhibitors. In conclusion, the results presented here showed that Lamp-2 deficiency in human lymphoblastoid cells did not modify the steady-state levels or the degradation of several protein substrates reported as selective substrates of the CMA pathway.

  3. Characteristic studies of non-homologous end joining in human cells irradiated with high LET radiation

    NASA Astrophysics Data System (ADS)

    Okayasu, R.; Okada, M.; Okabe, A.; Takakura, K.

    We studied the repair process of G0/G1 phase normal (HFL III) and non homologous end joining (NHEJ) deficient human fibroblasts (180 BR) exposed to X-rays and high LET carbon ions (70 keV/μ m) using a modified fusion-based premature chromosome condensation (PCC) technique. We have succeeded in increasing the sensitivity of the PCC method by adding a potent DNA double strand break repair inhibitor, wortmannin, during the incubation period of this assay. With x-ray exposure (2 Gy or less), the rejoining of G1 chromosome breaks in 180BR cells are significantly slower and less efficient than that in normal cells. On the other hand, the difference in rejoining kinetics between 180BR and normal cells with high LET carbon exposure is much smaller than that with x-ray exposure. These results seem to reflect the radiation cell survival responses using the same cell lines. We also studied the auto-phosphorylation status of DNA dependent protein kinase catalytic subunit (DNA-PKcs) protein in cells exposed to high and low LET radiation. Our immuno-staining results using an antibody to detect an auto-phosphorylation site of DNA-PKcs further reveal the difficulty in NHEJ for cells exposed to high LET radiation. The peak time for the auto-phosphorylation in x-irradiated normal human cells is one hour post-irradiation, but the peak in the same cells irradiated with high LET carbon beams shifted to two hours post-irradiation, reflecting much slower NHEJ processing associated with the high LET radiation. These data help understand the mechanism underlying the biological effect induced by heavy ion particles in the space environment.

  4. Low-fluence CO2 laser irradiation: selective epidermal damage to human skin.

    PubMed

    Kamat, B R; Tang, S V; Arndt, K A; Stern, R S; Noe, J M; Rosen, S

    1985-09-01

    The interaction of normal human skin with low-fluence CO2 laser irradiation was studied using a three-phase approach. In phase one, freshly excised skin was observed immediately after impact. In phase two, skin irradiated 2 h prior to excision was studied. In phase three, human volunteers were irradiated and biopsied at time zero, 24 h and 48 h. Seventy-five sites were exposed and 60 biopsies were performed. The earliest histologic changes were observed in the 6-10 J/cm2 fluence (radiant exposure) range and these changes included spindle and vacuolar changes in the basal layer of the epidermis. Papillary dermal coagulation was present to a maximum of 0.03 mm. At fluences of 10-25 J/cm2, superficial dermal necrosis (0.06-0.08 mm) was observed. At fluences above 25 J/cm2, transepidermal necrosis was present with increasing papillary dermal necrosis that was in proportion to the energy density delivered. At 2h, basal vacuolar changes were accompanied by diffuse keratinocytic cell death where contact was maintained between the epidermis and dermis, while where separation occurred limited keratinocytic death was observed. The earliest changes occurred at lower threshold fluences (4-6 J/cm2). After 24 h, these doses resulted in extensive epidermal necrosis with focal acute inflammatory infiltrates. At 48 h, the degree of epidermal "slough" was proportional to the energy density delivered and was maximal with a fluence of 5.7 J/cm2 delivered whereas with a fluence of 3.8 J/cm2 thin slough (0.02 mm) was observed. These findings suggest that low-dose CO2 laser irradiation may provide a new approach to selectively damage the epidermis with minimal dermal damage.

  5. Utilization of Lymphoblastoid Cell Lines as a System for the Molecular Modeling of Autism

    ERIC Educational Resources Information Center

    Baron, Colin A.; Liu, Stephenie Y.; Hicks, Chindo; Gregg, Jeffrey P.

    2006-01-01

    In order to provide an alternative approach for understanding the biology and genetics of autism, we performed statistical analysis of gene expression profiles of lymphoblastoid cell lines derived from children with autism and their families. The goal was to assess the feasibility of using this model in identifying autism-associated genes.…

  6. Utilization of Lymphoblastoid Cell Lines as a System for the Molecular Modeling of Autism

    ERIC Educational Resources Information Center

    Baron, Colin A.; Liu, Stephenie Y.; Hicks, Chindo; Gregg, Jeffrey P.

    2006-01-01

    In order to provide an alternative approach for understanding the biology and genetics of autism, we performed statistical analysis of gene expression profiles of lymphoblastoid cell lines derived from children with autism and their families. The goal was to assess the feasibility of using this model in identifying autism-associated genes.…

  7. Biscoclaurine alkaloid cepharanthine protects DNA in TK6 lymphoblastoid cells from constitutive oxidative damage

    PubMed Central

    Halicka, H. Dorota; Ita, Masamichi; Tanaka, Toshiki; Kurose, Akira; Darzynkiewicz, Zbigniew

    2008-01-01

    Cepharanthine (CEP), a biscoclaurine (bisbenzylisoquinoline) alkaloid isolated from Stephania cepharantha Hayata, is widely used in Japan to treat variety of diseases. Among a plethora of its biological activities CEP was reported to be able to scavenge radicals and prevent lipid peroxidation. We have recently described the phenomenon of constitutive ATM activation (CAA) and histone H2AX phosphorylation (CHP), the events that report DNA damage induced by endogenously generated radicals, the product of oxidative metabolism in otherwise healthy, untreated cells. The aim of the present study was to explore whether CEP can attenuate the level of CAA and CHP, which would indicate on its ability to protect DNA against endogenous oxidants. The data show that indeed the levels of CAA and CHP in human lymphoblastoid TK6 cells were distinctly lowered upon treatment with CEP. Thus, exposure of cells to 8.3 μM CEP for 4 h led to a reduction of the mean level of CAA and CHP by up to 60% and 50%, respectively. At 1.7 μM CEP the reduction of CAA and CHP after 4 h was 35% and 25%, respectively. Cells exposure to CEP led to a decrease in the level of ondogenous oxidants as measured by the ability to oxidate the fluorescent probe 5-(and-6)-carboxy-2′,7′-dichlorodihydro-fluorescein diacetate. No evidence of apoptosis was seen during the first 8 h of treatment with CEP but initiation of apoptosis (caspase-3 activation) was detected in relatively few (< 10%) cells after exposure to 8.3 μM CEP for 24 h. The data strongly suggest that the scavenging properties of CEP provide a protection of DNA from the radicals generated endogenously during oxidative metabolism. PMID:18276990

  8. Forward subtractive libraries containing genes transactivated by dexamethasone in ataxia-telangiectasia lymphoblastoid cells.

    PubMed

    Biagiotti, Sara; Menotta, Michele; Giacomini, Elisa; Radici, Lucia; Bianchi, Marzia; Bozzao, Cristina; Chessa, Luciana; Magnani, Mauro

    2014-07-01

    Ataxia telangiectasia (A-T) is a rare autosomal recessive disorder caused by biallelic mutations in the Ataxia Telangiectasia-mutated gene. A-T shows a complex phenotype ranging from early-onset progressive neurodegeneration to immunodeficiencies, high incidence of infections, and tumors. Unfortunately, no therapy is up to now available for treating this condition. Recently, the short term treatment of ataxia-telangiectasia patients with glucocorticoids was shown to improve their neurological symptoms and possibly reverse cerebellar atrophy. Thus, corticosteroids represent an attractive approach for the treatment of this neurodegenerative disease. However, the molecular mechanism involved in glucocorticoid action in A-T is yet unknown. The aim of our work is to construct cDNA libraries containing those genes which are transactivated by the glucocorticoid analogue, dexamethasone, in A-T human cells. For this purpose, suppression subtractive hybridization has been performed on ATM-null lymphoblastoid cell transcriptome extracted following drug administration. Annotation of whole genes contained in the libraries has been obtained by coupling subtractive hybridization with microarray analysis. Positive transcripts have been validated by quantitative PCR. Through in silico analyses, identified genes have been classified on the basis of the pathway in which they are involved, being able to address signaling required for dexamethasone action. Most of the induced transcripts are involved in metabolic processes and regulation of cellular processes. Our results can help to unravel the mechanism of glucocorticoid action in the reversion of A-T phenotype. Moreover, the induction of a specific region of the ATM transcript has been identified as putative biomarker predictive of dexamethasone efficacy on ataxic patients.

  9. The differential role of human macrophage in triggering secondary bystander effects after either gamma-ray or carbon beam irradiation

    PubMed Central

    Dong, Chen; He, Mingyuan; Tu, Wenzhi; Konishi, Teruaki; Liu, Weili; Xie, Yuexia; Dang, Bingrong; Li, Wenjian; Uchihori, Yukio; Hei, Tom K.; Shao, Chunlin

    2015-01-01

    The abscopal effect could be an underlying factor in evaluating prognosis of radiotherapy. This study established an in vitro system to examine whether tumor-generated bystander signals could be transmitted by macrophages to further trigger secondary cellular responses after different irradiations, where human lung cancer NCI-H446 cells were irradiated with either γ-rays or carbon ions and co-cultured with human macrophage U937 cells, then these U937 cells were used as a bystander signal transmitter and co-cultured with human bronchial epithelial cells BEAS-2B. Results showed that U937 cells were only activated by γ-irradiated NCI-H446 cells so that the secondary injuries in BEAS-2B cells under carbon ion irradiation were weaker than γ-rays. Both TNF-α and IL-1α were involved in γ-irradiation induced secondary bystander effect but only TNF-α contributed to the carbon ion induced response. Further assay disclosed that IL-1α but not TNF-α was largely responsible for the activation of macrophages and the formation of micronucleus in BEAS-2B cells. These data suggest that macrophages could transfer secondary bystander signals and play a key role in the secondary bystander effect of photon irradiation while carbon ion irradiation has conspicuous advantage due to its reduced secondary injury. PMID:25896631

  10. The differential role of human macrophage in triggering secondary bystander effects after either gamma-ray or carbon beam irradiation.

    PubMed

    Dong, Chen; He, Mingyuan; Tu, Wenzhi; Konishi, Teruaki; Liu, Weili; Xie, Yuexia; Dang, Bingrong; Li, Wenjian; Uchihori, Yukio; Hei, Tom K; Shao, Chunlin

    2015-07-10

    The abscopal effect could be an underlying factor in evaluating prognosis of radiotherapy. This study established an in vitro system to examine whether tumor-generated bystander signals could be transmitted by macrophages to further trigger secondary cellular responses after different irradiations, where human lung cancer NCI-H446 cells were irradiated with either γ-rays or carbon ions and co-cultured with human macrophage U937 cells, then these U937 cells were used as a bystander signal transmitter and co-cultured with human bronchial epithelial cells BEAS-2B. Results showed that U937 cells were only activated by γ-irradiated NCI-H446 cells so that the secondary injuries in BEAS-2B cells under carbon ion irradiation were weaker than γ-rays. Both TNF-α and IL-1α were involved in the γ-irradiation induced secondary bystander effect but only TNF-α contributed to the carbon ion induced response. Further assay disclosed that IL-1α but not TNF-α was largely responsible for the activation of macrophages and the formation of micronucleus in BEAS-2B cells. These data suggest that macrophages could transfer secondary bystander signals and play a key role in the secondary bystander effect of photon irradiation, while carbon ion irradiation has conspicuous advantage due to its reduced secondary injury. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  11. Molecular and cytogenetic analysis of lymphoblastoid and colon cancer cell lines from cotton-top tamarin (Sagiunus oedipus).

    PubMed

    Mao, X; McGuire, S; Hamoudi, R A

    2000-07-01

    The cotton-top tamarin (CTT) (Sagiunus oedipus) has been used as an animal model to investigate the etiology and pathophysiology of several human diseases, including ulcerative colitis and its associated colorectal carcinoma (CRC). Little is known, however, about genetic synteny between CTT and humans, and about chromosome aberrations in CTT CRC. To address these issues, we have analyzed CTT lymphoblastoid and CRC cell lines using cytogenetics, fluorescence in situ hybridization (Zoo-FISH), and direct sequencing. The CTT lymphocytes had pseudodiploid chromosomes of 46. The CTT CRC cells showed near-diploid chromosomes of 45. Several clonal structural aberrations were observed, including der(1), a marker chromosome, and double minutes. Zoo-FISH using human chromosome 2, 3, 5, 6, 9, 11, 13, 15, 16, 17, 19, 22, and X paints identified homologous chromosomes and subchromosomal regions in the CTT genome. Fluorescence in situ hybridization with human telomeric probe also detected a homologous sequence in CTT genome. Direct sequencing of CTT genomic DNA using primers amplifying exons 4 and 15 of the human APC gene identified DNA sequences in CTT genome with 99% and 95% homology, respectively. These results provide a basis for further comparative studies of CTT and human genome.

  12. The effect of 648 nm diode laser irradiation on second messengers in senescent human keratinocytes

    NASA Astrophysics Data System (ADS)

    Hawkins Evans, D.; Abrahamse, H.

    2009-02-01

    Background/purpose: Stress induced premature senescence (SIPS) is defined as the long-term effect of subcytotoxic stress on proliferative cell types. Cells in SIPS display differences at the level of protein expression which affect energy metabolism, defense systems, redox potential, cell morphology and transduction pathways. This study aimed to determine the effect of laser irradiation on second messengers in senescent cells and to establish if that effect can be directly linked to changes in cellular function such as cell viability or proliferation. Materials and Methods: Human keratinocyte cell cultures were modified to induce premature senescence using repeated sub-lethal stresses of 200 uM H2O2 or 5% OH every day for four days with two days recovery. SIPS was confirmed by senescence-associated β-galactosidase staining. Control conditions included normal, repeated stress of 500 uM H2O2 to induce apoptosis and 200 uM PBN as an anti-oxidant or free radical scavenger. Cells were irradiated with 1.5 J/cm2 on day 1 and 4 using a 648 nm diode laser (3.3 mW/cm2) and cellular responses were measured 1 h post irradiation. The affect on second messengers was assessed by measuring cAMP, cGMP, nitric oxide and intracellular calcium (Ca2+) while functional changes were assessed using cell morphology, ATP cell viability, LDH membrane integrity and WST-1 cell proliferation. Results: Results indicate an increase in NO and a decrease in cGMP and Ca2+ in 200 uM H2O2 irradiated cells while PBN irradiated cells showed a decrease in cAMP and an increase in ATP viability and cell proliferation. Conclusion: Laser irradiation influences cell signaling which ultimately changes the biological function of senescent cells. If laser therapy can stimulate the biological function of senescent cells it may be beneficial to conditions such as immune senescence, skin ageing, muscle atrophy, premature ageing of arteries in patients with advanced heart disease, neurodegenerative disorders and

  13. Cellular and molecular portrait of eleven human glioblastoma cell lines under photon and carbon ion irradiation.

    PubMed

    Ferrandon, S; Magné, N; Battiston-Montagne, P; Hau-Desbat, N-H; Diaz, O; Beuve, M; Constanzo, J; Chargari, C; Poncet, D; Chautard, E; Ardail, D; Alphonse, G; Rodriguez-Lafrasse, C

    2015-04-28

    This study aimed to examine the cellular and molecular long-term responses of glioblastomas to radiotherapy and hadrontherapy in order to better understand the biological effects of carbon beams in cancer treatment. Eleven human glioblastoma cell lines, displaying gradual radiosensitivity, were irradiated with photons or carbon ions. Independently of p53 or O(6)-methylguanine-DNA methyltransferase(1) status, all cell lines responded to irradiation by a G2/M phase arrest followed by the appearance of mitotic catastrophe, which was concluded by a ceramide-dependent-apoptotic cell death. Statistical analysis demonstrated that: (i) the SF2(2) and the D10(3) values for photon are correlated with that obtained in response to carbon ions; (ii) regardless of the p53, MGMT status, and radiosensitivity, the release of ceramide is associated with the induction of late apoptosis; and (iii) the appearance of polyploid cells after photon irradiation could predict the Relative Biological Efficiency(4) to carbon ions. This large collection of data should increase our knowledge in glioblastoma radiobiology in order to better understand, and to later individualize, appropriate radiotherapy treatment for patients who are good candidates.

  14. Thermal neutron irradiation field design for boron neutron capture therapy of human explanted liver.

    PubMed

    Bortolussi, S; Altieri, S

    2007-12-01

    The selective uptake of boron by tumors compared to that by healthy tissue makes boron neutron capture therapy (BNCT) an extremely advantageous technique for the treatment of tumors that affect a whole vital organ. An example is represented by colon adenocarcinoma metastases invading the liver, often resulting in a fatal outcome, even if surgical resection of the primary tumor is successful. BNCT can be performed by irradiating the explanted organ in a suitable neutron field. In the thermal column of the Triga Mark II reactor at Pavia University, a facility was created for this purpose and used for the irradiation of explanted human livers. The neutron field distribution inside the organ was studied both experimentally and by means of the Monte Carlo N-particle transport code (MCNP). The liver was modeled as a spherical segment in MCNP and a hepatic-equivalent solution was used as an experimental phantom. In the as-built facility, the ratio between maximum and minimum flux values inside the phantom ((phi(max)/phi(min)) was 3.8; this value can be lowered to 2.3 by rotating the liver during the irradiation. In this study, the authors proposed a new facility configuration to achieve a uniform thermal neutron flux distribution in the liver. They showed that a phi(max)/phi(min) ratio of 1.4 could be obtained without the need for organ rotation. Flux distributions and dose volume histograms were reported for different graphite configurations.

  15. Raman spectroscopic studies of CO2 laser-irradiated human dental enamel

    NASA Astrophysics Data System (ADS)

    Aminzadeh, A.; Shahabi, S.; Walsh, L. J.

    1999-06-01

    While the effects of carbon dioxide (CO2) laser radiation on the physical properties of human dental enamel are well characterized, little is known regarding laser-induced chemical changes. In this study, enamel was exposed to CO2 laser radiation to induce fusion and recrystallization, and the Raman spectra recorded using both dispersive and Fourier-transformed (FT) Raman spectroscopy. Spectra were compared to a heat-treated specimen of hydroxyapatite (HAP) and enamel. Laser irradiation induced chemical changes which differed from those induced by heat treatment. Comparing the Raman spectra of lased enamel to HAP and tricalcium phosphate (TCP), it is evident that CO2 laser irradiation of enamel causes the partial conversion of HAP to TCP. The effect of laser irradiation is not merely a simple local heating effect as previously thought, since simple heating of enamel leads to the formation of both TCP and Ca(OH)2, while laser treatment of enamel results in the formation of TCP but not Ca(OH)2.

  16. Helium-neon laser irradiation enhances DNA synthesis in a human neuroblastoma cell line

    NASA Astrophysics Data System (ADS)

    Condon, Michael R.; Gump, Frank; Wu, Wen-hsien

    1993-07-01

    To gain further insight into the mechanism of cell photostimulation by laser light ((lambda) equals 632.8 nm), DNA synthesis was measured in the human neuroblastoma cell line BE(2)-C. Cells were irradiated at high density to establish the characteristics of cellular energy into S- phase in response to laser stimulation. BE(2)-C cells after release from a quiescent, growth arrested state exhibited increased incorporation of isotope 12 hours after replating at subconfluent density in the presence of 2.5% fetal bovine serum (FBS) and [3H] thymidine. In contrast, cells replated under the same conditions, but stimulated with 15% FBS exhibited a time lag of approximately 16 hours in apparent DNA synthesis. These results were not corroborated by flow cytometry. Laser irradiation did not affect the fraction of cells entering S-phase. It therefore appears that the stimulatory effect of He-Ne laser irradiation on BE(2)-C cells is to enhance DNA synthesis while not altering the G1-S transition rate.

  17. Caffeine enhanced measurement of mutagenesis by low levels of [gamma]-irradiation in human lymphocytes

    SciTech Connect

    Puck, T.P.; Johnson, R.; Waldren, C.A. ); Morse, H. )

    1993-09-01

    The well-known action of caffeine in synergizing mutagenesis (including chromosome aberrations) of agents like ionizing radiation by inhibition of cellular repair processes has been incorporated into a rapid procedure for detection of mutagenicity with high sensitivity. Effects of 5-10 rads of [gamma]-irradiation, which approximate the human lifetime dose accumulation from background radiation, can be detected in a two-day procedure using an immortalized human WBC culture. Chromosomally visible lesions are scored on cells incubated for 2 h after irradiation in the presence and absence of 1.0 mg/ml of caffeine. An eightfold amplification of scorable lesions is achieved over the action of radiation alone. This approach provides a closer approximation to absolute mutagenicity unmitigated by repair processes, which can vary in different situations. It is proposed that mutagenesis testing of this kind, using caffiene or other repair-inhibitory agents, be employed to identify mutagens in their effective concentrations to which human populations may be exposed; to detect agents such as caffeine that may synergize mutagenic actions and pose epidemiologic threats; and to discover effective anti-mutagens. Information derived from the use of such procedures may help prevent cancer and newly acquired genetic disease.

  18. Atomic force microscopy analysis of human cornea surface after UV (λ=266 nm) laser irradiation

    NASA Astrophysics Data System (ADS)

    Spyratou, E.; Makropoulou, M.; Moutsouris, K.; Bacharis, C.; Serafetinides, A. A.

    2009-07-01

    Efficient cornea reshaping by laser irradiation for correcting refractive errors is still a major issue of interest and study. Although the excimer laser wavelength of 193 nm is generally recognized as successful in ablating corneal tissue for myopia correction, complications in excimer refractive surgery leads to alternative laser sources and methods for efficient cornea treatment. In this work, ablation experiments of human donor cornea flaps were conducted with the 4th harmonic of an Nd:YAG laser, with different laser pulses. AFM analysis was performed for examination of the ablated cornea flap morphology and surface roughness.

  19. Microwave irradiation of human brain tissue: production of microscopic slides within one day.

    PubMed Central

    Boon, M E; Marani, E; Adriolo, P J; Steffelaar, J W; Bots, G T; Kok, L P

    1988-01-01

    A three step method using microwave irradiation enabled microscopic slides of human brain tissue to be obtained within one working day: steps 1 and 2 hardened and solidified brain tissue; step 3 completed formalin fixation. The efficacy and precision of the method was compared with slides of conventionally processed brain tissue that had been fixed in formalin for six weeks. The microscopic quality of the sections was excellent with good presentation of brain tissue and equalled that of conventionally processed slides. Images Fig 1 Fig 2 Fig 3 PMID:3290268

  20. Shuttling of the autoantigen La between nucleus and cell surface after uv irradiation of human keratinocytes

    SciTech Connect

    Bachmann, M.; Chang, S.; Slor, H.; Kukulies, J.; Mueller, W.E. )

    1990-12-01

    During the past years we have established that the nuclear autoantigen La shuttles between the nucleus and the cytoplasm in tumor cells after inhibition of transcription or virus infection. We reinvestigated this shuttling using primary human keratinocytes from both healthy donors and patients with xeroderma pigmentosum. Ultraviolet irradiation resulted in both an inhibition of transcription and a translocation of La protein from the nucleus to the cytoplasm. After a prolonged inhibition of transcription La protein relocated into the nucleus and assembled with nuclear storage regions. The uv-induced shuttling included a translocation to the cell surface, where La protein colocalized with epidermal growth factor receptors.

  1. Chemical and morphological changes in human dentin after Er:YAGlaser irradiation: EDS and SEM analysis.

    PubMed

    Contreras-Arriaga, Belinda; Rodríguez-Vilchis, Laura Emma; Contreras-Bulnes, Rosalía; Olea-Mejìa, Oscar Fernando; Scougall-Vilchis, Rogelio José; Centeno-Pedraza, Claudia

    2015-11-01

    Sixty samples of human dentin were divided into six groups (n = 10) and were irradiated with Er:YAG laser at 100 mJ-19.9 J/cm(2), 150 mJ-29.8 J/cm(2), 100 mJ-35.3 J/cm(2), 150 mJ-53.0 J/cm(2), 200 mJ-70.7 J/cm(2), and 250 mJ-88.5 J/cm(2), respectively, at 7 Hz under a water spray. The atomic percentages of carbon, oxygen, magnesium, calcium, and phosphorus and the Ca-to-P molar ratio on the dentin were determined by energy dispersive X-ray spectroscopy. The morphological changes were observed using scanning electron microscopy. A paired t-test was used in statistical analysis before and after irradiation, and a one-way ANOVA was performed (P ≤ 0.05). The atomic percent of C tended to decrease in all of the groups after irradiation with statistically significant differences, O and Mg increased with significant differences in all of the groups, and the Ca-to-P molar ratio increased in groups IV, V, and VI, with statistically significant differences between groups II and VI. All the irradiated samples showed morphological changes. Major changes in the chemical composition of dentin were observed in trace elements. A significant increase in the Ca-to-P ratio was observed in the higher energy density groups. Morphological changes included loss of smear layer with exposed dentinal tubules. The changes produced by the different energy densities employed could have clinical implications, additional studies are required to clarify them. © 2015 Wiley Periodicals, Inc.

  2. Factors affecting ultraviolet-A photon emission from β-irradiated human keratinocyte cells.

    PubMed

    Le, M; Mothersill, C E; Seymour, C B; Ahmad, S B; Armstrong, A; Rainbow, A J; McNeill, F E

    2015-08-21

    The luminescence intensity of 340±5 nm photons emitted from HaCaT (human keratinocyte) cells was investigated using a single-photon-counting system during cellular exposure to (90)Y β-particles. Multiple factors were assessed to determine their influence upon the quantity and pattern of photon emission from β-irradiated cells. Exposure of 1 x 10(4) cells/5 mL to 703 μCi resulted in maximum UVA photoemission at 44.8 x 10(3)±2.5 x 10(3) counts per second (cps) from live HaCaT cells (background: 1-5 cps); a 16-fold increase above cell-free controls. Significant biophoton emission was achieved only upon stimulation and was also dependent upon presence of cells. UVA luminescence was measured for (90)Y activities 14 to 703 μCi where a positive relationship between photoemission and (90)Y activity was observed. Irradiation of live HaCaT cells plated at various densities produced a distinct pattern of emission whereby luminescence increased up to a maximum at 1 x 10(4) cells/5 mL and thereafter decreased. However, this result was not observed in the dead cell population. Both live and dead HaCaT cells were irradiated and were found to demonstrate different rates of photon emission at low β activities (⩽400 μCi). Dead cells exhibited greater photon emission rates than live cells which may be attributable to metabolic processes taking place to modulate the photoemissive effect. The results indicate that photon emission from HaCaT cells is perturbed by external stimulation, is dependent upon the activity of radiation delivered, the density of irradiated cells, and cell viability. It is postulated that biophoton emission may be modulated by a biological or metabolic process.

  3. Structural and Morphological Changes in Human Dentin after Ablative and Subablative Er:YAG Laser Irradiation

    PubMed Central

    Moosavi, Horieh; Ghorbanzadeh, Sajedeh; Ahrari, Farzaneh

    2016-01-01

    Introduction: This study investigated the influence of Erbium-Doped Yttrium Aluminum Garnet (Er:YAG) laser on microhardness, chemical composition and subsurface morphology of dentin cavity walls. Methods: Forty sound human premolars were selected and randomly assigned into four groups. Class V cavities were prepared either with an Er:YAG laser (groups 1 and 2; 15 Hz, 250 mJ for enamel, 10 Hz, 200 mJ for dentin) or with a high speed handpiece (groups 3 and 4). The specimens in groups 1 and 3 served as the control, whereas those in groups 2 and 4 were exposed to subablative laser irradiation following cavity preparation (10 Hz, 50 mJ). After bisecting the specimens, one half was subjected to microhardness assessment and the other half was evaluated by SEM-EDS analysis. Results: Microhardness was significantly greater in the specimens prepared by both ablative and subablative laser irradiation (group 2) than that of the bur-prepared cavities (groups 3 and 4) (P < 0.05). The quantity of calcium ion was significantly greater in cavities prepared by the Er:YAG laser (groups 1 and 2) compared to that of the bur cavities (groups 3 and 4) (P < 0.05). Subablative irradiation improved microhardness and weight percentage of calcium ion in both laser and bur cavities, but the difference was not significant compared to that of the relevant control group (P > 0.05). Conclusion: Cavity preparation with an Er:YAG laser could be considered as an alternative to the conventional method of drilling, as it enhances the mechanical and compositional properties of lased dentin, especially when combined by subablative irradiation. PMID:27330703

  4. Late ophthalmological complications after total body irradiation in non-human primates

    NASA Technical Reports Server (NTRS)

    Niemer-Tucker, M. M.; Sterk, C. C.; de Wolff-Rouendaal, D.; Lee, A. C.; Lett, J. T.; Cox, A.; Emmanouilidis-van der Spek, K.; Davelaar, J.; Lambooy, A. C.; Mooy, C. M.; Broerse, J. J.

    1999-01-01

    PURPOSE: To investigate the long-term effects of total body irradiation (TBI) on the incidence and time course of ocular complications. MATERIALS AND METHODS: Rhesus monkeys treated with TBI photon doses up to 8.5 Gy and proton doses up to 7.5 Gy were studied at intervals up to 25 years post-irradiation. They were compared with control groups with a similar age distribution. Cataract formation and ocular fundus lesions were scored according to a standardized protocol. Fluorescein angiography and histopathology was performed in selected animals. RESULTS: Cataract formation occurred after a latent period of 3-5 years. Significant cataract induction was observed for photon-doses of 8 and 8.5 Gy and beyond 20 years after proton irradiation. The severity of the lesions represents significant impairment of vision and would require cataract surgery if similar results occurred in human bone marrow transplant patients. Fluorescein angiography demonstrated a normal pattern of retinal vessels in 13 out of 14 animals (93%) from the irradiated group and in eight out of nine animals (89%) from the control group. No additional lesions apart from age-related degenerative changes could be demonstrated. Histological evaluation revealed no radiation-associated vasculopathy. CONCLUSIONS: Radiation alone for doses up to 8.5 Gy of photons does not carry a potential risk for fundus pathology, whereas clinically important cataract induction should be anticipated within 5 years after photon doses of 8.0 and 8.5 Gy and proton doses in excess of 2.5 Gy.

  5. Late ophthalmological complications after total body irradiation in non-human primates

    NASA Technical Reports Server (NTRS)

    Niemer-Tucker, M. M.; Sterk, C. C.; de Wolff-Rouendaal, D.; Lee, A. C.; Lett, J. T.; Cox, A.; Emmanouilidis-van der Spek, K.; Davelaar, J.; Lambooy, A. C.; Mooy, C. M.; hide

    1999-01-01

    PURPOSE: To investigate the long-term effects of total body irradiation (TBI) on the incidence and time course of ocular complications. MATERIALS AND METHODS: Rhesus monkeys treated with TBI photon doses up to 8.5 Gy and proton doses up to 7.5 Gy were studied at intervals up to 25 years post-irradiation. They were compared with control groups with a similar age distribution. Cataract formation and ocular fundus lesions were scored according to a standardized protocol. Fluorescein angiography and histopathology was performed in selected animals. RESULTS: Cataract formation occurred after a latent period of 3-5 years. Significant cataract induction was observed for photon-doses of 8 and 8.5 Gy and beyond 20 years after proton irradiation. The severity of the lesions represents significant impairment of vision and would require cataract surgery if similar results occurred in human bone marrow transplant patients. Fluorescein angiography demonstrated a normal pattern of retinal vessels in 13 out of 14 animals (93%) from the irradiated group and in eight out of nine animals (89%) from the control group. No additional lesions apart from age-related degenerative changes could be demonstrated. Histological evaluation revealed no radiation-associated vasculopathy. CONCLUSIONS: Radiation alone for doses up to 8.5 Gy of photons does not carry a potential risk for fundus pathology, whereas clinically important cataract induction should be anticipated within 5 years after photon doses of 8.0 and 8.5 Gy and proton doses in excess of 2.5 Gy.

  6. Factors affecting ultraviolet-A photon emission from β-irradiated human keratinocyte cells

    NASA Astrophysics Data System (ADS)

    Le, M.; Mothersill, C. E.; Seymour, C. B.; Ahmad, S. B.; Armstrong, A.; Rainbow, A. J.; McNeill, F. E.

    2015-08-01

    The luminescence intensity of 340+/- 5 nm photons emitted from HaCaT (human keratinocyte) cells was investigated using a single-photon-counting system during cellular exposure to 90Y β-particles. Multiple factors were assessed to determine their influence upon the quantity and pattern of photon emission from β-irradiated cells. Exposure of 1× {{10}4} cells/5 mL to 703 μCi resulted in maximum UVA photoemission at 44.8× {{10}3}+/- 2.5× {{10}3} counts per second (cps) from live HaCaT cells (background: 1-5 cps); a 16-fold increase above cell-free controls. Significant biophoton emission was achieved only upon stimulation and was also dependent upon presence of cells. UVA luminescence was measured for 90Y activities 14 to 703 μCi where a positive relationship between photoemission and 90Y activity was observed. Irradiation of live HaCaT cells plated at various densities produced a distinct pattern of emission whereby luminescence increased up to a maximum at 1× {{10}4} cells/5 mL and thereafter decreased. However, this result was not observed in the dead cell population. Both live and dead HaCaT cells were irradiated and were found to demonstrate different rates of photon emission at low β activities (⩽400 μCi). Dead cells exhibited greater photon emission rates than live cells which may be attributable to metabolic processes taking place to modulate the photoemissive effect. The results indicate that photon emission from HaCaT cells is perturbed by external stimulation, is dependent upon the activity of radiation delivered, the density of irradiated cells, and cell viability. It is postulated that biophoton emission may be modulated by a biological or metabolic process.

  7. High-frequency low-level diode laser irradiation promotes proliferation and migration of primary cultured human gingival epithelial cells.

    PubMed

    Ejiri, Kenichiro; Aoki, Akira; Yamaguchi, Yoko; Ohshima, Mitsuhiro; Izumi, Yuichi

    2014-07-01

    In periodontal therapy, the use of low-level diode lasers has recently been considered to improve wound healing of the gingival tissue. However, its effects on human gingival epithelial cells (HGECs) remain unknown. The aim of the present study was to examine whether high-frequency low-level diode laser irradiation stimulates key cell responses in wound healing, proliferation and migration, in primary cultured HGECs in vitro. HGECs were derived from seven independent gingival tissue specimens. Cultured HGECs were exposed to a single session of high-frequency (30 kHz) low-level diode laser irradiation with various irradiation time periods (fluence 5.7-56.7 J/cm(2)). After 20-24 h, cell proliferation was evaluated by WST-8 assay and [(3)H]thymidine incorporation assay, and cell migration was monitored by in vitro wound healing assay. Further, phosphorylation of the mitogen-activated protein kinase (MAPK) pathways after irradiation was investigated by Western blotting. The high-frequency low-level irradiation significantly increased cell proliferation and [(3)H]thymidine incorporation at various irradiation time periods. Migration of the irradiated cells was significantly accelerated compared with the nonirradiated control. Further, the low-level diode laser irradiation induced phosphorylation of MAPK/extracellular signal-regulated protein kinase (ERK) at 5, 15, 60, and 120 min after irradiation. Stress-activated protein kinases/c-Jun N-terminal kinase and p38 MAPK remained un-phosphorylated. The results show that high-frequency low-level diode laser irradiation promotes HGEC proliferation and migration in association with the activation of MAPK/ERK, suggesting that laser irradiation may accelerate gingival wound healing.

  8. Exploratory study of the prognostic value of microenvironmental parameters during fractionated irradiation in human squamous cell carcinoma xenografts.

    PubMed

    Yaromina, Ala; Kroeber, Theresa; Meinzer, Andreas; Boeke, Simon; Thames, Howard; Baumann, Michael; Zips, Daniel

    2011-07-15

    To explore the prognostic value of microenvironmental parameters for local tumor control determined before and during fractionated irradiation. Six human squamous cell carcinoma (hSCC) lines were transplanted subcutaneously into the right hind leg of nude mice. Tumors were irradiated with 30 fractions within 6 weeks. Local tumor control was determined 120 days after irradiation. Radiation response was quantified as dose to cure 50% of tumors (TCD(50)). In parallel, untreated and irradiated tumors were excised after injection of pimonidazole (hypoxia marker) and Hoechst 33342 (perfusion marker) for histological evaluation. Pimonidazole hypoxia decreased during fractionated irradiation in the majority of tumor lines. Fraction of perfused vessels and vascular area showed modest changes during fractionated irradiation. Histological parameters before treatment and after three and five fractions did not significantly correlate with TCD(50) after irradiation with 30 fractions within 6 weeks (p > 0.05). Hypoxic volume and perfused vessels after 10 fractions showed a significant association with local tumor control after fractionated irradiation (p = 0.018 and p = 0.019, respectively). None of these parameters remained statistically significant when the p value was adjusted for multiple comparisons. The results from this exploratory study suggest that determination of microenvironmental parameters during treatment provides better prognostic information for the outcome after fractionated radiotherapy than pretreatment parameters, which warrants further investigation and confirmation in experimental and clinical studies. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. Exploratory Study of the Prognostic Value of Microenvironmental Parameters During Fractionated Irradiation in Human Squamous Cell Carcinoma Xenografts

    SciTech Connect

    Yaromina, Ala; Kroeber, Theresa; Meinzer, Andreas; Boeke, Simon; Thames, Howard; Baumann, Michael; Zips, Daniel

    2011-07-15

    Purpose: To explore the prognostic value of microenvironmental parameters for local tumor control determined before and during fractionated irradiation. Methods and Materials: Six human squamous cell carcinoma (hSCC) lines were transplanted subcutaneously into the right hind leg of nude mice. Tumors were irradiated with 30 fractions within 6 weeks. Local tumor control was determined 120 days after irradiation. Radiation response was quantified as dose to cure 50% of tumors (TCD{sub 50}). In parallel, untreated and irradiated tumors were excised after injection of pimonidazole (hypoxia marker) and Hoechst 33342 (perfusion marker) for histological evaluation. Results: Pimonidazole hypoxia decreased during fractionated irradiation in the majority of tumor lines. Fraction of perfused vessels and vascular area showed modest changes during fractionated irradiation. Histological parameters before treatment and after three and five fractions did not significantly correlate with TCD{sub 50} after irradiation with 30 fractions within 6 weeks (p > 0.05). Hypoxic volume and perfused vessels after 10 fractions showed a significant association with local tumor control after fractionated irradiation (p = 0.018 and p = 0.019, respectively). None of these parameters remained statistically significant when the p value was adjusted for multiple comparisons. Conclusions: The results from this exploratory study suggest that determination of microenvironmental parameters during treatment provides better prognostic information for the outcome after fractionated radiotherapy than pretreatment parameters, which warrants further investigation and confirmation in experimental and clinical studies.

  10. Keratinocytes and fibroblasts in a human skin equivalent model enhance melanocyte survival and melanin synthesis after ultraviolet irradiation.

    PubMed

    Archambault, M; Yaar, M; Gilchrest, B A

    1995-05-01

    To investigate paracrine effects of fibroblasts and keratinocytes on melanocyte behavior after ultraviolet (UV) irradiation, we compared an in vitro skin equivalent model with melanocyte cultures. Human melanocytes were maintained alone in monolayer cultures or on dermal equivalents with or without keratinocytes and were irradiated daily with solar-simulated light. After seven daily UV irradiations, monolayer melanocytes displayed dose-dependent increases in cellular damage. In contrast, melanocytes on dermal equivalents survived strikingly better. Moreover, UV-irradiated skin equivalent melanocytes became highly dendritic as compared with sham-irradiated cells, closely mimicking their morphology in UV-irradiated skin. In addition, in skin equivalents melanocytes migrated from the center to the periphery of the keratinocyte layer after UV irradiation. Melanin production per culture, as measured by 14C-dihydroxyphenylalanine incorporation, was consistently higher in skin equivalent melanocytes than in monolayer melanocytes from the same donor, and it was highest in melanocytes from skin equivalents containing both keratinocytes and fibroblasts. Our data strongly suggest that fibroblasts and keratinocytes modulate melanocyte function in skin. The skin equivalent is a valuable model for investigating paracrine effects on melanocytes after UV irradiation.

  11. Irradiation combined with SU5416: Microvascular changes and growth delay in a human xenograft glioblastoma tumor line

    SciTech Connect

    Schuuring, Janneke; Bussink, Johan . E-mail: J.Bussink@rther.umcn.nl; Bernsen, Hans; Peeters, Wenny; Kogel, Albert J. van der

    2005-02-01

    Purpose: The combination of irradiation and the antiangiogenic compound SU5416 was tested and compared with irradiation alone in a human glioblastoma tumor line xenografted in nude mice. The aim of this study was to monitor microenvironmental changes and growth delay. Methods and materials: A human glioblastoma xenograft tumor line was implanted in nude mice. Irradiations consisted of 10 Gy or 20 Gy with and without SU5416. Several microenvironmental parameters (tumor cell hypoxia, tumor blood perfusion, vascular volume, and microvascular density) were analyzed after imunohistochemical staining. Tumor growth delay was monitored for up to 200 days after treatment. Results: SU5416, when combined with irradiation, has an additive effect over treatment with irradiation alone. Analysis of the tumor microenvironment showed a decreased vascular density during treatment with SU5416. In tumors regrowing after reaching only a partial remission, vascular characteristics normalized shortly after cessation of SU5416. However, in tumors regrowing after reaching a complete remission, permanent microenvironmental changes and an increase of tumor necrosis with a subsequent slower tumor regrowth was found. Conclusions: Permanent vascular changes were seen after combined treatment resulting in complete remission. Antiangiogenic treatment with SU5416 when combined with irradiation has an additive effect over treatment with irradiation or antiangiogenic treatment alone.

  12. Irradiated human endothelial progenitor cells induce bystander killing in human non-small cell lung and pancreatic cancer cells.

    PubMed

    Turchan, William T; Shapiro, Ronald H; Sevigny, Garrett V; Chin-Sinex, Helen; Pruden, Benjamin; Mendonca, Marc S

    2016-08-01

    Purpose To investigate whether irradiated human endothelial progenitor cells (hEPC) could induce bystander killing in the A549 non-small cell lung cancer (NSCLC) cells and help explain the improved radiation-induced tumor cures observed in A549 tumor xenografts co-injected with hEPC. Materials and methods We investigated whether co-injection of CBM3 hEPC with A549 NSCLC cells would alter tumor xenograft growth rate or tumor cure after a single dose of 0 or 5 Gy of X-rays. We then utilized dual chamber Transwell dishes, to test whether medium from irradiated CBM3 and CBM4 hEPC would induce bystander cell killing in A549 cells, and as an additional control, in human pancreatic cancer MIA PaCa-2 cells. The CBM3 and CBM4 hEPC were plated into the upper Transwell chamber and the A549 or MIA PaCa-2 cells were plated in the lower Transwell chamber. The top inserts with the CBM3 or CBM4 hEPC cells were subsequently removed, irradiated, and then placed back into the Transwell dish for 3 h to allow for diffusion of any potential bystander factors from the irradiated hEPC in the upper chamber through the permeable membrane to the unirradiated cancer cells in the lower chamber. After the 3 h incubation, the cancer cells were re-plated for clonogenic survival. Results We found that co-injection of CBM3 hEPC with A549 NSCLC cells significantly increased the tumor growth rate compared to A549 cells alone, but paradoxically also increased A549 tumor cure after a single dose of 5 Gy of X-rays (p < 0.05). We hypothesized that irradiated hEPC may be inducing bystander killing in the A549 NSCLC cells in tumor xenografts, thus improving tumor cure. Bystander studies clearly showed that exposure to the medium from irradiated CBM3 and CBM4 hEPC induced significant bystander killing and decreased the surviving fraction of A549 and MIA PaCa-2 cells to 0.46 (46%) ± 0.22 and 0.74 ± 0.07 (74%) respectively (p < 0.005, p < 0.0001). In addition, antibody depletion

  13. Characterization of Treefoil Peptide Genes in Iron-Ion or X-Irradiated Human Cells

    NASA Technical Reports Server (NTRS)

    Balcer-Kubiczek, E. K.; Harrison, G. H.; Xu, J. F.; Zhou, X. F.

    1999-01-01

    The gastrointestinal (GI) tract is especially sensitive to ionizing radiation, probably because of its high rate of cell turn over. Most of the data in the literature concerns the histological/anatomical description of damage rather than functional studies. In fact, previous reports in humans have shown that, at doses of 2 Gy or more, functional abnormalities appear indicating that in radiation sensitive tissues the effects of radiation are not limited to cell death. GI functions are controlled in particular by GI peptides. One hypothesis is that ionizing radiation may modulate the synthesis and release of these peptides and consequently may contribute largely to abnormalities in GI function. However, no previous studies have been concerned with GI-specific gene expression in irradiated GI tissues. The family of human trefoil peptides comprises three members thus far, all of which are expressed in specific regions of the GI tract. In addition, two trefoil peptides, pS2 (TFFI) and HITF (TFF2) are expressed in breast tissue. Their exact function in GI and breast tissues is unclear but mucosal integrity, repair, mucin secretion and responsiveness to hormones have been shown. We recently isolated and characterized pS2 as a novel p53- and estrogen receptor-independent gene whose MRNA expression in several cells lines was found to be delayed 4 to 7 days after irradiation with X-rays, fission neutrons or 1 GeV/n Fe-ions. The aim of the present study was to determine whether pS2 and HITF have a similar induction kinetics in irradiated gastric and breast cell lines, and whether they have the phorbol ester (TPA) responsive element (TRE).

  14. Motor Stereotypies and Cognitive Perseveration in Non-human Primates Exposed to Early Gestational Irradiation

    PubMed Central

    Selemon, Lynn D.; Friedman, Harriet R.

    2013-01-01

    A number of psychiatric illnesses have been associated with prenatal disturbance of brain development, including autism, attention deficit hyperactivity disorder, and schizophrenia. Individuals afflicted with these disorders exhibit both repetitive motor and cognitive behavior. The potential role that environmental insult to the developing brain may play in generating these aberrant behaviors is unclear. Here we examine the behavioral consequences of an early gestational insult in the non-human primate. Rhesus macaques were exposed to x-irradiation during the first trimester of development to disrupt neurogenesis. The behavior of five fetally irradiated monkeys (FIMs) and five control monkeys (CONs) was observed as they matured from juvenile (1.5 years) to adult ages (4–5 years). Home-cage behavior was indistinguishable in the two groups. In the testing cage, circling was prevalent in both groups at juvenile ages,persisting to adulthood in three of the five FIMs. One FIM executed a ritualized motor sequence marked by semi-circling and undulating head movements. Seven macaques (4 FIMs, 3 CONs)were tested on a spatial Delayed Alternation (DA) task as adults. Perseverative errors and non-perseverative errors were recorded in early stages of the testing, at the 0 delay interval. while performing DA, FIMs made more errors of perseveration than CONs yet the number of total errors committed did not differ between groups. The presence of motor stereotypies and cognitive perseveration in fetally irradiated non-human primates suggests that environmental insult to the embryonic brain may contribute to repetitive motor and cognitive behaviors in neuropsychiatric diseases. PMID:23769911

  15. Motor stereotypies and cognitive perseveration in non-human primates exposed to early gestational irradiation.

    PubMed

    Selemon, L D; Friedman, H R

    2013-09-17

    A number of psychiatric illnesses have been associated with prenatal disturbance of brain development, including autism, attention deficit hyperactivity disorder, and schizophrenia. Individuals afflicted with these disorders exhibit both repetitive motor and cognitive behavior. The potential role that environmental insult to the developing brain may play in generating these aberrant behaviors is unclear. Here we examine the behavioral consequences of an early gestational insult in the non-human primate. Rhesus macaques were exposed to x-irradiation during the first trimester of development to disrupt neurogenesis. The behavior of five fetally irradiated monkeys (FIMs) and five control monkeys (CONs) was observed as they matured from juvenile (1.5 years) to adult ages (4-5 years). Home-cage behavior was indistinguishable in the two groups. In the testing cage, circling was prevalent in both groups at juvenile ages, persisting to adulthood in three of the five FIMs. One FIM executed a ritualized motor sequence marked by semi-circling and undulating head movements. Seven macaques (4 FIMs, 3 CONs) were tested on a spatial Delayed Alternation (DA) task as adults. Perseverative errors and non-perseverative errors were recorded in early stages of the testing, at the 0 delay interval. While performing DA, FIMs made more errors of perseveration than CONs yet the number of total errors committed did not differ between groups. The presence of motor stereotypies and cognitive perseveration in fetally irradiated non-human primates suggests that environmental insult to the embryonic brain may contribute to repetitive motor and cognitive behaviors in neuropsychiatric diseases. Published by Elsevier Ltd.

  16. Characterization of Treefoil Peptide Genes in Iron-Ion or X-Irradiated Human Cells

    NASA Technical Reports Server (NTRS)

    Balcer-Kubiczek, E. K.; Harrison, G. H.; Xu, J. F.; Zhou, X. F.

    1999-01-01

    The gastrointestinal (GI) tract is especially sensitive to ionizing radiation, probably because of its high rate of cell turn over. Most of the data in the literature concerns the histological/anatomical description of damage rather than functional studies. In fact, previous reports in humans have shown that, at doses of 2 Gy or more, functional abnormalities appear indicating that in radiation sensitive tissues the effects of radiation are not limited to cell death. GI functions are controlled in particular by GI peptides. One hypothesis is that ionizing radiation may modulate the synthesis and release of these peptides and consequently may contribute largely to abnormalities in GI function. However, no previous studies have been concerned with GI-specific gene expression in irradiated GI tissues. The family of human trefoil peptides comprises three members thus far, all of which are expressed in specific regions of the GI tract. In addition, two trefoil peptides, pS2 (TFFI) and HITF (TFF2) are expressed in breast tissue. Their exact function in GI and breast tissues is unclear but mucosal integrity, repair, mucin secretion and responsiveness to hormones have been shown. We recently isolated and characterized pS2 as a novel p53- and estrogen receptor-independent gene whose MRNA expression in several cells lines was found to be delayed 4 to 7 days after irradiation with X-rays, fission neutrons or 1 GeV/n Fe-ions. The aim of the present study was to determine whether pS2 and HITF have a similar induction kinetics in irradiated gastric and breast cell lines, and whether they have the phorbol ester (TPA) responsive element (TRE).

  17. Particle irradiation induces FGF2 expression in normal human lens cells

    NASA Technical Reports Server (NTRS)

    Chang, P. Y.; Bjornstad K, A.; Chang, E.; McNamara, M.; Barcellos-Hoff, M. H.; Lin, S. P.; Aragon, G.; Polansky, J. R.; Lui, G. M.; Blakely, E. A.

    2000-01-01

    Particle Irradiation Induces FGF2 Expression in Normal Human Lens Cells. Particle radiations, including both proton and helium-ion beams, have been used to successfully treat choroidal melanoma, but with the complication of radiation-induced cataract. We have investigated a role for radiation-induced changes in the expression of basic fibroblast growth factor (FGF2) gene expression as part of the mechanism(s) underlying lens cell injury associated with cataract. Normal human lens epithelial (HLE) cells were cultured in vitro on extracellular matrix (ECM) originated from bovine corneal endothelial cells. This study reports evidence for rapid but transient induction of FGF2 transcripts, an increase of between 5- and 8-fold, within 0.5 h after exposure to particle radiation, followed by another wave of increased transcription at 2-3 h postirradiation. Immunofluorescence results confirm the enhanced levels of FGF2 protein rapidly after exposure to protons or helium ions, followed by another wave of increased activity unique to helium at 6 h postirradiation. This second wave of increased immunoreactivity was not observed in the proton-irradiated samples. Total FGF2 protein analysis after helium-ion exposures shows induced expression of three FGF2 isoforms, with an increase of up to 2-fold in the 18-kDa low-molecular-weight species. Studies of the effects of protons on individual FGF2 protein isoforms are in progress. Several mechanisms involving a role for FGF2 in radiation-induced cataract are discussed.

  18. Particle irradiation induces FGF2 expression in normal human lens cells

    NASA Technical Reports Server (NTRS)

    Chang, P. Y.; Bjornstad K, A.; Chang, E.; McNamara, M.; Barcellos-Hoff, M. H.; Lin, S. P.; Aragon, G.; Polansky, J. R.; Lui, G. M.; Blakely, E. A.

    2000-01-01

    Particle Irradiation Induces FGF2 Expression in Normal Human Lens Cells. Particle radiations, including both proton and helium-ion beams, have been used to successfully treat choroidal melanoma, but with the complication of radiation-induced cataract. We have investigated a role for radiation-induced changes in the expression of basic fibroblast growth factor (FGF2) gene expression as part of the mechanism(s) underlying lens cell injury associated with cataract. Normal human lens epithelial (HLE) cells were cultured in vitro on extracellular matrix (ECM) originated from bovine corneal endothelial cells. This study reports evidence for rapid but transient induction of FGF2 transcripts, an increase of between 5- and 8-fold, within 0.5 h after exposure to particle radiation, followed by another wave of increased transcription at 2-3 h postirradiation. Immunofluorescence results confirm the enhanced levels of FGF2 protein rapidly after exposure to protons or helium ions, followed by another wave of increased activity unique to helium at 6 h postirradiation. This second wave of increased immunoreactivity was not observed in the proton-irradiated samples. Total FGF2 protein analysis after helium-ion exposures shows induced expression of three FGF2 isoforms, with an increase of up to 2-fold in the 18-kDa low-molecular-weight species. Studies of the effects of protons on individual FGF2 protein isoforms are in progress. Several mechanisms involving a role for FGF2 in radiation-induced cataract are discussed.

  19. A simple assay for the study of human hair follicle damage induced by ionizing irradiation.

    PubMed

    Poeggeler, Burkhard; Bodó, Enikö; Nadrowitz, Roger; Dunst, Juergen; Paus, Ralf

    2010-08-01

    Due to its rapidly proliferating matrix keratinocytes, the hair follicle is highly sensitive to ionizing irradiation (IR)-induced skin damage and thus an instructive and clinically relevant model organ for investigating the effects of IR on rapidly dividing epithelial-mesenchymal interaction systems. Here, we have assessed the impact of IR on organ-cultured human scalp hair follicles. We show that IR significantly inhibits the proliferation and induces apoptosis of hair follicle matrix keratinocytes, disrupts normal hair follicle pigmentation, and upregulates a number of quantitative toxicity and viability markers (oxidative stress indicators, DNA oxidative damage, LDH release). This introduces human hair follicle organ culture as an excellent novel research tool for radiobiology and invites exploitation as a preclinical assay system for testing candidate radioprotectants.

  20. Electron paramagnetic resonance dose response studies for neutron irradiated human teeth

    NASA Astrophysics Data System (ADS)

    Khan, Rao F. H.; Aslam; Rink, W. J.; Boreham, D. R.

    2004-10-01

    The dosimetric response of neutron irradiated human tooth enamel has been investigated using electron paramagnetic resonance (EPR) dosimetry. Continuous energy fast neutrons of mean energy less than 450 keV were produced from the McMaster University 3 MV K.N. Van de Graaff accelerator employing a thick lithium target via 7Li(p,n) 7Be interaction. Prior to its use for various experiments, the gamma dose contamination of the neutron beams was determined at the selected proton beam energies using the tissue-equivalent proportional counter (TEPC). The neutron sensitivity (/Gy-100 mg) of human tooth enamel remained constant for various mean neutron energies ranging from 167 to 450 keV. Similarly, the EPR signal intensity remained independent of the neutron dose rate variation from 0.5 to 2.4 Gy/h.

  1. Effect of cell-derived growth factors and cytokines on the clonal outgrowth of EBV-infected B cells and established lymphoblastoid cell lines.

    PubMed

    Ifversen, P; Zhang, X M; Ohlin, M; Zeuthen, J; Borrebaeck, C A

    1993-07-01

    Epstein-Barr virus (EBV) is a potent inducer of polyclonal B lymphocyte proliferation and is widely used as a tool for the establishment of B cell lines producing human monoclonal antibodies. However, because of low transformability, low clonability, and the inherent instability of EBV-infected B cells, valuable antibody-producing B cells are often lost during this procedure. We have here examined various cell-derived cytokines for their ability to enhance both the cellular outgrowth of newly infected B cells and the clonability of infected B cells and lymphoblastoid cell lines. Our results show that the murine thymoma cell line EL-4 is superior to peripheral blood mononuclear cells in both cellular outgrowth and cloning experiments, whereas monocyte-derived factors and monocyte cell lines were less capable than peripheral blood mononuclear cells in enhancing cellular outgrowth and cloning. Furthermore, the human T cell hybridoma cell line MP6 that secretes a B cell growth and differentiation factor, recently identified as an isoform of thioredoxin, is also capable of stimulating EBV-infected B cells and lymphoblastoid cell lines. Co-cultivation of EBV-infected B cells with MP6 cells significantly enhanced the cloning efficiency at the 1 cell/well level. The present results also suggest that one potential role of the MP6-derived thioredoxin could be the up regulation of IL-6 receptor expression in EBV-infected B cells.

  2. IL-1 receptor antagonist attenuates MAP kinase/AP-1 activation and MMP1 expression in UVA-irradiated human fibroblasts induced by culture medium from UVB-irradiated human skin keratinocytes.

    PubMed

    Wang, Xiaoyong; Bi, Zhigang; Chu, Wenming; Wan, Yinsheng

    2005-12-01

    Solar UV light comprises UVB wavelengths (290-320 nm) and UVA wavelengths (320-400 nm). UVB radiation reaches the epidermis and, to a lesser extent, the upper part of the dermis, while UVA radiation penetrates more deeply into human skin. Existing studies have demonstrated that UV-irradiated epidermal keratinocytes release cytokines that indirectly promote MMP-1 production in dermal fibroblasts. In this study, we first investigated the effect of IL-1 on MAPK activity, c-Jun and c-Fos mRNA expression, and MMP-1 and MMP-2 production in UVA-irradiated human dermal fibroblasts. The results showed that UVA irradiation dose-dependently increased MMP-1 but not MMP-2 production in human skin fibroblasts. IL-1alpha and IL-1beta promoted MMP-1 but not MMP-2 production in UVA-irradiated fibroblasts. Both IL-1alpha and IL-1beta activated MAP kinase, significantly elevating c-Jun and c-Fos mRNA expression. We then investigated the indirect effect of UVB-irradiated keratinocyte culture medium on MMP-1 production in UVA-irradiated primary cultured human dermal fibroblasts and the effect of IL-1Ra. The results showed that cell culture medium from UVB-irradiated keratinocytes increased MMP-1 production in UVA-irradiated fibroblasts, and IL-1Ra dose-dependently inhibited MMP-1 production. IL-1Ra dose-dependently inhibited c-Jun mRNA expression of fibroblasts with no significant effect on c-Fos mRNA expression. These results demonstrate that UVB-irradiated keratinocytes promoted MMP-1 production in UVA-irradiated fibroblasts in a paracrine manner while IL-1Ra reduced MMP-1 production through inhibiting c-Jun mRNA expression. Collectively, our data suggest that IL-1 plays an important role in the dermal collagen degradation associated with UV-induced premature aging of the skin and IL-1Ra may be applied for the prevention and treatment of photoaging.

  3. Cell growth kinetics of the human cell line Colo-205 irradiated with photons and astatine-211 alpha-particles.

    PubMed

    Palm, S; Andersson, H; Bäck, T; Claesson, I; Delle, U; Hultborn, R; Jacobsson, L; Köpf, I; Lindegren, S

    2000-01-01

    Cell growth kinetics following Astatine-211 (211At, alpha-particle emitter) and photon irradiation were studied for the human colorectal cell line Colo-205. A growth assay using 96-well plates was chosen. The growth kinetics could be simulated by assuming certain fractions of cells with various proliferative capacities, i.e. from none up to 5 cell doublings, in addition to the defined survivors with remaining unlimited clonogenic capacity. No significant difference in cell growth characteristics was seen between 211At and photon irradiation. The cell doubling time, as calculated from the increment in optical density, was compared with the results from BrdU experiments in the early phases of growth (Tpot = 18.5 +/- 0.6 h for LDR (low dose rate) photon irradiated and 20.3 +/- 0.8 hours for sham-irradiated cells 40-45 hours post-irradiation) confirming the transient accelerated growth of irradiated cells. No statistically significant difference in growth was found between LDR, MDR (medium dose rate) and HDR (high dose rate) photon irradiation.

  4. Differential responses to x-irradiation of subpopulations of two heterogeneous human carcinomas in vitro.

    PubMed

    Leith, J T; Dexter, D L; DeWyngaert, J K; Zeman, E M; Chu, M Y; Calabresi, P; Glicksman, A S

    1982-07-01

    The responses of two heterogeneous human cancer cell lines and their derivative clones to graded single doses of X-rays were examined in vitro. One system consisted of the human colon carcinoma line DLD-1 and two subpopulations (clones A and D). The second system consisted of the human lung carcinoma line (LX1) and four subpopulations (LX1-1, LX1-2, LX1-3, and LX1-9). These subpopulations have previously been shown to be markedly heterogeneous in terms of such characteristics as karyotype, morphology, drug sensitivity, tumorigenicity, and expression of membrane glycoproteins (such as carcinoembryonic antigen and tumor colonic mucoprotein antigen). Exponentially growing cultures were irradiated with graded single doses of 100-kVp X-rays. Survival was assessed using colony formation as the end point, and responses from multiple experiments were fitted to the single-hit, multitarget equation of cell survival. Values for the mean lethal dose (D0, grays), quasithreshold dose (Dq, grays), and extrapolation number (n) were obtained. For the human colon adenocarcinoma system, these values for the three tumor lines were: DLD-1, 0.95, 2.34, and 11.7; clone A, 1.06, 2.23 and 8.20; and clone D, 1.08, 1.89, and 5.80. For the human lung carcinoma system, these values for the five sublines were: LX1, 1.14, 0.19, and 1.20; LX1-1, 0.96, 2.06, and 8.54; LX1-2, 0.98, 0.88, and 2.48; LX1-3, 0.68, 2.05, and 20.3; and LX1-9, 1.12, 0.00, and 1.00. These two human tumor systems therefore exhibit variability in their intrinsic sensitivity to X-irradiation. The data indicate that failure of some human carcinomas to respond to physical treatment modalities can be due to preexisting resistant subpopulations.

  5. Low-Dose Irradiation Enhances Gene Targeting in Human Pluripotent Stem Cells

    PubMed Central

    Hatada, Seigo; Subramanian, Aparna; Mandefro, Berhan; Ren, Songyang; Kim, Ho Won; Tang, Jie; Funari, Vincent; Baloh, Robert H.; Sareen, Dhruv

    2015-01-01

    Human pluripotent stem cells (hPSCs) are now being used for both disease modeling and cell therapy; however, efficient homologous recombination (HR) is often crucial to develop isogenic control or reporter lines. We showed that limited low-dose irradiation (LDI) using either γ-ray or x-ray exposure (0.4 Gy) significantly enhanced HR frequency, possibly through induction of DNA repair/recombination machinery including ataxia-telangiectasia mutated, histone H2A.X and RAD51 proteins. LDI could also increase HR efficiency by more than 30-fold when combined with the targeting tools zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats. Whole-exome sequencing confirmed that the LDI administered to hPSCs did not induce gross genomic alterations or affect cellular viability. Irradiated and targeted lines were karyotypically normal and made all differentiated lineages that continued to express green fluorescent protein targeted at the AAVS1 locus. This simple method allows higher throughput of new, targeted hPSC lines that are crucial to expand the use of disease modeling and to develop novel avenues of cell therapy. Significance The simple and relevant technique described in this report uses a low level of radiation to increase desired gene modifications in human pluripotent stem cells by an order of magnitude. This higher efficiency permits greater throughput with reduced time and cost. The low level of radiation also greatly increased the recombination frequency when combined with developed engineered nucleases. Critically, the radiation did not lead to increases in DNA mutations or to reductions in overall cellular viability. This novel technique enables not only the rapid production of disease models using human stem cells but also the possibility of treating genetically based diseases by correcting patient-derived cells. PMID:26185257

  6. Host cell reactivation of uv- and X-ray-damaged herpes simplex virus by Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines

    SciTech Connect

    Henderson, E.E.; Long, W.K.

    1981-12-01

    The efficacy of using an infected centers assay, employing herpes simplex virus-infected, Epstein-Barr virus-transformed lymphoblastoid cell lines (LCLs) as components, to study host cell reactivation has been explored. Herpes simplex virus type 1 (HSV-1) was shown through the infected centers assay to have detectable but varying ability to lytically infect LCLs established from chromosomal breakage syndromes or closely related genetic disorders. The rate of HSV inactivation by ultraviolet (uv) irradiation was faster in LCLs established from Cockaynes's syndrome than in normal LCLs, and faster still in LCLs established from xeroderma pigmentosum. These results indicate that Cockayne's syndrome, while having what appears to be quantitatively normal levels of uv-induced DNA repair replication, shows decreased ability to host cell reactivated uv-damaged HSV. In direct contrast, X-irradiated HSV showed identical survival when assayed on normal LCLs or LCLs established from ataxia telangiectasia showing increased sensitivity to X irradiation as measured by colony formation. Through the infected centers assay, it has also been possible to demonstrate low levels of multiplicity reactivation of mutagen-damaged HSV in permanently proliferating LCLs.

  7. Abnormal segregation of alleles in CEPH pedigree DNAs arising from allele loss in lymphoblastoid DNA

    SciTech Connect

    Royle, N.J.; Armour, J.A.L.; Crosier, M.; Jeffreys, A.J. )

    1993-01-01

    Somatic events that result in the reduction to hemior homozygosity at all loci affected by the event have been identified in lymphoblastoid DNA from mothers of two CEPH families. Using suitably informative probes, the allele deficiencies were detected by the abnormal transmission of alleles from grandparents to grandchildren, with the apparent absence of the alleles from the parent. Undetected somatic deficiencies in family DNAs could result in misscoring of recombination events and consequently introduce errors into linkage analysis. 15 refs., 2 figs.

  8. Cytogenetic damage, oncogenic transformation and p53 induction in human epithelial cells in response to irradiation

    NASA Astrophysics Data System (ADS)

    Armitage, Mark

    Ionizing radiation can have several different effects on cells, some are almost instantaneous such as the generation of DNA damage, other cellular responses take a matter of minutes or hours - DNA repair protein induction/activation, and others may take months or even years to be manifested - carcinogenesis. Human epithelial cell lines derived from both normal, non-neoplastic tissues and from a malignant source were cultured in order to examine several effects of ionizing radiation on such cell types. Cells not from a malignant source were previously immortalized by viral infection or by transfection with viral sequences. Simian virus 40 immortalised uroepithelial cells (SV-HUC) were found to be approximately a factor of two fold more radioresistant than cells of malignant origin (T24) in terms of unrepaired clastogenic damage i.e. assessment of micronuclei levels following irradiation. SV-HUC lines unlike T24 cells are non-tumourigenic when inoculated into nude athymic mice. SV-HUC lines proved very resistant to full oncogenic transformation using radiation and chemical carcinogens. However, morphological alterations and decreased anchorage dependant growth was observed in post carcinogen treated cells after appropriate cell culture conditions were utilized. The progression from this phenotype to a fully tumourigenic one was not recorded in this study. The ability of ionizing radiation to induce increased levels of the nuclear phosphoprotein p53 was also assessed using several different cell lines. SV- HUC and T24 cell lines failed to exhibit any increased p53 stabilization following irradiation. One cell line, a human papilloma virus transformed line (HPV) did show an approximate two fold increase of the wild type p53 protein after treatment with radiation. Only the cell line HPV showed any cell cycle delay, resulting in accumulation of cells in the G2/M compartment in post irradiation cell cycle analysis. The status of p53 was also assessed i.e. wild type or

  9. Cellular and molecular effects for mutation induction in normal human cells irradiated with accelerated neon ions.

    PubMed

    Suzuki, Masao; Tsuruoka, Chizuru; Kanai, Tatsuaki; Kato, Takeshi; Yatagai, Fumio; Watanabe, Masami

    2006-02-22

    We investigated the linear energy transfer (LET) dependence of mutation induction on the hypoxanthine-guanine phosphoribosyl transferase (HPRT) locus in normal human fibroblast-like cells irradiated with accelerated neon-ion beams. The cells were irradiated with neon-ion beams at various LETs ranging from 63 to 335 keV/microm. Neon-ion beams were accelerated by the Riken Ring Cyclotron at the Institute of Physical and Chemical Research in Japan. Mutation induction at the HPRT locus was detected to measure 6-thioguanine-resistant clones. The mutation spectrum of the deletion pattern of exons of mutants was analyzed using the multiplex polymerase chain reaction (PCR). The dose-response curves increased steeply up to 0.5 Gy and leveled off or decreased between 0.5 and 1.0 Gy, compared to the response to (137)Cs gamma-rays. The mutation frequency increased up to 105 keV/microm and then there was a downward trend with increasing LET values. The deletion pattern of exons was non-specific. About 75-100% of the mutants produced using LETs ranging from 63 to 335 keV/mum showed all or partial deletions of exons, while among gamma-ray-induced mutants 30% showed no deletions, 30% partial deletions and 40% complete deletions. These results suggested that the dose-response curves of neon-ion-induced mutations were dependent upon LET values, but the deletion pattern of DNA was not.

  10. Specific toxicity of aphidicolin to ultraviolet-irradiated excision proficient human skin fibroblasts

    SciTech Connect

    Tyrrell, R.M.

    1983-01-01

    Aphidicolin, a specific inhibitor of the eucaryotic alpha polymerase, has been employed to study the role of this enzyme in repair of potentially lethal damage (PLD) induced by far u.v. (254 nm) radiation in normal and repair defective primary human fibroblasts. There is strong concentration dependent specific toxicity to cells treated with a fluence of 6 Jm-2 of far-u.v. radiation and incubated with aphidicolin for 2 days over the concentration range 0.0025-2.5 micrograms/ml. A similar effect is seen with a xeroderma pigmentosum (XP) variant (excision proficient) strain but there is no specific toxicity to u.v. irradiated excision deficient XP cells of complementation group A. Inactivation of irradiated excision proficient fibroblasts is rapid over the first 6 h of aphidicolin (1 microgram/ml) treatment but the reaction takes 2 days or longer to complete depending on the u.v. dose. These results demonstrate that the apparent uncoupling of excision repair seen previously by other investigators prevents repair of PLD and is lethal to the cells.

  11. A Quantitative Proteomic Analysis of Urine from Gamma-Irradiated Non-Human Primates

    PubMed Central

    Byrum, Stephanie D; Burdine, Marie S; Orr, Lisa; Moreland, Linley; Mackintosh, Samuel G; Authier, Simon; Pouliot, Mylene; Hauer-Jensen, Martin; Tackett, Alan J

    2016-01-01

    The molecular effects of total body gamma-irradiation exposure are of critical importance as large populations of people could be exposed either by terrorists, nuclear blast, or medical therapy. In this study, we aimed to identify changes in the urine proteome using a non-human primate model system, Rhesus macaque, in order to characterize effects of acute radiation syndrome following whole body irradiation (Co-60) at 6.7 Gy and 7.4 Gy with a twelve day observation period. The urine proteome is potentially a valuable and non-invasive diagnostic for radiation exposure. Using high-resolution mass spectrometry, we identified 2346 proteins in the urine proteome. We show proteins involved in disease, cell adhesion, and metabolic pathway were significantly changed upon exposure to differing levels and durations of radiation exposure. Cell damage increased at a faster rate at 7.4 Gy compared with 6.7 Gy exposures. We report sets of proteins that are putative biomarkers of time- and dose-dependent radiation exposure. The proteomic study presented here is a comprehensive analysis of the urine proteome following radiation exposure. PMID:26962295

  12. Multi-frequency electron paramagnetic resonance study of irradiated human finger phalanxes

    NASA Astrophysics Data System (ADS)

    Zdravkova, M.; Vanhaelewyn, G.; Callens, F.; Gallez, B.; Debuyst, R.

    2005-10-01

    Electron paramagnetic resonance (EPR) is often used in dosimetry using biological samples such as teeth and bones. It is generally assumed that the radicals, formed after irradiation, are similar in both tissues as the mineral part of bone and tooth is carbonated hydroxyapatite. However, there is a lack of experimental evidence to support this assumption. The aim of the present study was to contribute to that field by studying powder and block samples of human finger phalanxes that were irradiated and analyzed by multi-frequency EPR. The results obtained from bones are different from the ones obtained in enamel by several respects: the ordering of the apatite crystallites is much smaller in bone, complicating the assignment of the observed CO 2- radicals to a specific location, and one type of CO 33- radical was only found in enamel. Moreover, a major difference was found in the non-CO 2- and non-CO 33- signals. The elucidation of the nature of these native signals (in bone and tooth enamel) still represents a big challenge.

  13. The effects of daily irradiation with polychromatic visible polarized light on human lymphocyte populations.

    PubMed

    Lim, Jeong H; Lee, Jongmin; Lee, In S; Kim, Youn J; Song, Eun Y; Choi, Young S; Yun, Yeo M

    2008-08-01

    The goal of this randomized, placebo controlled, double-blind study was to investigate the effects of transcutaneous irradiation with polychromatic visible polarized light (540-780 nm; 68% polarization; power density 3.0 E-10 W/cm(2)) on a subset population of human lymphocytes using flow cytometry. The biomodulation and therapeutic effects of visible light of different wavelengths are well known, but the immunological effects of polychromatic visible polarized light have not been investigated sufficiently. Before and after 28 consecutive days of irradiation, blood samples were collected from the subjects and the population count of the lymphocyte subset was measured. The absolute count of total lymphocytes, CD3(+) lymphocytes, and CD3(+)CD4(+) lymphocytes increased by 7% (p = 0.023), 9% (p = 0.058), and 13% (p = 0.021), respectively. Yet the absolute count of WBCs, CD3(+)CD8(+), CD19(+), and CD16(+)56(+) lymphocytes did not change significantly. The application of polychromatic visible polarized light with the aforementioned features increases the CD3(+)CD4(+) lymphocyte population. It is suggested that this regimen may be useful for the promotion of natural defenses in cell-mediated immunity.

  14. Replication of UV-irradiated DNA in human cell extracts: Evidence for mutagenic bypass of pyrimidine dimers

    SciTech Connect

    Thomas, D.C.; Kunkel, T.A. )

    1993-08-15

    The authors have examined the efficiency and fidelity of simian virus 40-origin-dependent replication of UV-irradiated double-stranded DNA in extracts of human cells. Using as a mutational target the [alpha]-complementation domain of the Escherichia coli lacZ gene in bacteriophage M13mp2DNA, replication of undamaged DNA in HeLa cell extracts was highly accurate, whereas replication of DNA irradiated with UV light (280-320 nm) was both less efficient and less accurate. Replication was inhibited by irradiation in a dose-dependent manner. Nonetheless, covalently closed, monomer-length circular products were generated that were resistant to digestion by Dpn I, showing that they resulted from semiconservative replication. These products were incised by T4 endonuclease V, whereas the undamaged replication products were not, suggesting that pyrimidine dimers were bypassed during replication. When replicated, UV-irradiated DNA was used to transfect an E. coli [alpha]-complementation host strain to score mutant M13mp2 plaques, the mutant plaque frequency was substantially higher than that obtained with either unirradiated, replicated DNA, or unreplicated, UV-irradiated DNA. Both the increased mutagenicity and the inhibition of replication associated with UV irradiation were reversed by treatment of the irradiated DNA with photolyase before replication. Sequence analysis of mutants resulting from replication of UV-irradiated DNA demonstrated that most mutants contained C [yields] T transition errors at dipyrimidine sites. A few mutants contained 1-nt frameshift errors or tandem double CC [yields] TT substitutions. The data are consistent with the interpretation that pyrimidine dimers are bypassed during replication by the multiprotein replication apparatus in human cell extracts and that this bypass is mutagenic primarily via misincorporation of dAMP opposite a cytosine (or uracil) in the dimer. 56 refs., 2 figs., 3 tabs.

  15. Structure of the replication fork in ultraviolet light-irradiated human cells.

    PubMed Central

    Cordeiro-Stone, M; Schumacher, R I; Meneghini, R

    1979-01-01

    The DNA extracted from xeroderma pigmentosum human fibroblasts previously irradiated with 12.5 J/m2 of UV light and pulse-labeled for 45 min with radioactive and (or) heavy precursors, was used to determine the structural characteristics of the replication fork. Density equilibrium centrifugation experiments showed that a fork moved 6 micrometer in 45 min and bypassed 3 pyrimidine dimers in both strands. The same length was covered in 15-20 min in control cells. The delay in irradiated cells was apparently due to pyrimidine dimers acting as temporary blocks to the fork movement. Evidence for this interpretation comes from kinetics of incorporation of [3H]thymidine into DNA, which show that the time necessary to attain a new stable level of DNA synthesis in irradiated cells is equivalent to that required for the replication fork to cover the interdimer distance in one strand. On the other hand, the action of S1 nuclease on DNA synthesized soon after irradiation gives rise to a bimodal distribution in neutral sucrose gradients, one peak corresponding to 43 X 10(6) daltons and the other to 3 X 10(6) daltons. These two DNA species are generated by the attack of the S1 nuclease on single-stranded regions associated with the replication fork. A possible explanation for these results is given by a model according to which there is a delayed bypass of the dimer in the leading strand and the appearance of gaps opposite pyrimidine dimers in the lagging strand, as a direct consequence of the discontinuous mode of DNA replication. In terms of the model, the DNA of 43 X 10(6) daltons corresponds to the leading strand, linked to the unreplicated branch of the forks, whereas the piece of 3 X 10(6) daltons is the intergap DNA coming from the lagging strand. Pulse and chase experiments reveal that the low molecular weight DNA grows in a pattern that suggests that more than one gap may be formed per replication fork. PMID:233582

  16. Mean Organ Doses Resulting From Non-Human Primate Whole Thorax Lung Irradiation Prescribed to Mid-Line Tissue.

    PubMed

    Prado, Charlotte; Kazi, Abdul; Bennett, Alexander; MacVittie, Thomas; Prado, Karl

    2015-11-01

    Multi-organ dose evaluations and the effects of heterogeneous tissue dose calculations have been retrospectively evaluated following irradiation to the whole thorax and lung in non-human primates (NHP). A clinical-based approach was established to evaluate actual doses received in the heart and lungs during whole thorax lung irradiation. Anatomical structure and organ densities have been introduced in the calculations to show the effects of dose distribution through heterogeneous tissue. Mean organ doses received by non-human primates undergoing whole thorax lung irradiations were calculated using a treatment planning system that is routinely used in clinical radiation oncology. The doses received by non-human primates irradiated following conventional dose calculations have been retrospectively reconstructed using computerized tomography-based, heterogeneity-corrected dose calculations. The use of dose volume descriptors for irradiation to organs at risk and tissue exposed to radiation is introduced. Mean and partial-volume doses to lung and heart are presented and contrasted. The importance of exact dose definitions is highlighted, and the relevance of precise dosimetry to establish organ-specific dose response relationships in NHP models of acute and delayed effects of acute radiation exposure is emphasized.

  17. [Assessment of oncogenic risk of the irradiation of the thyroid gland in humans].

    PubMed

    Zvonova, I A; Likhtarev, I A; Filiushkin, I V; Shandala, N K; Gul'ko, G M

    1991-01-01

    The paper deals with one of the most urgent aspects of irradiation hygiene, namely assessment of risk for irradiation-induced cancers of the thyroid. A model is described to predict high mortality rates of thyroidal cancer in the population due to the catastrophe at the Chernobyl Atomic Power Station. With the model, life-time risk rates involving sex and age at the moment of irradiation, as well as an irradiation mode.

  18. Gamma irradiation of human dendritic cells influences proliferation and cytokine profile of T cells in autologous mixed lymphocyte reaction.

    PubMed

    Cao, Meng-De; Chen, Zong-De; Xing, Ying

    2004-01-01

    Dendritic cells (DC) are the most potent antigen-presenting cells (APC); their ability to induce proliferation of T cells in a mixed lymphocyte reaction (MLR) assay is commonly used for the evaluation of their function. It is a general thought that gamma irradiation of APC does not influence their ability to activate T-cell proliferation, but the data from several studies are controversial. To further determine the mechanisms involved in DC-induced T-cell activation in MLR assay, human DC induced from peripheral blood mononuclear cells (PBMC) were gamma-irradiated and determine their effects on the proliferation and cytokine profiles of T cells in an autologous MLR. DC were induced from the PBMC of 11 multiple sclerosis (MS) patients with RMPI 640 medium containing recombinant human GM-CSF (rhGM-CSF; 800 U/ml) and recombinant human IL-4 (rhIL-4; 500 U/ml). DC harvested on day 7 were divided into two equal parts. One part was not irradiated (naive DC); the other was gamma-irradiated at a dose of 30 Gy. Cell surface molecules were analyzed by flow cytometry. T-cell proliferation was determined using a beta-scintillation counter. The levels of IL-2, IL-4, IL-6 and IL-10 in co-culture supernatants were measured by ELISA. The results indicated that gamma irradiation reduced expression of CD86, CD80 and HLA-DR molecules on DC, especially CD86 (P=0.0072). DC, irradiated or non-irradiated, effectively stimulated autologous T-cell proliferation. Compared to naive DC, irradiated DC showed a markedly lower capacity to promote T-cell proliferation (P=0.0073), and strikingly up-regulated secretion of IL-4 (P=0.0145) and IL-2 (P=0.0323) by autologous T cells. No significant differences were noted in IL-6 and IL-10 production between T cells co-cultured with naive DC and irradiated DC (P>0.05). It is concluded that gamma irradiation of DC not only influences the phenotype of DC but also alters their capacity to stimulate the proliferation and the cytokine profiles of autologous T

  19. Dose-dependent microRNA expression in human fibroblasts after LET irradiation

    NASA Astrophysics Data System (ADS)

    Maes, Olivier Charles; An, Jin; Wu, Honglu; Wang, Eugenia; Sarojini, Harshini

    Humans are exposed to various levels of radiation during spaceflight voyages. In cells, exposure to linear energy transfer (LET) radiation causes cellular damage and triggers responses controlled by unique gene-directed signaling pathways. MicroRNAs (miRNAs) are small ( 22- nucleotide) non-coding RNAs, which regulate gene expression generally by either degrading the messager RNA or inhibiting translation. Their implication in specific cellular response pathways is largely unknown. Here, we investigated the role of radiation-dependent changes in miRNA expression patterns after low (0.1 Gy) and high (2.0 Gy) doses of X-ray exposure in human fibroblasts, and correlated their predicted targets with the cells' genomics and proteomics profiles. A differential miRNA expression pattern was observed between low and high doses of irradiation, with early (0.5 and 2 hrs) significant changes mostly after a high dose and, late (6 and 24 hrs) significant changes after both low and high doses of irradiation. The results suggest that miRNAs may act as ‘hub' regulators of signaling pathways initially to derepress their target genes for cellular responses such as DNA repair, followed by up-regulation to suppress apoptosis, and finally down-regulation to reestablish cellular normalcy. Functional attributions are made to key microRNAs, potentially regulating known radiation biomarkers as well as radiation-responsive mechanisms of cell cycle checkpoint, proliferation and apoptosis. In summary, radiation-responsive miRNAs may have functional roles in the regulation of cell death or survival, and may become biodosimeters for radiation dose exposure. Specific microRNAs may exert a hormetic effect after low-dose radiation, and prove useful in future applications for radiation adaptive therapy and in the prevention and treatment of radiation-induced damage. The confirmation of specific miRNAs as biodosimetry markers with therapeutic applications will be necessary in future functional

  20. Electron spin resonance detection of oxygen radicals released by UVA-irradiated human fibroblasts

    NASA Astrophysics Data System (ADS)

    Souchard, J. P.; Pierlot, G.; Barbacanne, M. A.; Charveron, M.; Bonafé, J.-L.; Nepveu, F.

    1999-01-01

    This work reports the electron spin resonance (ESR) detection of oxygenated radicals (OR) released by cultured human fibroblasts after UVA (365 nm) exposure. 5,5-dimethyl-pyrroline-N-oxide (DMPO) was used as spin trap. After a UVA irradiation of one hour, followed by a latent period of at least 45 min., and an incubation time of 30 min. in a trapping medium containing DMPO, glucose, Na^+, K+ and Ca2+ an ESR signal was recorded. By contrast, an ESR signal was produced after only 15 min. incubation when calcium ionophore A23187 was used. Although the ESR signal was characteristic of the hydroxyl adduct DMPO-OH, the use of catalase and superoxide dismutase (SOD) revealed that UVA stimulated fibroblasts released the superoxide anion O2- in the medium. SOD, vitamin C and (+)-catechin inhibited the release of superoxide generated by human fibroblasts stimulated with A23187 calcium ionophore at 5 units/ml, 10-5 M and 2× 10-4 M, respectively. Dans ce travail nous présentons la détection par résonance de spin électronique (RSE) de radicaux oxygénés (RO) libérés par des fibroblastes humains en culture après irradiation aux UVA (365 nm). Le 5,5-diméthyl-1-pyrroline-N-oxyde (DMPO) a été utilisé comme piégeur de spin. Après une irradiation aux UVA d'une heure, suivie d'une période de latence d'au moins 45 min. et d'une incubation de 30 min. dans un milieu de piégeage composé de DMPO, glucose, Na^+, K+ et Ca2+, un signal RPE est enregistré. L'ionophore calcique A23187 entraîne l'apparition d'un signal RPE après seulement 15 min. d'incubation. Bien que le signal RPE obtenu corresponde à l'adduit DMPO-OH du radical hydroxyle, l'utilisation de catalase et de superoxyde dismutase (SOD) a révélé que les fibroblastes libéraient l'anion superoxyde dans le milieu de culture. Sur ce modèle cellulaire la SOD, la vitamine C et la (+) catéchine inhibent la production du radical superoxyde aux concentrations respectivement de 5 unités/ml, 10-5 M et 2× 10-4M.

  1. Ultraviolet irradiation induces the accumulation of chondroitin sulfate, but not other glycosaminoglycans, in human skin.

    PubMed

    Werth, Benjamin Boegel; Bashir, Muhammad; Chang, Laura; Werth, Victoria P

    2011-01-01

    Ultraviolet (UV) light alters cutaneous structure and function. Prior work has shown loss of dermal hyaluronan after UV-irradiation of human skin, yet UV exposure increases total glycosaminoglycan (GAG) content in mouse models. To more fully describe UV-induced alterations to cutaneous GAG content, we subjected human volunteers to intermediate-term (5 doses/week for 4 weeks) or single-dose UV exposure. Total dermal uronyl-containing GAGs increased substantially with each of these regimens. We found that UV exposure substantially increased dermal content of chondroitin sulfate (CS), but not hyaluronan, heparan sulfate, or dermatan sulfate. UV induced the accumulation of both the 4-sulfated (C4S) and 6-sulfated (C6S) isoforms of CS, but in distinct distributions. Next, we examined several CS proteoglycan core proteins and found a significant accumulation of dermal and endothelial serglycin, but not of decorin or versican, after UV exposure. To examine regulation in vitro, we found that UVB in combination with IL-1α, a cytokine upregulated by UV radiation, induced serglycin mRNA in cultured dermal fibroblasts, but did not induce the chondroitin sulfate synthases. Overall, our data indicate that intermediate-term and single-dose UVB exposure induces specific GAGs and proteoglycan core proteins in human skin in vivo. These molecules have important biologic functions and contribute to the cutaneous response to UV.

  2. Effects of low power violet laser irradiation on red blood cells volume and erythrocyte sedimentation rate in human blood

    NASA Astrophysics Data System (ADS)

    Al Musawi, Mustafa S.; Jafaar, M. S.; Ahmed, Naser M.; Al-Gailani, B. T.; Suhaimi, Fatanah M.

    2017-08-01

    This study is designed in vitro to examine the effects of low power violet laser irradiation on some human blood samples rheological factors such as mean red blood cell volume (MCV) and erythrocyte sedimentation rate (ESR). Blood samples were collected into EDTA contained tubes and separated into two equal aliquots to be attended as irradiated and control. Samples were irradiated for 20, 30, 40 or 50 min with a laser of power 10 mW. The measurements were done directly after irradiation by applying westergen method and using a computerized hemtoanalyzer. The RBCs volume and ESR were decreased after irradiation for 40min by 0.44% and 6.7% respectively. It is possible to suggest that laser irradiation can reduction red blood cells volume because of the increased concentrations of free intracellular Ca+². The result shows that ESR reduction exposed to low power laser is mostly by reason of the effect of laser on composition of the plasma that finally affects in ESR of whole blood.

  3. Cytoplasmic Irradiation Induces Metabolic Shift in Human Small Airway Epithelial Cells via Activation of Pim-1 Kinase.

    PubMed

    Wu, Jinhua; Zhang, Qin; Wuu, Yen-Ruh; Zou, Sirui; Hei, Tom K

    2017-04-01

    The unique cellular and molecular consequences of cytoplasmic damage caused by ionizing radiation were studied using a precision microbeam irradiator. Our results indicated that targeted cytoplasmic irradiation induced metabolic shift from an oxidative to glycolytic phenotype in human small airway epithelial cells (SAE). At 24 h postirradiation, there was an increase in the mRNA expression level of key glycolytic enzymes as well as lactate secretion in SAE cells. Using RNA-sequencing analysis to compare genes that were responsive to cytoplasmic versus nuclear irradiation, we found a glycolysis related gene, Pim-1, was significantly upregulated only in cytoplasmic irradiated SAE cells. Inhibition of Pim-1 activity using the selective pharmaceutic inhibitor Smi-4a significantly reduced the level of lactate production and glucose uptake after cytoplasmic irradiation. In addition, Pim-1 also inhibited AMPK activity, which is a well-characterized negative regulator of glycolysis. Distinct from the glycolysis induced by cytoplasmic irradiation, targeted nuclear irradiation also induced a transient and minimal increase in glycolysis that correlated with increased expression of Hif-1α. In an effort to explore the underline mechanism, we found that inhibition of mitochondria fission using the cell-permeable inhibitor mdivi-1 suppressed the induction of Pim-1, thus confirming Pim-1 upregulation as a downstream effect of mitochondrial dysfunction. Our data show and, for the first time, that cytoplasmic irradiation mediate expression level of Pim-1, which lead to glycolytic shift in SAE cells. Additionally, since glycolysis is frequently linked to cancer cell metabolism, our findings further suggest a role of cytoplasmic damage in promoting neoplastic changes.

  4. Chromosome aberrations induced in human lymphocytes by U-235 fission neutrons: I. Irradiation of human blood samples in the "dry cell" of the TRIGA Mark II nuclear reactor.

    PubMed

    Fajgelj, A; Lakoski, A; Horvat, D; Remec, I; Skrk, J; Stegnar, P

    1991-11-01

    A set-up for irradiation of biological samples in the TRIGA Mark II research reactor in Ljubljana is described. Threshold activation detectors were used for characterisation of the neutron flux, and the accompanying gamma dose was measured by TLDs. Human peripheral blood samples were irradiated "in vitro" and biological effects evaluated according to the unstable chromosomal aberrations induced. Biological effects of two types of cultivation of irradiated blood samples, the first immediately after irradiation and the second after 96 h storage, were studied. A significant difference in the incidence of chromosomal aberrations between these two types of samples was obtained, while our dose-response curve fitting coefficients alpha 1 = (7.71 +/- 0.09) x 10(-2) Gy-1 (immediate cultivation) and alpha 2 = (11.03 +/- 0.08) x 10(-2) Gy-1 (96 h delayed cultivation) are in both cases lower than could be found in the literature.

  5. UV irradiation-induced methionine oxidation in human skin keratins: Mass spectrometry-based non-invasive proteomic analysis.

    PubMed

    Lee, Seon Hwa; Matsushima, Keita; Miyamoto, Kohei; Oe, Tomoyuki

    2016-02-05

    Ultraviolet (UV) radiation is the major environmental factor that causes oxidative skin damage. Keratins are the main constituents of human skin and have been identified as oxidative target proteins. We have recently developed a mass spectrometry (MS)-based non-invasive proteomic methodology to screen oxidative modifications in human skin keratins. Using this methodology, UV effects on methionine (Met) oxidation in human skin keratins were investigated. The initial screening revealed that Met(259), Met(262), and Met(296) in K1 keratin were the most susceptible oxidation sites upon UVA (or UVB) irradiation of human tape-stripped skin. Subsequent liquid chromatography/electrospray ionization-MS and tandem MS analyses confirmed amino acid sequences and oxidation sites of tryptic peptides D(290)VDGAYMTK(298) (P1) and N(258)MQDMVEDYR(267) (P2). The relative oxidation levels of P1 and P2 increased in a time-dependent manner upon UVA irradiation. Butylated hydroxytoluene was the most effective antioxidant for artifactual oxidation of Met residues. The relative oxidation levels of P1 and P2 after UVA irradiation for 48 h corresponded to treatment with 100mM hydrogen peroxide for 15 min. In addition, Met(259) was oxidized by only UVA irradiation. The Met sites identified in conjunction with the current proteomic methodology can be used to evaluate skin damage under various conditions of oxidative stress. We demonstrated that the relative Met oxidation levels in keratins directly reflected UV-induced damages to human tape-stripped skin. Human skin proteins isolated by tape stripping were analyzed by MS-based non-invasive proteomic methodology. Met(259), Met(262), and Met(296) in K1 keratin were the most susceptible oxidation sites upon UV irradiation. Met(259) was oxidized by only UVA irradiation. Quantitative LC/ESI-SRM/MS analyses confirmed a time-dependent increase in the relative oxidation of target peptides (P1 and P2) containing these Met residues, upon UVA irradiation

  6. Observation of DNA damage of human hepatoma cells irradiated by heavy ions using comet assay

    PubMed Central

    Qiu, Li-Mei; Li, Wen-Jian; Pang, Xin-Yue; Gao, Qing-Xiang; Feng, Yan; Zhou, Li-Bin; Zhang, Gao-Hua

    2003-01-01

    AIM: Now many countries have developed cancer therapy with heavy ions, especially in GSI (Gesellschaft fürSchwerionenforschung mbH, Darmstadt, Germany), remarkable results have obtained, but due to the complexity of particle track structure, the basic theory still needs further researching. In this paper, the genotoxic effects of heavy ions irradiation on SMMC-7721 cells were measured using the single cell gel electrophoresis (comet assay). The information about the DNA damage made by other radiations such as X-ray, γ-ray, UV and fast neutron irradiation is very plentiful, while little work have been done on the heavy ions so far. Hereby we tried to detect the reaction of liver cancer cells to heavy ion using comet assay, meanwhile to establish a database for clinic therapy of cancer with the heavy ions. METHODS: The human hepatoma cells were chosen as the test cell line irradiated by 80Mev/u 20Ne10+ on HIRFL (China), the radiation-doses were 0, 0.5, 1, 2, 4 and 8 Gy, and then comet assay was used immediately to detect the DNA damages, 100-150 cells per dose-sample (30-50 cells were randomly observed at constant depth of the gel). The tail length and the quantity of the cells with the tail were put down. EXCEL was used for statistical analysis. RESULTS: We obtained clear images by comet assay and found that SMMC-7721 cells were all damaged apparently from the dose 0.5 Gy to 8 Gy (t-test: P < 0.001, vs control). The tail length and tail moment increased as the doses increased, and the number of cells with tails increased with increasing doses. When doses were higher than 2 Gy, nearly 100% cells were damaged. Furthermore, both tail length and tail moment, showed linear equation. CONCLUSION: From the clear comet assay images, our experiment proves comet assay can be used to measure DNA damages by heavy ions. Meanwhile DNA damages have a positive correlation with the dose changes of heavy ions and SMMC-7721 cells have a great radiosensitivity to 20Ne10+. Different

  7. Chromosome aberration yields and apoptosis in human lymphocytes irradiated with Fe-ions of differing LET

    NASA Astrophysics Data System (ADS)

    Lee, R.; Nasonova, E.; Ritter, S.

    In the present paper the relationship between cell cycle delays induced by Fe-ions of differing LET and the aberration yield observable in human lymphocytes at mitosis was examined. Cells of the same donor were irradiated with 990 MeV/n Fe-ions (LET = 155 keV/μm), 200 MeV/n Fe-ions (LET = 440 keV/μm) and X-rays and aberrations were measured in first cycle mitoses harvested at different times after 48 84 h in culture and in prematurely condensed G2-cells (PCCs) collected at 48 h using calyculin A. Analysis of the time-course of chromosomal damage in first cycle metaphases revealed that the aberration frequency was similar after X-ray irradiation, but increased two and seven fold after exposure to 990 and 200 MeV/n Fe-ions, respectively. Consequently, RBEs derived from late sampling times were significantly higher than those obtained at early times. The PCC-data suggest that the delayed entry of heavily damaged cells into mitosis results especially from a prolonged arrest in G2. Preliminary data obtained for 4.1 MeV/n Cr-ions (LET = 3160 keV/μm) revealed, that these delays are even more pronounced for low energy Fe-like particles. Additionally, for the different radiation qualities, BrdU-labeling indices and apoptotic indices were determined at several time-points. Only the exposure to low energy Fe-like particles affected the entry of lymphocytes into S-phase and generated a significant apoptotic response indicating that under this particular exposure condition a large proportion of heavily damaged cells is rapidly eliminated from the cell population. The significance of this observation for the estimation of the health risk associated with space radiation remains to be elucidated.

  8. Relationship between the chemical and morphological characteristics of human dentin after Er:YAG laser irradiation.

    PubMed

    Soares, Luís Eduardo Silva; Martin, Ovídio César Lavesa; Moriyama, Lilian Tan; Kurachi, Cristina; Martin, Airton Abrahão

    2013-06-01

    The effects of laser etching on dentin are studied by microenergy-dispersive x-ray fluorescence spectrometry (μ-EDXRF) and scanning electron microscopy (SEM) to establish the correlation of data obtained. Fifteen human third molars are prepared, baseline μ-EDXRF mappings are performed, and ten specimens are selected. Each specimen received four treatments: acid etching (control-CG) or erbium:yttrium-aluminum-garnet (Er:YAG) laser irradiation (I-100 mJ, II-160 mJ, and III-220 mJ), and maps are done again. The Ca and P content are significantly reduced after acid etching (p<0.0001) and increased after laser irradiation with 220 mJ (Ca: p<0.0153 and P: p=0.0005). The Ca/P ratio increased and decreased after CG (p=0.0052) and GI (p=0.0003) treatments, respectively. CG treatment resulted in lower inorganic content (GI: p<0.05, GII: p<0.01, and GIII: p<0.01) and higher Ca/P ratios than laser etching (GI: p<0.001, GII: p<0.01, and GIII: p<0.01). The SEM photomicrographies revealed open (CG) and partially open dentin tubules (GI, GII, and GIII). μ-EDXRF mappings illustrated that acid etching created homogeneous distribution of inorganic content over dentin. Er:YAG laser etching (220 mJ) produced irregular elemental distribution and changed the stoichiometric proportions of hydroxyapatite, as showed by an increase of mineral content. Decreases and increases of mineral content in the μ-EDXRF images are correlated to holes and mounds, respectively, as found in SEM images.

  9. Enzyme plus light therapy to repair DNA damage in ultraviolet-B-irradiated human skin

    PubMed Central

    Stege, Helger; Roza, Len; Vink, Arie A.; Grewe, Markus; Ruzicka, Thomas; Grether-Beck, Susanne; Krutmann, Jean

    2000-01-01

    Ultraviolet-B (UVB) (290–320 nm) radiation-induced cyclobutane pyrimidine dimers within the DNA of epidermal cells are detrimental to human health by causing mutations and immunosuppressive effects that presumably contribute to photocarcinogenesis. Conventional photoprotection by sunscreens is exclusively prophylactic in nature and of no value once DNA damage has occurred. In this paper, we have therefore assessed whether it is possible to repair UVB radiation-induced DNA damage through topical application of the DNA-repair enzyme photolyase, derived from Anacystis nidulans, that specifically converts cyclobutane dimers into their original DNA structure after exposure to photoreactivating light. When a dose of UVB radiation sufficient to induce erythema was administered to the skin of healthy subjects, significant numbers of dimers were formed within epidermal cells. Topical application of photolyase-containing liposomes to UVB-irradiated skin and subsequent exposure to photoreactivating light decreased the number of UVB radiation-induced dimers by 40–45%. No reduction was observed if the liposomes were not filled with photolyase or if photoreactivating exposure preceded the application of filled liposomes. The UVB dose administered resulted in suppression of intercellular adhesion molecule-1 (ICAM-1), a molecule required for immunity and inflammatory events in the epidermis. In addition, in subjects hypersensitive to nickel sulfate, elicitation of the hypersensitivity reaction in irradiated skin areas was prevented. Photolyase-induced dimer repair completely prevented these UVB radiation-induced immunosuppressive effects as well as erythema and sunburn-cell formation. These studies demonstrate that topical application of photolyase is effective in dimer reversal and thereby leads to immunoprotection. PMID:10660687

  10. Bog blueberry anthocyanins alleviate photoaging in ultraviolet-B irradiation-induced human dermal fibroblasts.

    PubMed

    Bae, Ji-Young; Lim, Soon Sung; Kim, Sun Ju; Choi, Jung-Suk; Park, Jinseu; Ju, Sung Mi; Han, Seoung Jun; Kang, Il-Jun; Kang, Young-Hee

    2009-06-01

    Fruits of bog blueberry (Vaccinium uliginosum L.) are rich in anthocyanins that contribute pigmentation. Anthocyanins have received much attention as agents with potentials preventing chronic diseases. This study investigated the capacity of anthocyanin-rich extract from bog blueberry (ATH-BBe) to inhibit photoaging in UV-B-irradiated human dermal fibroblasts. BBe anthocyanins were detected as cyanidin-3-glucoside, petunidin-3-glucoside, malvidin-3-glucoside, and delphinidin3-glucoside. ATH-BBe attenuated UV-B-induced toxicity accompanying reactive oxygen species (ROS) production and the resultant DNA damage responsible for activation of p53 and Bad. Preincubation of ATH-BBe markedly suppressed collagen degradation via blunting production of collagenolytic matrix metalloproteinases (MMP). Additionally, ATH-BBe enhanced UV-B-downregulated procollagen expression at transcriptional levels. We next attempted to explore whether ATH-BBe mitigated the MMP-promoted collagen degradation through blocking nuclear factor kappaB (NF-kappaB) activation and MAPK-signaling cascades. UV-B radiation enhanced nuclear translocation of NF-kappaB, which was reversed by treatment with ATH-BBe. The UV-B irradiation rapidly activated apoptosis signal-regulating kinase-1 (ASK-1)-signaling cascades of JNK and p38 mitogen-activated protein kinase (p38 MAPK), whereas ATH-BBe hampered phosphorylation of c-Jun, p53, and signal transducers and activators of transcription-1 (STAT-1) linked to these MAPK signaling pathways. ATH-BBe diminished UV-B augmented-release of inflammatory interleukin (IL)-6 and IL-8. These results demonstrate that ATH-BBe dampens UV-B-triggered collagen destruction and inflammatory responses through modulating NF-kappaB-responsive and MAPK-dependent pathways. Therefore, anthocyanins from edible bog blueberry may be protective against UV-induced skin photoaging.

  11. [Growth factor production and autocrine mechanism of cell proliferation regulation in the RPMI-6410t lymphoblastoid line].

    PubMed

    Seregina, T M; Mekshenkov, M I

    1988-03-01

    The human lymphoblastoid B-cell line RPMI-6410t was found to synthesize and secrete into the growth medium a factor necessary to maintain the reproduction of these cells. In the condition of low plating density (concentration 1-1000 cells per ml) cell proliferation can be maintained only in the presence of a definite dose of medium conditioned by 6410t cell growth under high concentration. Using such a medium guaranteed almost 100% cloning efficiency of these cells by the method of limiting dilutions. The cloning of 6410t cells in the presence of feeder cells, such as mouse splenocytes and peritoneal cells, failed. The 6410t cells were shown to bind specifically the growth factor secreted by them, thus suggesting the presence of a growth factor acceptor on their surface. With the help of special selective method some clones were derived which did not secrete growth factor but were likely to have growth factor acceptors on their surface. A comparison of growth properties of clones GF- and GF+ supported the idea of autocrine control of proliferation as one of the mechanisms of malignant cell transformation.

  12. Fluoroquinolones Lower Constitutive H2AX and ATM Phosphorylation in TK6 Lymphoblastoid Cells via Modulation of Intracellular Redox Status

    PubMed Central

    Halicka, H. Dorota; Smart, Daniel J.; Traganos, Frank; Williams, Gary M.; Darzynkiewicz, Zbigniew

    2008-01-01

    Accumulation of reactive oxygen species (ROS)-induced damage and mutations in genomic DNA is considered the primary etiology of aging and age-related pathologies including cancer. Strategies aimed at slowing these conditions often involve protecting against oxidative DNA damage via modulation of the intracellular redox state. Recently, a biomarker of DNA double-strand breaks (DSBs), serine-139-phosphorylated histone H2AX (γH2AX), and its upstream mediator, activated PI-3-related kinase ATM (ATMP1981), were shown to be constitutively expressed in cells and modulated by antioxidant treatment. Thus, both constitutive histone H2AX phosphorylation (CHP) and constitutive ATM activation (CAA) are thought to reflect a cell’s response to endogenous ROS-induced DSBs. In the present study, we investigated the effects of a battery of fluoroquinolone (FQ) compounds, namely Ciprofloxacin, Enrofloxacin, Gatifloxacin, Lomefloxacin and Ofloxacin, on CHP and CAA in human TK6 lymphoblastoid cells. All FQs tested reduced CHP and CAA compared to controls following 6 and 24 h treatment, with CAA being more sensitive to their effects at both time points. In addition, intracellular ROS levels and mitochondrial activities were also lowered in FQ-treated cells at 6 and 24 h. We believe that FQs mediate this effect via a combination of ROS-scavenging and mitochondrial suppression, and therefore may protect against the onset or slow the progression of numerous oxidative pathophysiological conditions. PMID:19815954

  13. Arginine to lysine 108 substitution in recombinant CYP1A2 abolishes methoxyresorufin metabolism in lymphoblastoid cells

    PubMed Central

    Hadjokas, Nicholas E; Dai, Renke; Friedman, Fred K; Spence, Michael J; Cusack, Barry J; Vestal, Robert E; Ma, Yongsheng

    2002-01-01

    Cytochrome P4501A2 (CYP1A2) activates a large number of procarcinogens to carcinogens. Phytochemicals such as flavones can inhibit CYP1A2 activity competitively, and hydroxylated derivatives of flavone (galangin) may be potent, selective inhibitors of CYP1A2 activity relative to CYP1A1 activity. Molecular modelling of the CYP1A2 interaction with hydroxylated derivatives of flavone suggests that a number of hydrophobic residues of the substrate-binding domain engage in hydrogen bonding with such inhibitors.We have tested this model using site-directed mutagenesis of these residues in expression plasmids transfected into the human B-lymphoblastoid cell line, AHH-1 TK+/−.Consistent with the molecular model's predicted placement in the active site, amino acid substitutions at the predicted residues abolished CYP1A2 enzymatic activity.Transfected cell lines contained equal amounts of immunoreactive CYP1A2.Our results support the molecular model's prediction of the critical amino acid residues present in the hydrophobic active site, residues that can hydrogen bond with CYP1A2 inhibitors and modify substrate binding and/or turnover. PMID:12023936

  14. Host Genetic Variants and Gene Expression Patterns Associated with Epstein-Barr Virus Copy Number in Lymphoblastoid Cell Lines

    PubMed Central

    Houldcroft, Charlotte J.; Petrova, Velislava; Liu, Jimmy Z.; Frampton, Dan; Anderson, Carl A.; Gall, Astrid; Kellam, Paul

    2014-01-01

    Lymphoblastoid cell lines (LCLs) are commonly used in molecular genetics, supplying DNA for the HapMap and 1000 Genomes Projects, used to test chemotherapeutic agents, and informing the basis of a number of population genetics studies of gene expression. The process of transforming human B cells into LCLs requires the presence of Epstein-Barr virus (EBV), a double-stranded DNA virus which through B-cell immortalisation maintains an episomal virus genome in every cell of an LCL at variable copy numbers. Previous studies have reported that EBV alters host-gene expression and EBV copy number may be under host genetic control. We performed a genome-wide association study of EBV genome copy number in LCLs and found the phenotype to be highly heritable, although no individual SNPs achieved a significant association with EBV copy number. The expression of two host genes (CXCL16 and AGL) was positively correlated and expression of ADARB2 was negatively correlated with EBV copy number in a genotype-independent manner. This study shows an association between EBV copy number and the gene expression profile of LCLs, and suggests that EBV copy number should be considered as a covariate in future studies of host gene expression in LCLs. PMID:25290448

  15. Lymphoblastoid Cell lines: a Continuous in Vitro Source of Cells to Study Carcinogen Sensitivity and DNA Repair

    PubMed Central

    Hussain, Tabish; Mulherkar, Rita

    2012-01-01

    Obtaining a continuous source of normal cells or DNA from a single individual has always been a rate limiting step in biomedical research. Availability of Lymphoblastoid cell lines (LCLs) as a surrogate for isolated or cryopreserved peripheral blood lymphocytes has substantially accelerated the process of biological investigations. LCLs can be established by in vitro infection of resting B cells from peripheral blood with Epstein Barr Virus (EBV) resulting in a continuous source, bearing negligible genetic and phenotypic alterations. Being a spontaneous replicating source, LCLs fulfil the requirement of constant supply of starting material for variety of assays, sparing the need of re-sampling. There is a reason to believe that LCLs are in close resemblance with the parent lymphocytes based on the ample supporting observations from a variety of studies showing significant level of correlation at molecular and functional level. LCLs, which carry the complete set of germ line genetic material, have been instrumental in general as a source of biomolecules and a system to carry out various immunological and epidemiological studies. Furthermore, in recent times their utility for analysing the whole human genome has extensively been documented. This proves the usefulness of LCLs in various genetic and functional studies. There are a few contradictory reports that have questioned the employment of LCLs as parent surrogate. Regardless of some inherent limitations LCLs are increasingly being considered as an important resource for genetic and functional research. PMID:24551762

  16. Biological effects of helium-neon laser irradiation on normal and wounded human skin fibroblasts.

    PubMed

    Hawkins, D; Abrahamse, H

    2005-06-01

    This study aimed to investigate a number of structural, cellular, and molecular responses to heliumneon (632.8 nm) laser irradiation following a single dose of 0.5, 2.5, 5, or 10 J/cm2 on normal and wounded human skin fibroblasts. Low-level laser therapy (LLLT) is a form of phototherapy, involving the application of low-power monochromatic and coherent light to injuries and lesions to stimulate healing. 1 This therapy has been successfully used for pain attenuation and to induce wound healing in nonhealing defects. Changes in normal and wounded fibroblast cell morphology were evaluated by light microscopy. Cellular parameters evaluated cell proliferation, cell viability, and cytotoxicity while molecular parameters assessed the extent of DNA damage. The results clearly demonstrate that LLLT has an effect on normal and wounded(3) human skin fibroblasts. The parameters showed that doses of 0.5, 2.5, 5, and 10 J/cm2 were sufficient to produce measurable changes in fibroblast cells. A dose of 10 J/cm2 appeared to produce a significant amount of cellular and molecular damage, which could be an important consideration for other therapies, such as photodynamic therapy.

  17. Long-Term Quantitative Biodistribution and Side Effects of Human Mesenchymal Stem Cells (hMSCs) Engraftment in NOD/SCID Mice following Irradiation.

    PubMed

    François, Sabine; Usunier, Benoit; Douay, Luc; Benderitter, Marc; Chapel, Alain

    2014-01-01

    There is little information on the fate of infused mesenchymal stem cells (MSCs) and long-term side effects after irradiation exposure. We addressed these questions using human MSCs (hMSCs) intravenously infused to nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice submitted to total body irradiation (TBI) or local irradiation (abdominal or leg irradiation). The animals were sacrificed 3 to 120 days after irradiation and the quantitative and spatial distribution of hMSCs were studied by polymerase chain reaction (PCR). Following their infusion into nonirradiated animals, hMSCs homed to various tissues. Engraftment depended on the dose of irradiation and the area exposed. Total body irradiation induced an increased hMSC engraftment level compared to nonirradiated mice, while local irradiations increased hMSC engraftment locally in the area of irradiation. Long-term engraftment of systemically administered hMSCs in NOD/SCID mice increased significantly in response to tissue injuries produced by local or total body irradiation until 2 weeks then slowly decreased depending on organs and the configuration of irradiation. In all cases, no tissue abnormality or abnormal hMSCs proliferation was observed at 120 days after irradiation. This work supports the safe and efficient use of MSCs by injection as an alternative approach in the short- and long-term treatment of severe complications after radiotherapy for patients refractory to conventional treatments.

  18. [Boron neutron capture therapy of human gastric cancer by boron-containing immunoliposomes under thermal neutron irradiation].

    PubMed

    Xu, L

    1991-10-01

    Boron neutron capture therapy (BNCT) is based on the nuclear reaction yielding high LET Li-7 and alpha particles when boron-10 is irradiated with thermal neutrons. (Et4N)2(10)B10H10 was entrapped in 40 nm liposomes coating the monoclonal antibody, MGb 2, against human gastric cancer. There were 1.4 x 10(4) 10B atoms encapsulated and 20 molecules of MGb 2 incorporated per liposomes ELISA indicated that the immunoreactivity of antibodies on liposomes retained 80%. Preferred binding to human gastric cancer cell line SGC-7901 was observed as many as 15.1 x 10(9) 10B atoms/tumor cell, 38-fold more than that to normal human embryonic lung cell line SL 7. The fluorescent immunoliposome-stained tumor cells showed membrane-fluorescence while SL 7 cells showed no obvious fluorescence. Irradiated with thermal neutrons (0.025 eV, 3.12 x 10(11)n/cm2, gamma-ray 0.84 Gy), 10B-containing immunoliposomes pretreated SGC-7901 cells survived 27%, significantly lower than non-irradiated cells or non-pretreated cells with irradiation (P less than 0.001). The results demonstrated that boron-containing immunoliposomes could bind selectively and deliver sufficient amount of boron-10 to the target tumor cells.

  19. Carboxylated nanodiamonds inhibit γ-irradiation damage of human red blood cells

    NASA Astrophysics Data System (ADS)

    Santacruz-Gomez, K.; Silva-Campa, E.; Melendrez-Amavizca, R.; Teran Arce, F.; Mata-Haro, V.; Landon, P. B.; Zhang, C.; Pedroza-Montero, M.; Lal, R.

    2016-03-01

    Nanodiamonds when carboxylated (cNDs) act as reducing agents and hence could limit oxidative damage in biological systems. Gamma (γ)-irradiation of whole blood or its components is required in immunocompetent patients to prevent transfusion-associated graft versus host disease (TA-GVHD). However, γ-irradiation of blood also deoxygenates red blood cells (RBCs) and induces oxidative damage, including abnormalities in cellular membranes and hemolysis. Using atomic force microscopy (AFM) and Raman spectroscopy, we examined the effect of cNDs on γ-irradiation mediated deoxygenation and morphological damage of RBCs. γ-Radiation induced several morphological phenotypes, including stomatocytes, codocytes and echinocytes. While stomatocytes and codocytes are reversibly damaged RBCs, echinocytes are irreversibly damaged. AFM images show significantly fewer echinocytes among cND-treated γ-irradiated RBCs. The Raman spectra of γ-irradiated RBCs had more oxygenated hemoglobin patterns when cND-treated, resembling those of normal, non-irradiated RBCs, compared to the non-cND-treated RBCs. cND inhibited hemoglobin deoxygenation and morphological damage, possibly by neutralizing the free radicals generated during γ-irradiation. Thus cNDs have the therapeutic potential to preserve the quality of stored blood following γ-irradiation.Nanodiamonds when carboxylated (cNDs) act as reducing agents and hence could limit oxidative damage in biological systems. Gamma (γ)-irradiation of whole blood or its components is required in immunocompetent patients to prevent transfusion-associated graft versus host disease (TA-GVHD). However, γ-irradiation of blood also deoxygenates red blood cells (RBCs) and induces oxidative damage, including abnormalities in cellular membranes and hemolysis. Using atomic force microscopy (AFM) and Raman spectroscopy, we examined the effect of cNDs on γ-irradiation mediated deoxygenation and morphological damage of RBCs. γ-Radiation induced several

  20. Ultraviolet light-emitting diode irradiation-induced cell death in HL-60 human leukemia cells in vitro

    PubMed Central

    XIE, DONG; SUN, YAN; WANG, LINGZHEN; LI, XIAOLING; ZANG, CHUANNONG; ZHI, YUNLAI; SUN, LIRONG

    2016-01-01

    Ultraviolet (UV) radiation is considered to be a potent cell-damaging agent in various cell lineages; however, the effect of UV light-emitting diode (LED) irradiation on human cells remains unclear. The aim of the present study was to examine the effect of UV LED irradiation emitting at 280 nm on cultured HL-60 human leukemia cells, and to explore the underlying mechanisms. HL-60 cells were irradiated with UV LED (8, 15, 30 and 60 J/m2) and incubated for 2 h after irradiation. The rates of cell proliferation and apoptosis, the cell cycle profiles and the mRNA expression of B-cell lymphoma 2 (Bcl-2) were detected using cell counting kit-8, multicaspase assays, propidium iodide staining and reverse transcription-quantitative polymerase chain reaction, respectively. The results showed that UV LED irradiation (8–60 J/m2) inhibited the proliferation of HL-60 cells in a dose-dependent manner. UV LED at 8–30 J/m2 induced dose-dependent apoptosis and G0/G1 cell cycle arrest, and inhibited the expression of Bcl-2 mRNA, while UV LED at 60 J/m2 induced necrosis. In conclusion, 280 nm UV LED irradiation inhibits proliferation and induces apoptosis and necrosis in cultured HL-60 cells. In addition, the cell cycle arrest at the G0/G1 phase and the downregulation of Bcl-2 mRNA expression were shown to be involved in UV LED-induced apoptosis. PMID:26820261

  1. Effects of high-dose gamma irradiation on tensile properties of human cortical bone: Comparison of different radioprotective treatment methods.

    PubMed

    Allaveisi, Farzaneh; Mirzaei, Majid

    2016-08-01

    There are growing interests in the radioprotective methods that can reduce the damaging effects of ionizing radiation on sterilized bone allografts. The aim of this study was to investigate the effects of 50kGy (single dose, and fractionated) gamma irradiation, in presence and absence of l-Cysteine (LC) free radical scavenger, on tensile properties of human femoral cortical bone. A total of 48 standard tensile test specimens was prepared from diaphysis of femurs of three male cadavers (age: 52, 52, and 54 years). The specimens were assigned to six groups (n=8) according to different irradiation schemes, i.e.; Control (Non-irradiated), LC-treated control, a single dose of 50kGy (sole irradiation), a single dose of 50kGy in presence of LC, 10 fractions of 5kGy (sole irradiation), and 10 fractions of 5kGy in presence of LC. Uniaxial tensile tests were carried out to evaluate the variations in tensile properties of the specimens. Fractographic analysis was performed to examine the microstructural features of the fracture surfaces. The results of multivariate analysis showed that fractionation of the radiation dose, as well as the LC treatment of the 50kGy irradiated specimens, significantly reduced the radiation-induced impairment of the tensile properties of the specimens (P<0.05). The fractographic observations were consistent with the mechanical test results. In summary, this study showed that the detrimental effects of gamma sterilization on tensile properties of human cortical bone can be substantially reduced by free radical scavenger treatment, dose fractionation, and the combined treatment of these two methods. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Alterations of human acellular tissue matrix by gamma irradiation: histology, biomechanical property, stability, in vitro cell repopulation, and remodeling.

    PubMed

    Gouk, Sok-Siam; Lim, Tit-Meng; Teoh, Swee-Hin; Sun, Wendell Q

    2008-01-01

    AlloDerm, a processed acellular human tissue matrix, is used in a number of surgical applications for tissue repair and regeneration. In the present work, AlloDerm serves as a model system for studying gamma radiation-induced changes in tissue structure and stability as well as the effect of such changes on the cell-matrix interactions, including cell repopulation and matrix remodeling. AlloDerm tissue matrix was treated with 2-30 kGy gamma irradiation at room temperature. Gamma irradiation reduced the swelling of tissue matrix upon rehydration and caused significant structural modifications, including collagen condensation and hole formation in collagen fibres. The tensile strength of AlloDerm increased at low gamma dose but decreased with increasing gamma dosage. The elasticity of irradiated AlloDerm was reduced significantly. Calorimetric study showed that gamma irradiation destabilized the tissue matrix, resulting in greater susceptibility to proteolytic enzyme degradation. Although gamma irradiation did not affect in vitro proliferation of fibroblast cells, it promoted tissue degradation upon cell repopulation and influenced synthesis and deposition of new collagen.

  3. Cytopathic Effects of X-ray Irradiation and MnO Nanoparticles on Human Glioblastoma (U87)

    NASA Astrophysics Data System (ADS)

    Kuper, K. E.; Zavjalov, E. L.; Razumov, I. A.; Romaschenko, A. V.; Stupak, A. S.; Troicky, S. Yu; Goldenberg, B. G.; Legkodymov, A. G.; Lemzyakov, A. A.; Moshkin, M. P.

    Glioblastoma is a leader among the most malignant brain tumors with the average lifespan of patients around 9-12 months. For prevention and treatment of neuropathology, a variety of therapeutic and surgical approaches are being developed and improved, including radiation and chemical therapy methods. In our work, we investigated cytopathic effect of X-ray irradiation with application of metal oxides nanoparticles such as manganese oxide (MnO) on U87 human glioblastoma cells. We used the X-ray irradiation dose of 0.5, 4, 40 and 100 Gy in combination with nanoparticles at the concentration of 0.5 ng/ml. The irradiation of glioma cell was carried out at the synchrotron radiation source VEPP-4. After cells treatments with nanoparticles for about 24 h and radiation the results were assessed by MTT assay test with 106/ml cells densities. We demonstrate that preincubation of the glioblastoma cell lines U87 with MnO nanoparticles allows reducing dose of irradiation. This combination of nanoparticles and X-ray irradiation provides new possibilities for the treatment of brain tumors.

  4. Antioxidant enzymes and the mechanism of the bystander effect induced by ultraviolet C irradiation of A375 human melanoma cells.

    PubMed

    Ghosh, Rita; Guha, Dipanjan; Bhowmik, Sudipta; Karmakar, Sayantani

    2013-09-18

    Irradiated cells generate dynamic responses in non-irradiated cells; this signaling phenomenon is known as the bystander effect (BE). Factors secreted by the irradiated cells communicate some of these signals. Conditioned medium from UVC-irradiated A375 human melanoma cells was used to study the BE. Exposure of cells to conditioned medium induce cell-cycle arrest at the G2/M transition. Although conditioned medium treatment, by itself, did not alter cell viability, treated cells were more resistant to the lethal action of UVC or H2O2. This protective effect of conditioned medium was lost within 8h. Apoptotic or autophagic cell death was not involved in this resistance. Exposure to conditioned medium did not influence the rate of DNA repair, as measured by NAD(+) depletion. The activities of catalase and superoxide dismutase were elevated in cells exposed to conditioned medium, but returned to normal levels by 8h post-treatment. These results indicate a close correlation between BE-stimulated antioxidant activity and cellular sensitivity. Cell-cycle arrest and stimulation of antioxidant activity may account for the resistance to killing that was observed in bystander cells exposed to UVC or H2O2 treatment and are consistent with the role of the BE as a natural defense function triggered by UVC irradiation. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. [Optical properties of human normal bladder tissue at five different wavelengths of laser and their linearly polarized laser irradiation in vitro].

    PubMed

    Wei, Hua-jiang; Xing, Da; Wu, Guo-yong; Jin, Ying; Gu, Huai-min

    2004-09-01

    A double-integrating-spheres system, the basic principle of measuring technology of radiation, and an optical model of biological tissues were used for the study. Optical properties of human normal bladder tissue at 476.5, 488, 496.5, 514.5 and 532 nm of laser and their linearly polarized laser irradiation were studied. The results of measurement showed that total attenuation coefficient and scattering coefficient of human normal bladder tissue at these wavelengths of laser and their linearly polarized laser irradiation increased with decreasing wavelengths. And these was an obvious distinction between the results at these wavelengths of laser and their linearly polarized laser irradiation. Absorption coefficient of human normal bladder tissue at these wavelengths of laser and their linearly polarized laser irradiation was tardily increased with decreasing wavelengths. But there were a number of gurgitations. And these were independent of the wavelengths of laser or their linearly polarized laser irradiation. Mean cosine of scattering of human normal bladder tissue at these wavelengths of laser and their linearly polarized laser irradiation also increased with decreasing wavelengths. And these was an obvious distinction with these wavelengths of laser and their linearly polarized laser irradiation. But penetration depth of human normal bladder tissue at these wavelengths of laser and their linearly polarized laser irradiation also increased with increasing wavelengths. But there were a number of gurgitations. Refractive index of human normal bladder tissue at these wavelengths of laser ranged from 1.37 to 1.44. Absorption coefficient, scattering coefficient, total attenuation coefficient, and effective attenuation coefficients of human normal bladder tissue in Kubelka-Munk two-flux model at the same wavelength of laser and the linearly polarized laser irradiation do not exhibit prominent distinction (P > 0.05). Some absorption coefficient, scattering coefficient

  6. Total Body Irradiation in the "Hematopoietic" Dose Range Induces Substantial Intestinal Injury in Non-Human Primates.

    PubMed

    Wang, Junru; Shao, Lijian; Hendrickson, Howard P; Liu, Liya; Chang, Jianhui; Luo, Yi; Seng, John; Pouliot, Mylene; Authier, Simon; Zhou, Daohong; Allaben, William; Hauer-Jensen, Martin

    2015-11-01

    The non-human primate has been a useful model for studies of human acute radiation syndrome (ARS). However, to date structural changes in various parts of the intestine after total body irradiation (TBI) have not been systematically studied in this model. Here we report on our current study of TBI-induced intestinal structural injury in the non-human primate after doses typically associated with hematopoietic ARS. Twenty-four non-human primates were divided into three groups: sham-irradiated control group; and total body cobalt-60 (60Co) 6.7 Gy gamma-irradiated group; and total body 60Co 7.4 Gy gamma-irradiated group. After animals were euthanized at day 4, 7 and 12 postirradiation, sections of small intestine (duodenum, proximal jejunum, distal jejunum and ileum) were collected and fixed in 10% formalin. The intestinal mucosal surface length, villus height and crypt depths were assessed by computer-assisted image analysis. Plasma citrulline levels were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Total bone marrow cells were counted and hematopoietic stem/progenitor cells in bone marrow were analyzed by flow cytometer. Histopathologically, all segments exhibited conspicuous disappearance of plicae circulares and prominent atrophy of crypts and villi. Intestinal mucosal surface length was significantly decreased in all intestinal segments on day 4, 7 and 12 after irradiation (P < 0.02-P < 0.001). Villus height was significantly reduced in all segments on day 4 and 7 (P = 0.02-0.005), whereas it had recovered by day 12 (P > 0.05). Crypt depth was also significantly reduced in all segments on day 4, 7 and 12 after irradiation (P < 0.04-P < 0.001). Plasma citrulline levels were dramatically reduced after irradiation, consistent with intestinal mucosal injury. Both 6.7 and 7.4 Gy TBI reduced total number of bone marrow cells. And further analysis showed that the number and function of CD45(+)CD34(+) hematopoietic stem/progenitors in bone

  7. Adenylate pool and energy charge in human lymphocytes and granulocytes irradiated at 632 nm (HeNe laser)

    NASA Astrophysics Data System (ADS)

    Bolognani, Lorenzo; Venturelli, T.; Volpi, N.; Zirilli, O.

    1995-05-01

    Aim of this report was to investigate the adenylate pool and the energy charge in human white blood cells exposed to increasing time (15, 30 and 60 min) of HeNe laser treatment. EDTA treated human blood diluted 1:1 with 0.88% KCl was added (1:5) with NaCl-dextran solution to allow sedimentation of red blood cells. 6 ml of the white cells floating in the supernatant were layered on 3 ml of Lymphoprep in plastic tubes and each tube was centrifuged (from 50 to 5000 X g for 5 min). Granulocytes were concentrated in the lower phase, whilst lymphocytes were in the intermediated phase. After further purification cytological homogeneity was tested by a cell counter. Granulocytes and lymphocytes were irradiated at +22°C with HeNe (Space, Valfivre equipment). On these population ATP was tested by luminometric procedure, the adenylate pool was separated by HPLC (Jasco) on neutralyzed perchloric extracts. ATP concentration increased in lymphocytes (+63.9%, p < 0.01) and in granulocytes (+25.0%, p < 0.05) after 60 min irradiation. The adenylate pool (tested by HPLC) does not change significatively in lymphocytes or granulocytes after 30 min irradiation, whilst in 60 min irradiated lymphocytes and granulocytes a significative increment was observed in nucleotide concentration. No changes were observed in energy charge according to Atkinson.

  8. Effect of ultrasound irradiation on α-SMA and TGF-β1 expression in human dermal fibroblasts.

    PubMed

    Maeshige, Noriaki; Terashi, Hiroto; Aoyama, Michiko; Torii, Kazuhiro; Sugimoto, Masaharu; Usami, Makoto

    2011-05-11

    Ultrasound therapy is used to promote pressure ulcer healing as an adjunctive therapy. However, the efficacy and the scientific basis of this treatment are unclear. We investigated the effect of ultrasound irradiation on alpha-smooth muscle actin (α-SMA) and transforming growth factor-beta1 (TGF-β1) expression in human dermal fibroblasts. These are important factors for acceleration of wound closure. We used pulsed ultrasound of 0, 0.1, 0.5, and 1.0 W/cm2. TGF-β1 and α-SMA mRNA was measured by quantitative real-time polymerase chain reaction, α-SMA protein was examined by western blot, and localization of α-SMA was evaluated by immunofluorescence staining. Expression of α-SMA and TGF-β1 mRNA was increased at 24 h but not at 48 h after ultrasound irradiation. There were significant differences between controls of 0 W/cm² and 0.1 W/cm² with a 1.34 ± 0.26 fold increase in α-SMA (P < 0.05) and a 1.78 ± 0.57 fold increase in TGF-β1 (P < 0.05). Protein levels of α-SMA were also increased and detected in ultrasound irradiated fibroblasts at 24 h. Ultrasound irradiation promotes α-SMA expression in human dermal fibroblasts and this suggests the biological mechanism of ultrasound efficacy on chronic wound treatment.

  9. Gastrodia elata Blume Extract Modulates Antioxidant Activity and Ultraviolet A-Irradiated Skin Aging in Human Dermal Fibroblast Cells.

    PubMed

    Song, Eunju; Chung, Haeyon; Shim, Eugene; Jeong, Jung-Ky; Han, Bok-Kyung; Choi, Hyuk-Joon; Hwang, Jinah

    2016-11-01

    Gastrodia elata Blume (GEB), a traditional herbal medicine, has been used to treat a wide range of neurological disorders (e.g., paralysis and stroke) and skin problems (e.g., atopic dermatitis and eczema) in oriental medicine. This study was designed to investigate the antioxidant ability of GEB and its antiaging effect on human dermal fibroblast cells (HDF). The total phenolic and flavonoid contents of GEB were 21.8 and 0.43 mg/g dry weight (DW), respectively. The ergothioneine content of GEB was 0.41 mg/mL DW. The DPPH and ABTS radical scavenging activities of GEB at 5 and 10 mg/mL approximately ranged between 31% and 44%. The superoxide dismutase activity of GEB at 10 and 25 mg/mL was 57% and 76%, respectively. GEB increased procollagen type 1 (PC1) production and inhibited matrix metalloproteinase-1 (MMP-1) production and elastase-1 activity in UVA-irradiated HDF. PC1 messenger RNA (mRNA) levels decreased upon UVA irradiation, but recovered in response to high doses of GEB in HDF. On the contrary, GEB significantly decreased MMP-1 and elastase-1 mRNA levels, which were markedly induced in UVA-irradiated HDF. Collectively, these results suggest that GEB has sufficient antioxidant ability to prevent the signs of skin aging in UVA-irradiated human skin cells, suggesting its potential as a natural antiaging product.

  10. Cyclobutane-type pyrimidine photodimer formation and induction of ornithine decarboxylase in human skin fibroblasts after UV irradiation

    SciTech Connect

    Niggli, H.J.; Roethlisberger, R.

    1988-12-01

    Cyclobutane-type pyrimidine photodimers as well as the induction of ornithine decarboxylase (ODC) may serve as biochemical markers of the mutagenic and carcinogenic effects of ultraviolet light (UV). For this reason, it is important to compare the formation of pyrimidine dimers with the induction of ODC in human skin fibroblasts after irradiation with UVC (200-290 nm) and UVB (290-320 nm). In our studies we determined cytosine-thymine (C-T) as well as thymine-thymine dimer yields (T-T) by high-pressure liquid chromatography in cultures of neonatal normal human foreskin-derived fibroblasts after irradiation with UVC and UVB light. It was found that the yield of dimerization and the ratio of T-T/C-T decreased from the UVC to the UVB region. Time-course studies of ODC-induction in the same cells indicated that the maximal activity after UVB irradiation was retarded compared to UVC exposure. For the UV-induced ODC-levels, however, no significant difference in maximal induction could be measured after UVC and UVB irradiation at fluences where comparable yields of thymine dimerization are produced. Similar ODC-maxima were obtained with strains from children, while cells from adults showed significantly less pronounced ODC induction, indicating that ODC-response decreases with age and may therefore be used as a marker of aging.

  11. Genetic factors affecting EBV copy number in lymphoblastoid cell lines derived from the 1000 Genome Project samples

    PubMed Central

    Rodríguez, Juan Antonio; Farré, Xavier; Layouni, Hafid; Marigorta, Urko M.; Cundiff, Caitlin; Heredia-Genestar, Jose Maria; Navarro, Arcadi

    2017-01-01

    Epstein-Barr virus (EBV), human herpes virus 4, has been classically associated with infectious mononucleosis, multiple sclerosis and several types of cancers. Many of these diseases show marked geographical differences in prevalence, which points to underlying genetic and/or environmental factors. Those factors may include a different susceptibility to EBV infection and viral copy number among human populations. Since EBV is commonly used to transform B-cells into lymphoblastoid cell lines (LCLs) we hypothesize that differences in EBV copy number among individual LCLs may reflect differential susceptibility to EBV infection. To test this hypothesis, we retrieved whole-genome sequenced EBV-mapping reads from 1,753 LCL samples derived from 19 populations worldwide that were sequenced within the context of the 1000 Genomes Project. An in silico methodology was developed to estimate the number of EBV copy number in LCLs and validated these estimations by real-time PCR. After experimentally confirming that EBV relative copy number remains stable over cell passages, we performed a genome wide association analysis (GWAS) to try detecting genetic variants of the host that may be associated with EBV copy number. Our GWAS has yielded several genomic regions suggestively associated with the number of EBV genomes per cell in LCLs, unraveling promising candidate genes such as CAND1, a known inhibitor of EBV replication. While this GWAS does not unequivocally establish the degree to which genetic makeup of individuals determine viral levels within their derived LCLs, for which a larger sample size will be needed, it potentially highlighted human genes affecting EBV-related processes, which constitute interesting candidates to follow up in the context of EBV related pathologies. PMID:28654678

  12. Genetic factors affecting EBV copy number in lymphoblastoid cell lines derived from the 1000 Genome Project samples.

    PubMed

    Mandage, Rajendra; Telford, Marco; Rodríguez, Juan Antonio; Farré, Xavier; Layouni, Hafid; Marigorta, Urko M; Cundiff, Caitlin; Heredia-Genestar, Jose Maria; Navarro, Arcadi; Santpere, Gabriel

    2017-01-01

    Epstein-Barr virus (EBV), human herpes virus 4, has been classically associated with infectious mononucleosis, multiple sclerosis and several types of cancers. Many of these diseases show marked geographical differences in prevalence, which points to underlying genetic and/or environmental factors. Those factors may include a different susceptibility to EBV infection and viral copy number among human populations. Since EBV is commonly used to transform B-cells into lymphoblastoid cell lines (LCLs) we hypothesize that differences in EBV copy number among individual LCLs may reflect differential susceptibility to EBV infection. To test this hypothesis, we retrieved whole-genome sequenced EBV-mapping reads from 1,753 LCL samples derived from 19 populations worldwide that were sequenced within the context of the 1000 Genomes Project. An in silico methodology was developed to estimate the number of EBV copy number in LCLs and validated these estimations by real-time PCR. After experimentally confirming that EBV relative copy number remains stable over cell passages, we performed a genome wide association analysis (GWAS) to try detecting genetic variants of the host that may be associated with EBV copy number. Our GWAS has yielded several genomic regions suggestively associated with the number of EBV genomes per cell in LCLs, unraveling promising candidate genes such as CAND1, a known inhibitor of EBV replication. While this GWAS does not unequivocally establish the degree to which genetic makeup of individuals determine viral levels within their derived LCLs, for which a larger sample size will be needed, it potentially highlighted human genes affecting EBV-related processes, which constitute interesting candidates to follow up in the context of EBV related pathologies.

  13. Carboxylated nanodiamonds inhibit γ-irradiation damage of human red blood cells.

    PubMed

    Santacruz-Gomez, K; Silva-Campa, E; Melendrez-Amavizca, R; Teran Arce, F; Mata-Haro, V; Landon, P B; Zhang, C; Pedroza-Montero, M; Lal, R

    2016-04-07

    Nanodiamonds when carboxylated (cNDs) act as reducing agents and hence could limit oxidative damage in biological systems. Gamma (γ)-irradiation of whole blood or its components is required in immunocompetent patients to prevent transfusion-associated graft versus host disease (TA-GVHD). However, γ-irradiation of blood also deoxygenates red blood cells (RBCs) and induces oxidative damage, including abnormalities in cellular membranes and hemolysis. Using atomic force microscopy (AFM) and Raman spectroscopy, we examined the effect of cNDs on γ-irradiation mediated deoxygenation and morphological damage of RBCs. γ-Radiation induced several morphological phenotypes, including stomatocytes, codocytes and echinocytes. While stomatocytes and codocytes are reversibly damaged RBCs, echinocytes are irreversibly damaged. AFM images show significantly fewer echinocytes among cND-treated γ-irradiated RBCs. The Raman spectra of γ-irradiated RBCs had more oxygenated hemoglobin patterns when cND-treated, resembling those of normal, non-irradiated RBCs, compared to the non-cND-treated RBCs. cND inhibited hemoglobin deoxygenation and morphological damage, possibly by neutralizing the free radicals generated during γ-irradiation. Thus cNDs have the therapeutic potential to preserve the quality of stored blood following γ-irradiation.

  14. The Effect of Lycopene Preexposure on UV-B-Irradiated Human Keratinocytes

    PubMed Central

    Ascenso, Andreia; Pedrosa, Tiago; Pinho, Sónia; Pinho, Francisco; de Oliveira, José Miguel P. Ferreira; Cabral Marques, Helena; Oliveira, Helena; Simões, Sandra; Santos, Conceição

    2016-01-01

    Lycopene has been reported as the antioxidant most quickly depleted in skin upon UV irradiation, and thus it might play a protective role. Our goal was to investigate the effects of preexposure to lycopene on UV-B-irradiated skin cells. Cells were exposed for 24 h to 10 M lycopene, and subsequently irradiated and left to recover for another 24 h period. Thereafter, several parameters were analyzed by FCM and RT-PCR: genotoxicity/clastogenicity by assessing the cell cycle distribution; apoptosis by performing the Annexin-V assay and analyzing gene expression of apoptosis biomarkers; and oxidative stress by ROS quantification. Lycopene did not significantly affect the profile of apoptotic, necrotic and viable cells in nonirradiated cells neither showed cytostatic effects. However, irradiated cells previously treated with lycopene showed an increase in both dead and viable subpopulations compared to nonexposed irradiated cells. In irradiated cells, lycopene preexposure resulted in overexpression of BAX gene compared to nonexposed irradiated cells. This was accompanied by a cell cycle delay at S-phase transition and consequent decrease of cells in G0/G1 phase. Thus, lycopene seems to play a corrective role in irradiated cells depending on the level of photodamage. Thus, our findings may have implications for the management of skin cancer. PMID:26664697

  15. Raman study of human dentin irradiated with Er:YAG laser

    NASA Astrophysics Data System (ADS)

    S. Soares, Luis E.; Martin, Airton A.; Brugnera, Aldo, Jr.; Zanin, Fatima A.; Arisawa, Emilia A.; T. Pacheco, Marcos T.

    2004-09-01

    Raman Spectroscopy was used to examine the distribution of the mineral and organic components in the human dentin before and after the chemical and thermal etching process. Polished dentin disks (n = 6/group) with 4mm thickness from twelve third molars were irradiated with Er:YAG laser. The dentin disks were prepared by polishing through a series of SiO2 papers with water and cleaned by ultrasonic system. Four pretreatment were performed. The disks were etched with 37% phosphoric acid (group I), Er:YAG laser 80mJ, 3Hz, 30s. (group II), Er:YAG laser 120mJ, 3Hz, 30s. (group III) and Er:YAG laser 180mJ, 3Hz, 30s. (group IV). The Raman spectra obtained from normal and treated dentin were analyzed. Attention was paid to the mineral PO4 (962 cm-1), CO3 (1073 cm-1) and to the organic component (1453cm-1). Raman spectroscopy showed that the mineral and organic dentin content were more affected in autoclaved teeth than in the specimens treated by Thymol. Peak area reduction in the specimens treated by Thymol in group I and II showed to be the most conservative procedures regarding to changes in organic and inorganic dentin components. Pulse energies of 120 and 180mJ showed to produce more reduction in the organic and inorganic content associated with more reduction in the peak areas at 960 and 1453cm-1.

  16. Er:YAG laser irradiation of human dentin: Raman study of collagen

    NASA Astrophysics Data System (ADS)

    Soares, Luis E. S.; Martin, Airton A.; Brugnera, Aldo, Jr.; Zanin, Fatima; Arisawa, Emilia A.; Pacheco, Marcos T. T.

    2004-05-01

    Raman Spectroscopy was used to examine the distribution of the organic components in the human dentin before and after the chemical and thermal etching process. Polished dentin disks (n = 6/group) with 4mm thickness from twelve third molars were irradiated with Er:YAG laser. The dentin disks were prepared by polishing through a series of SiO2 papers with water and cleaned by ultrasonic system. Four pretreatment were performed. The disks were etched with 37% phoshporic acid for 15 s (group 1), Er:YAG laser 80 mJ, 3Hz, 30s. (group II), Er:YAG laser 120 mJ, 3Hz, 30s. (group III) and Er:YAG laser 180mJ, 3Hz, 30s. (group IV). The Raman spectra obtained from normal and treated dentin were analyzed. Attention was paid to the organic component (1453cm-1). Raman spectroscopy showed that the organic dentin content were more affected in autoclaved teeth than in the specimens treated by Thymol. Peak area reduction in the specimens treated by Thymol in group I and II showed to be the most conservative procedures regarding to changes in organic dentin components. Pulse energies of 120 and 180 mJ showed to preduce more reduction in the organic content associated with more reduction in the peak areas at 1453 cm-1.

  17. DNA repair within nucleosome cores of UV-irradiated human cells

    SciTech Connect

    Jensen, K.A.; Smerdon, M.J. )

    1990-05-22

    We have compared the distributions of repair synthesis and pyrimidine dimers (PD) in nucleosome core DNA during the early (fast) repair phase and the late (slow) repair phase of UV-irradiated human fibroblasts. As shown previously, repair synthesis is nonuniform in nucleosome core particles during the fast repair phase, and the distribution curve can be approximated by a model where repair synthesis occurs preferentially in the 5' and 3' end regions. In this report, we show that, during the slow repair phase, (3H)dThd-labeled repair patches are much more uniformly distributed in core DNA, although they appear to be preferentially located in sequences degraded slowly by exonuclease III. This change in distribution cannot be explained by an increase in patch size during slow repair, since the size of these patches actually decreases to about half the size measured during the fast repair phase. Furthermore, PD mapping within core DNA at the single-nucleotide level demonstrated that, at least within the 30-130-base region from the 5' end, there is little (or no) selective removal of PD during the fast repair phase. However, the nonuniform distribution of repair synthesis obtained during fast repair throughout most of the core DNA region (approximately 40-146 bases) is accounted for by the nonuniform distribution of PD in core DNA. The near-uniform distribution of repair synthesis observed during slow repair may result from more extensive nucleosome rearrangement and/or nucleosome modification during this phase.

  18. Protective effects of sodium selenite supplementation against irradiation-induced damage in non-cancerous human esophageal cells.

    PubMed

    Puspitasari, Irma M; Yamazaki, Chiho; Abdulah, Rizky; Putri, Mirasari; Kameo, Satomi; Nakano, Takashi; Koyama, Hiroshi

    2017-01-01

    The administration of radioprotective compounds is one approach to preventing radiation damage in non-cancerous tissues. Therefore, radioprotective compounds are crucial in clinical radiotherapy. Selenium is a radioprotective compound that has been used in previous clinical studies of radiotherapy. However, evidence regarding the effectiveness of selenium in radiotherapy and the mechanisms underlying the selenium-induced reduction of the side effects of radiotherapy remains insufficient. To further investigate the effectiveness of selenium in radiotherapy, the present study examined the protective effects of sodium selenite supplementation administered prior to X-ray radiation treatment in CHEK-1 non-cancerous human esophageal cells. Sodium selenite supplementation increased glutathione peroxidase 1 (GPx-1) activity in a dose- and time-dependent manner. The sodium selenite dose that induced the highest GPx-1 activity was determined to be 50 nM for 72 h prior to radiotherapy. The half-maximal inhibitory concentration of sodium selenite in CHEK-1 cells was 3.6 µM. Sodium selenite supplementation increased the survival rate of the cells in a dose-dependent manner and enhanced the degree of cell viability at 72 h post-irradiation (P<0.05). Combined treatment with 50 nM sodium selenite and 2 gray (Gy) X-ray irradiation decreased the number of sub-G1 cells from 5.9 to 4.2% (P<0.05) and increased the proportion of G1 cells from 58.8 to 62.1%, compared with 2 Gy X-ray irradiation alone; however, this difference was not statistically significant (P=1.00). Western blot analysis revealed that treatment with 2 Gy X-ray irradiation significantly increased the expression levels of cleaved poly (ADP-ribose) polymerase (PARP; P<0.05). In addition, combined treatment with 50 nM sodium selenite and 2 Gy X-ray irradiation reduced the expression levels of cleaved PARP protein, compared with 2 Gy X-ray irradiation alone; however, this reduction was not statistically significant (P=0

  19. Protective effects of sodium selenite supplementation against irradiation-induced damage in non-cancerous human esophageal cells

    PubMed Central

    Puspitasari, Irma M.; Yamazaki, Chiho; Abdulah, Rizky; Putri, Mirasari; Kameo, Satomi; Nakano, Takashi; Koyama, Hiroshi

    2017-01-01

    The administration of radioprotective compounds is one approach to preventing radiation damage in non-cancerous tissues. Therefore, radioprotective compounds are crucial in clinical radiotherapy. Selenium is a radioprotective compound that has been used in previous clinical studies of radiotherapy. However, evidence regarding the effectiveness of selenium in radiotherapy and the mechanisms underlying the selenium-induced reduction of the side effects of radiotherapy remains insufficient. To further investigate the effectiveness of selenium in radiotherapy, the present study examined the protective effects of sodium selenite supplementation administered prior to X-ray radiation treatment in CHEK-1 non-cancerous human esophageal cells. Sodium selenite supplementation increased glutathione peroxidase 1 (GPx-1) activity in a dose- and time-dependent manner. The sodium selenite dose that induced the highest GPx-1 activity was determined to be 50 nM for 72 h prior to radiotherapy. The half-maximal inhibitory concentration of sodium selenite in CHEK-1 cells was 3.6 µM. Sodium selenite supplementation increased the survival rate of the cells in a dose-dependent manner and enhanced the degree of cell viability at 72 h post-irradiation (P<0.05). Combined treatment with 50 nM sodium selenite and 2 gray (Gy) X-ray irradiation decreased the number of sub-G1 cells from 5.9 to 4.2% (P<0.05) and increased the proportion of G1 cells from 58.8 to 62.1%, compared with 2 Gy X-ray irradiation alone; however, this difference was not statistically significant (P=1.00). Western blot analysis revealed that treatment with 2 Gy X-ray irradiation significantly increased the expression levels of cleaved poly (ADP-ribose) polymerase (PARP; P<0.05). In addition, combined treatment with 50 nM sodium selenite and 2 Gy X-ray irradiation reduced the expression levels of cleaved PARP protein, compared with 2 Gy X-ray irradiation alone; however, this reduction was not statistically significant (P=0

  20. Salidroside protects against premature senescence induced by ultraviolet B irradiation in human dermal fibroblasts.

    PubMed

    Mao, G-X; Xing, W-M; Wen, X-L; Jia, B-B; Yang, Z-X; Wang, Y-Z; Jin, X-Q; Wang, G-F; Yan, J

    2015-06-01

    Salidroside, the predominant component of a Chinese herbal medicine, Rhodiola rosea L., becomes an attractive bio-agent due to its multifunction. Although it is well proposed that this herbal medicine may have photoprotective effect according to the folk hearsay, the direct supportive experimental evidences linking the drug with skin ageing have rarely been reported so far. The study was conducted to investigate the photoprotective role of salidrosdie and its related mechanisms in vitro. First, a premature senescence model induced by UVB irradiation (250 mJ cm(-2)) in human dermal fibroblasts (HDFs) was established, and senescent phenotypes were evaluated by cell morphology, cell proliferation, senescence-associated beta-galactosidase (SA-β-gal) activity and cell cycle distribution. Then the photoprotective effect of salidroside was investigated. Cells were pre-treated with various doses of salidroside (1, 5 and 10 μM) followed by the sublethal dosage of UVB exposure and then were harvested for various detections, including senescence-associated phenotypes and molecules, alteration of oxidative stress, matrix metalloproteinase-1 (MMP-1) secretion and inflammatory response. Pre-treatment of salidroside dose dependently reversed the senescent state of HDFs induced by UVB as evidenced by elevated cell viability, decreased SA-β-gal activity and relieving of G1/G0 cell cycle arrest. UVB-induced increased protein expression of cyclin-dependent kinase (CDK) inhibitors p21(WAF) (1) and p16(INK) (4) was also repressed by salidrosdie treatment in a dose-dependent manner. Meanwhile, the increment of malondialdehyde (MDA) level in UVB-irradiated HDFs was inhibited upon salidroside treatment. Additionally, salidroside significantly attenuated UVB-induced synthesis of MMP-1 as well as the production of IL-6 and TNF-α in HDFs. Our data provided the evidences for the protective role of salidroside against UVB-induced premature senescence in HDFs probably via its anti

  1. Effects of gamma irradiation and repetitive freeze-thaw cycles on the biomechanical properties of human flexor digitorum superficialis tendons.

    PubMed

    Ren, Dejie; Sun, Kang; Tian, Shaoqi; Yang, Xu; Zhang, Cailong; Wang, Wenhao; Huang, Hongjie; Zhang, Jihua; Deng, Yujie

    2012-01-10

    An increasing number of tissue banks have begun to focus on gamma irradiation and freeze-thaw in the reconstruction of anterior cruciate ligaments using allografts. The purpose of this study was to evaluate the biomechanical properties of human tendons after exposure to gamma radiation and repeated freeze-thaw cycles and to compare them with fresh specimens. Forty flexor digitorum superficialis tendons were surgically procured from five fresh cadavers and divided into four groups: fresh tendon, gamma irradiation, freeze-thaw and gamma irradiation+freeze-thaw. The dose of gamma irradiation was 25 kGy. Each freeze-thaw cycle consisted of freezing at -80 °C for 7 day and thawing at 25 °C for 6 h. These tendons underwent 4 freeze-thaw cycles. Biomechanical properties were analyzed during load-to-failure testing. The fresh tendons were found to be significantly different in ultimate load, stiffness and ultimate stress relative to the other three groups. The tendons of the gamma+freeze-thaw group showed a significant decrease in ultimate load, ultimate stress and stiffness compared with the other three groups. Gamma irradiation and repeated freezing-thawing (4 cycles) can change the biomechanical properties. However, no significant difference was found between these two processes on the effect of biomechanical properties. It is recommended that gamma irradiation (25 kGy) and repetitive freeze-thaw cycles (4 cycles) should not be adopted in the processing of the allograft tendons. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Cell killing and chromatid damage in primary human bronchial epithelial cells irradiated with accelerated 56Fe ions

    NASA Technical Reports Server (NTRS)

    Suzuki, M.; Piao, C.; Hall, E. J.; Hei, T. K.

    2001-01-01

    We examined cell killing and chromatid damage in primary human bronchial epithelial cells irradiated with high-energy 56Fe ions. Cells were irradiated with graded doses of 56Fe ions (1 GeV/nucleon) accelerated with the Alternating Gradient Synchrotron at Brookhaven National Laboratory. The survival curves for cells plated 1 h after irradiation (immediate plating) showed little or no shoulder. However, the survival curves for cells plated 24 h after irradiation (delayed plating) had a small initial shoulder. The RBE for 56Fe ions compared to 137Cs gamma rays was 1.99 for immediate plating and 2.73 for delayed plating at the D10. The repair ratio (delayed plating/immediate plating) was 1.67 for 137Cs gamma rays and 1.22 for 56Fe ions. The dose-response curves for initially measured and residual chromatid fragments detected by the Calyculin A-mediated premature chromosome condensation technique showed a linear response. The results indicated that the induction frequency for initially measured fragments was the same for 137Cs gamma rays and 56Fe ions. On the other hand, approximately 85% of the fragments induced by 137Cs gamma rays had rejoined after 24 h of postirradiation incubation; the corresponding amount for 56Fe ions was 37%. Furthermore, the frequency of chromatid exchanges induced by gamma rays measured 24 h after irradiation was higher than that induced by 56Fe ions. No difference in the amount of chromatid damage induced by the two types of radiations was detected when assayed 1 h after irradiation. The results suggest that high-energy 56Fe ions induce a higher frequency of complex, unrepairable damage at both the cellular and chromosomal levels than 137Cs gamma rays in the target cells for radiation-induced lung cancers.

  3. Cell killing and chromatid damage in primary human bronchial epithelial cells irradiated with accelerated 56Fe ions

    NASA Technical Reports Server (NTRS)

    Suzuki, M.; Piao, C.; Hall, E. J.; Hei, T. K.

    2001-01-01

    We examined cell killing and chromatid damage in primary human bronchial epithelial cells irradiated with high-energy 56Fe ions. Cells were irradiated with graded doses of 56Fe ions (1 GeV/nucleon) accelerated with the Alternating Gradient Synchrotron at Brookhaven National Laboratory. The survival curves for cells plated 1 h after irradiation (immediate plating) showed little or no shoulder. However, the survival curves for cells plated 24 h after irradiation (delayed plating) had a small initial shoulder. The RBE for 56Fe ions compared to 137Cs gamma rays was 1.99 for immediate plating and 2.73 for delayed plating at the D10. The repair ratio (delayed plating/immediate plating) was 1.67 for 137Cs gamma rays and 1.22 for 56Fe ions. The dose-response curves for initially measured and residual chromatid fragments detected by the Calyculin A-mediated premature chromosome condensation technique showed a linear response. The results indicated that the induction frequency for initially measured fragments was the same for 137Cs gamma rays and 56Fe ions. On the other hand, approximately 85% of the fragments induced by 137Cs gamma rays had rejoined after 24 h of postirradiation incubation; the corresponding amount for 56Fe ions was 37%. Furthermore, the frequency of chromatid exchanges induced by gamma rays measured 24 h after irradiation was higher than that induced by 56Fe ions. No difference in the amount of chromatid damage induced by the two types of radiations was detected when assayed 1 h after irradiation. The results suggest that high-energy 56Fe ions induce a higher frequency of complex, unrepairable damage at both the cellular and chromosomal levels than 137Cs gamma rays in the target cells for radiation-induced lung cancers.

  4. Cellular and mitochondrial glutathione redox imbalance in lymphoblastoid cells derived from children with autism

    PubMed Central

    James, S. Jill; Rose, Shannon; Melnyk, Stepan; Jernigan, Stefanie; Blossom, Sarah; Pavliv, Oleksandra; Gaylor, David W.

    2009-01-01

    Research into the metabolic phenotype of autism has been relatively unexplored despite the fact that metabolic abnormalities have been implicated in the pathophysiology of several other neurobehavioral disorders. Plasma biomarkers of oxidative stress have been reported in autistic children; however, intracellular redox status has not yet been evaluated. Lymphoblastoid cells (LCLs) derived from autistic children and unaffected controls were used to assess relative concentrations of reduced glutathione (GSH) and oxidized disulfide glutathione (GSSG) in cell extracts and isolated mitochondria as a measure of intracellular redox capacity. The results indicated that the GSH/GSSG redox ratio was decreased and percentage oxidized glutathione increased in both cytosol and mitochondria in the autism LCLs. Exposure to oxidative stress via the sulfhydryl reagent thimerosal resulted in a greater decrease in the GSH/GSSG ratio and increase in free radical generation in autism compared to control cells. Acute exposure to physiological levels of nitric oxide decreased mitochondrial membrane potential to a greater extent in the autism LCLs, although GSH/GSSG and ATP concentrations were similarly decreased in both cell lines. These results suggest that the autism LCLs exhibit a reduced glutathione reserve capacity in both cytosol and mitochondria that may compromise antioxidant defense and detoxification capacity under prooxidant conditions.—James, S. J., Rose, S., Melnyk, S., Jernigan, S., Blossom, S., Pavliv, O., Gaylor, D. W. Cellular and mitochondrial glutathione redox imbalance in lymphoblastoid cells derived from children with autism. PMID:19307255

  5. Duck lymphocytes. VIII. T-lymphoblastoid cell lines from reticuloendotheliosis virus-induced tumours.

    PubMed

    Chan, S W; Bando, Y; Warr, G W; Middleton, D L; Higgins, D A

    1999-04-01

    The T strain of reticuloendotheliosis virus (REV-T) obtained, along with the helper chicken syncytia virus (CSV), from the CSO4 cell line was highly oncogenic and rapidly fatal in ducks. Tumours were mainly seen in spleen, but neoplastic cells were observed microscopically in many organs. In vitro REV transformation of duck lymphocytes failed to yield stable cell lines, so cells from organs (blood, bone marrow, spleen, lymph node, bursa of Fabricius) of infected birds were used to establish cell lines. Some of these cell lines have been cloned. The success rates of establishment and cloning were increased if cells were cultured in a range of media containing different supplements; however, medium containing 5% foetal calf serum (FCS) and 5% duck serum was generally most efficacious for initial establishment, while spent medium from the parental line supplemented with a further 20% FCS gave best results for cloning. Cloned cell lines had the morphology of lymphoblastoid cells, with irregular nuclei and diffuse chromatin. Analysis of mRNA extracted from these cell lines showed that the uncloned lines were strongly expressing the β chain of the T cell antigen receptor (TCR) and weakly expressing immunoglobulin (Ig) polypeptides [λ light chain and μ, υ, υ (ΔFc) and α heavy chains in various proportions], suggesting the presence of T and B cells. The cloned cell lines that could be classified were TCR β+ ve T cells. This is the first report of the establishment, cloning and partial characterization of duck lymphoblastoid cell lines.

  6. Exosome-Mediated Telomere Instability in Human Breast Epithelial Cancer Cells after X Irradiation.

    PubMed

    Al-Mayah, Ammar H J; Bright, Scott J; Bowler, Debbie A; Slijepcevic, Predrag; Goodwin, Edwin; Kadhim, Munira A

    2017-01-01

    In directly irradiating cells, telomere metabolism is altered and similar effects have been observed in nontargeted cells. Exosomes and their cargo play dominant roles in communicating radiation-induced bystander effects with end points related to DNA damage. Here we report novel evidence that exosomes are also responsible for inducing telomere-related bystander effects. Breast epithelial cancer cells were exposed to either 2 Gy X rays, or exposed to irradiated cell conditioned media (ICCM), or exosomes purified from ICCM. Compared to control cells, telomerase activity decreased in the 2 Gy irradiated cells and both bystander samples after one population doubling. At the first population doubling, telomere length was shorter in the 2 Gy irradiated sample but not in the bystander samples. By 24 population doublings telomerase activity recovered to control levels in all samples; however, the 2 Gy irradiated sample continued to demonstrate short telomeres and both bystander samples acquired shorter telomeres. RNase treatment of exosomes prevented the bystander effects on telomerase and telomere length that were observed at 1 population doubling and 24 population doublings, respectively. Thermal denaturation by boiling eliminated the reduction of telomere length in bystander samples, suggesting that the protein fraction of exosomes also contributes to the telomeric effect. RNase treatment plus boiling abrogated all telomere-related effects in directly irradiated and bystander cell populations. These findings suggest that both proteins and RNAs of exosomes can induce alterations in telomeric metabolism, which can instigate genomic instability in epithelial cancer cells after X-ray irradiation.

  7. Chromosome aberrations induced in human lymphocytes after partial-body irradiation

    SciTech Connect

    Fong, L.; Lai-Lei Ting; Po-Ming Wang

    1995-10-01

    Chromosomal aberrations in peripheral blood lymphocytes obtained from two patients before and after they received one fraction of partial-body irradiation for palliative treatment were analyzed. Blood samples were taken 30 min and 24 h after radiation treatment. The yield of dicentrics obtained from case A 30 min after a partial-body (about 21%) treatment with 8 Gy was 0.066/cell, while the yield obtained 24 h radiation treatment was 0.071/cell. The fraction of irradiated lymphocytes that reached metaphase at 52 h was 0.08 as evaluated by mixing cultures of in vitro irradiated and unirradiated blood. The yield of dicentrics for blood from case B 30 min after 6 Gy partial-body (about 24%) irradiation was 0.655/cell, while the yield 24 h after irradiation was 0.605/cell. The fraction of irradiated cells was 0.29. Estimation of doses and irradiated fractions for the two cases using the method proposed by Dolphin and the Qdr method is discussed. Although there was no significant difference between the mean yields of dicentrics per cell obtained 30 min and 24 h after radiation treatment, the data obtained at 24 h seemed more useful for the purpose of dose estimation. When a higher dose (8 Gy) was delivered to a smaller percentage of the body, underestimation of the dose was encountered. 18 refs., 4 tabs.

  8. FTIR and SEM analysis of CO2 laser irradiated human enamel.

    PubMed

    Corrêa-Afonso, Alessandra Marques; Bachmann, Luciano; Almeida, Cíntia Guimarães de; Corona, Silmara Aparecida Milori; Borsatto, Maria Cristina

    2012-09-01

    Considering the enamel chemical structure, especially carbonate band, which has a major role in the caries prevention, the objective of the present study was to assess the chemical alterations on the enamel irradiated with CO(2) laser by means of FTIR spectroscopy and SEM analysis. The enamel surfaces were analysed on a spectrometer for acquisition of the absorption spectrum relative to the chemical composition of the control sample. The irradiation was conducted with a 10.6-μm CO(2) laser (0.55W, 660W/cm(2)). The carbonate absorption band at 1600-1291cm(-1) as well as the water absorption band at 3793-2652cm(-1) was measured in each sample after the irradiation. The water band was measured again 24-h after the irradiation. The band area of each chemical compound was delimited, the background was subtracted, and the area under each band was integrated. Each area was normalized by the phosphate band (1190-702cm(-1)). There was a statistically significant decrease (p<0.05) in the water content after irradiation (control: 0.184±0.04; irradiated: 0.078±0.026), which increased again after rehydration (0.145±0.038). The carbonate/phosphate ratio was measured initially (0.112±0.029) and its reduction after irradiation indicated the carbonate loss (0.088±0.014) (p<0.05). The 10.6-μm CO(2) laser irradiation diminishes the carbonate and water contents in the enamel after irradiation. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. Depletion of Securin Induces Senescence After Irradiation and Enhances Radiosensitivity in Human Cancer Cells Regardless of Functional p53 Expression

    SciTech Connect

    Chen Wenshu; Yu Yichu; Lee Yijang; Chen, J.-H.; Hsu, H.-Y.; Chiu, S.-J.

    2010-06-01

    Purpose: Radiotherapy is one of the best choices for cancer treatment. However, various tumor cells exhibit resistance to irradiation-induced apoptosis. The development of new strategies to trigger cancer cell death besides apoptosis is necessary. This study investigated the role of securin in radiation-induced apoptosis and senescence in human cancer cells. Methods and Materials: Cell survival was determined using clonogenic assays. Western blot analysis was used to analyze levels of securin, caspase-3, PARP, p53, p21, Rb, gamma-H2AX, and phospho-Chk2. Senescent cells were analyzed using a beta-galactosidase staining assay. A securin-expressed vector (pcDNA-securin) was stably transfected into securin-null HCT116 cells. Securin gene knockdown was performed by small interfering RNA and small hairpin RNA in HCT116 and MDA-MB-231 cells, respectively. Results: Radiation was found to induce apoptosis in securin wild type HCT116 cells but induced senescence in securin-null cells. Restoration of securin reduced senescence and increased cell survival in securin-null HCT116 cells after irradiation. Radiation-induced gamma-H2AX and Chk2 phosphorylation were induced transiently in securin-wild-type cells but exhibited sustained activation in securin-null cells. Securin gene knockdown switches irradiation-induced apoptosis to senescence in both HCT116 p53-null and MDA-MB-231 cells. Conclusions: Our results demonstrated that the level of securin expression plays a determining role in the radiosensitivity and fate of cells. Depletion of securin impairs DNA repair after irradiation, increasing DNA damage and promoting senescence in the residual surviving cells regardless of functional p53 expression. The knockdown of securin may contribute to a novel radiotherapy protocol for the treatment of human cancer cells that are resistant to irradiation.

  10. A branching process model for the analysis of abortive colony size distributions in carbon ion-irradiated normal human fibroblasts.

    PubMed

    Sakashita, Tetsuya; Hamada, Nobuyuki; Kawaguchi, Isao; Hara, Takamitsu; Kobayashi, Yasuhiko; Saito, Kimiaki

    2014-05-01

    A single cell can form a colony, and ionizing irradiation has long been known to reduce such a cellular clonogenic potential. Analysis of abortive colonies unable to continue to grow should provide important information on the reproductive cell death (RCD) following irradiation. Our previous analysis with a branching process model showed that the RCD in normal human fibroblasts can persist over 16 generations following irradiation with low linear energy transfer (LET) γ-rays. Here we further set out to evaluate the RCD persistency in abortive colonies arising from normal human fibroblasts exposed to high-LET carbon ions (18.3 MeV/u, 108 keV/µm). We found that the abortive colony size distribution determined by biological experiments follows a linear relationship on the log-log plot, and that the Monte Carlo simulation using the RCD probability estimated from such a linear relationship well simulates the experimentally determined surviving fraction and the relative biological effectiveness (RBE). We identified the short-term phase and long-term phase for the persistent RCD following carbon-ion irradiation, which were similar to those previously identified following γ-irradiation. Taken together, our results suggest that subsequent secondary or tertiary colony formation would be invaluable for understanding the long-lasting RCD. All together, our framework for analysis with a branching process model and a colony formation assay is applicable to determination of cellular responses to low- and high-LET radiation, and suggests that the long-lasting RCD is a pivotal determinant of the surviving fraction and the RBE.

  11. A branching process model for the analysis of abortive colony size distributions in carbon ion-irradiated normal human fibroblasts

    PubMed Central

    Sakashita, Tetsuya; Hamada, Nobuyuki; Kawaguchi, Isao; Hara, Takamitsu; Kobayashi, Yasuhiko; Saito, Kimiaki

    2014-01-01

    A single cell can form a colony, and ionizing irradiation has long been known to reduce such a cellular clonogenic potential. Analysis of abortive colonies unable to continue to grow should provide important information on the reproductive cell death (RCD) following irradiation. Our previous analysis with a branching process model showed that the RCD in normal human fibroblasts can persist over 16 generations following irradiation with low linear energy transfer (LET) γ-rays. Here we further set out to evaluate the RCD persistency in abortive colonies arising from normal human fibroblasts exposed to high-LET carbon ions (18.3 MeV/u, 108 keV/µm). We found that the abortive colony size distribution determined by biological experiments follows a linear relationship on the log–log plot, and that the Monte Carlo simulation using the RCD probability estimated from such a linear relationship well simulates the experimentally determined surviving fraction and the relative biological effectiveness (RBE). We identified the short-term phase and long-term phase for the persistent RCD following carbon-ion irradiation, which were similar to those previously identified following γ-irradiation. Taken together, our results suggest that subsequent secondary or tertiary colony formation would be invaluable for understanding the long-lasting RCD. All together, our framework for analysis with a branching process model and a colony formation assay is applicable to determination of cellular responses to low- and high-LET radiation, and suggests that the long-lasting RCD is a pivotal determinant of the surviving fraction and the RBE. PMID:24501383

  12. Morphological degradation of human hair cuticle due to simulated sunlight irradiation and washing.

    PubMed

    Richena, M; Rezende, C A

    2016-08-01

    Morphological changes in hair surface are undesirable, since they cause shine loss, roughness increase and split ends. These effects occur more frequently in the cuticle, which is the outermost layer of the hair strand, and thus the most exposed to the environmental damages. Sunlight irradiation contributes significantly to these morphological alterations, which motivates the investigation of this effect on hair degradation. In this work, the influence of irradiation and hand-washing steps on the morphology of pigmented and non-pigmented hair cuticle was investigated using field emission scanning electron microscopy (FESEM) and transmission electron microscopy (TEM). To simulate daily conditions, where hair is hand-washed and light exposed, samples of dark brown and gray hair underwent three different conditions: 1) irradiation with a mercury lamp for up to 600h; 2) irradiation with the mercury lamp combined with washes with a sodium lauryl sulphate solution; and 3) only washing. A new preparation procedure was applied for TEM samples to minimize natural variations among different hair strands: a single hair strand was cut into two neighbouring halves and only one of them underwent irradiation and washing. The non-exposed half was used as a control, so that the real effects caused by the controlled irradiation and washing procedures could be highlighted in samples that had very similar morphologies initially. More than 25images/sample were analysed using FESEM (total of 300 images) and ca. 150images/sample were obtained with TEM (total of 900 images). The results presented herein show that the endocuticle and the cell membrane complex (CMC) are the cuticle structures more degraded by irradiation. Photodegradation alone results in fracturing, cavities (Ø≈20-200nm) and cuticle cell lifting, while the washing steps were able to remove cuticle cells (≈1-2 cells removed after 60 washes). Finally, the combined action of irradiation and washing caused the most severe

  13. Effect of gamma irradiation on mechanical properties of human cortical bone: influence of different processing methods.

    PubMed

    Kaminski, Artur; Jastrzebska, Anna; Grazka, Ewelina; Marowska, Joanna; Gut, Grzegorz; Wojciechowski, Artur; Uhrynowska-Tyszkiewicz, Izabela

    2012-08-01

    The secondary sterilisation by irradiation reduces the risk of infectious disease transmission with tissue allografts. Achieving sterility of bone tissue grafts compromises its biomechanical properties. There are several factors, including dose and temperature of irradiation, as well as processing conditions, that may influence mechanical properties of a bone graft. The purpose of this study was to evaluate the effect of gamma irradiation with doses of 25 or 35 kGy, performed on dry ice or at ambient temperature, on mechanical properties of non-defatted or defatted compact bone grafts. Left and right femurs from six male cadaveric donors aged from 46 to 54 years, were transversely cut into slices of 10 mm height, parallel to the longitudinal axis of the bone. Compact bone rings were assigned to the eight experimental groups according to the different processing method (defatted or non-defatted), as well as gamma irradiation dose (25 or 35 kGy) and temperature conditions of irradiation (ambient temperature or dry ice). Axial compression testing was performed with a material testing machine. Results obtained for elastic and plastic regions of stress-strain curves examined by univariate analysis are described. Based on multivariate analysis it was found that defatting of bone rings had no significant effect on any mechanical parameter studied, whereas irradiation with both doses decreased significantly the ultimate strain and its derivative toughness. The elastic limit and resilience were significantly increased by irradiation with the dose 25 kGy, but not 35 kGy, when the time of irradiation was longer. Additionally, irradiation at ambient temperature decreased maximum load, elastic limit, resilience, and ultimate stress. As strain in the elastic region was not affected, decreased elastic limit resulted in lower resilience. The opposite phenomenon was observed in the plastic region, where in spite of the lower ultimate stress, the toughness was increased due to the

  14. Comparison of the Effects of Carbon Ion and Photon Irradiation on the Angiogenic Response in Human Lung Adenocarcinoma Cells

    SciTech Connect

    Kamlah, Florentine; Haenze, Joerg; Arenz, Andrea; Seay, Ulrike; Hasan, Diya; Gottschald, Oana R.; Seeger, Werner; Rose, Frank

    2011-08-01

    Purpose: Radiotherapy resistance is a commonly encountered problem in cancer treatment. In this regard, stabilization of endothelial cells and release of angiogenic factors by cancer cells contribute to this problem. In this study, we used human lung adenocarcinoma (A549) cells to compare the effects of carbon ion and X-ray irradiation on the cells' angiogenic response. Methods and Materials: A549 cells were irradiated with biologically equivalent doses for cell survival of either carbon ions (linear energy transfer, 170 keV/{mu}m; energy of 9.8 MeV/u on target) or X-rays and injected with basement membrane matrix into BALB/c nu/nu mice to generate a plug, allowing quantification of angiogenesis by blood vessel enumeration. The expression of angiogenic factors (VEGF, PlGF, SDF-1, and SCF) was assessed at the mRNA and secreted protein levels by using real-time reverse transcription-PCR and enzyme-linked immunosorbent assay. Signal transduction mediated by stem cell factor (SCF) was assessed by phosphorylation of its receptor c-Kit. For inhibition of SCF/c-Kit signaling, a specific SCF/c-Kit inhibitor (ISCK03) was used. Results: Irradiation of A549 cells with X-rays (6 Gy) but not carbon ions (2 Gy) resulted in a significant increase in blood vessel density (control, 20.71 {+-} 1.55; X-ray, 36.44 {+-} 3.44; carbon ion, 16.33 {+-} 1.03; number per microscopic field). Concordantly, irradiation with X-rays but not with carbon ions increased the expression of SCF and subsequently caused phosphorylation of c-Kit in endothelial cells. ISCK03 treatment of A549 cells irradiated with X-rays (6 Gy) resulted in a significant decrease in blood vessel density (X-ray, 36.44 {+-} 3.44; X-ray and ISCK03, 4.33 {+-} 0.71; number of microscopic field). These data indicate that irradiation of A549 cells with X-rays but not with carbon ions promotes angiogenesis. Conclusions: The present study provides evidence that SCF is an X-ray-induced mediator of angiogenesis in A549 cells, a

  15. THE PRODUCTION OF VESICULAR STOMATITIS VIRUS BY ANTIGEN- OR MITOGEN-STIMULATED LYMPHOCYTES AND CONTINUOUS LYMPHOBLASTOID LINES

    PubMed Central

    Nowakowski, Maja; Feldman, Joseph D.; Kano, Shogo; Bloom, Barry R.

    1973-01-01

    A variety of lymphoid cell populations were examined in terms of their ability to replicate vesicular stomatitis virus (VSV), a lytic, RNA-containing virus maturing at the cell surface. The number of cells capable of producing VSV was estimated in terms of infectious centers by the virus plaque assay (VPA), and morphologically by electron microscopy (EM). The lymphoid cells examined in this study included: (a) lymph node cells from delayed hypersensitive guinea pigs stimulated by specific antigen, (b) mouse spleen cells activated by selective bone marrow-derived (B) cell and thymus derived (T) cell mitogens, and (c) cells of human and murine continuous lymphoblastoid or lymphoma lines. In unstimulated cultures of guinea pig lymph node cells there is a background of approximately 1 in 1,000 cells which produces VSV; in purified protein derivative (PPD)-stimulated cultures the number of cells producing virus was 1.6% in the VPA and 1.9% by EM. These cells were large lymphocytes with some morphological features of transformed lymphocytes but were not typical blast cells. A few macrophages were associated with virus in both stimulated and control cultures. These observations indicate that (a) cells responsive to antigens, as detected by a marker virus, were lymphocytes; (b) cells other than lymphocytes (macrophages) were capable of replicating VSV even without antigenic stimulation; and (c) the correlation of results obtained by VPA and morphologic examination was usually quite good. Of the total number of mouse spleen cells stimulated with concanavalin (Con A), a T cell mitogen, 4.5 (EM)–5.7% (VPA) were associated with VSV. These were characteristic transformed lymphocytes, similar to phytohemagglutinin (PHA)-stimulated human lymphocytes. In contrast Escherichia coli lipopolysaccharide (LPS)-treated mouse spleen cultures contained lower numbers of virus plaque-forming cells. The majority of such cells associated with virus displayed extensive rough endoplasmic

  16. Cytokine expression and ultrastructural alterations in fresh-frozen, freeze-dried and γ-irradiated human amniotic membranes.

    PubMed

    Paolin, Adolfo; Trojan, Diletta; Leonardi, Antonio; Mellone, Stefano; Volpe, Antonio; Orlandi, Augusto; Cogliati, Elisa

    2016-09-01

    The aim of this work was to compare the effects on human amniotic membrane of freeze-drying and γ-irradiation at doses of 10, 20 and 30 kGy, with freezing. For this purpose, nine cytokines (interleukin 10, platelet-derived growth factor-AA, platelet-derived growth factor-BB, basic fibroblast growth factor, epidermal growth factor, transforming growth factor beta 1, and tissue inhibitors of metalloproteinase-1, -2, and -4) were titrated in 5 different preparations for each of 3 amniotic membranes included in the study. In addition, the extracellular matrix structure of each sample was assessed by transmission electron microscopy. While freeze-drying did not seem to affect the biological structure or cytokine content of the different amniotic membrane samples, γ-irradiation led to a significant decrease in the tissue inhibitors of metalloproteinase-4, basic fibroblast growth factor and epidermal growth factor, and induced structural damage to the epithelium, basement membrane and lamina densa. The higher the irradiation dose the more severe the damage to the amniotic membrane structure. In conclusion, the Authors recommend processing amniotic membrane under sterile conditions to guarantee safety at every step rather than final sterilization with γ-irradiation, thereby avoiding alteration to the biological characteristics of the amniotic membrane.

  17. Pyrimidine dimer induction and repair in cultured human skin keratinocytes or melanocytes after irradiation with monochromatic ultraviolet radiation

    SciTech Connect

    Schothorst, A.A.; Evers, L.M.; Noz, K.C.; Filon, R.; van Zeeland, A.A. )

    1991-06-01

    We compared the susceptibilities of cultured melanocytes and keratinocytes to dimer induction in DNA by monochromatic ultraviolet (UV) radiation. Keratinocytes as well as melanocytes were derived from human foreskin, grown as a monolayer in petri dishes, covered with phosphate-buffered saline containing 0.1% glucose, and irradiated. UV irradiation was carried out at 254, 297, and 302 nm as well as with a light source emitting predominantly 312 nm. The induction of pyrmidine dimers was assessed by determination of the number of T4 endonuclease V-sensitive sites (ESS). We found a slightly higher response for dimer induction in melanocytes at 254, 297, and 302 nm; this difference was only significant at the 297-nm wavelength. Action spectra for pyrimidine dimer induction were derived from the exposure-response data obtained. The action spectra mimic to a large degree the action spectra for dimer induction in other cultured mammalian cells. The repair rate during a post-irradiation period lasting up to 24 h was substantially the same for the two cell types. The percentage of T4 endonuclease V-sensitive sites (ESS) remaining 9 and 24 h after irradiation was 45% and 30%, respectively.

  18. Integrin {beta}1-dependent invasive migration of irradiation-tolerant human lung adenocarcinoma cells in 3D collagen matrix

    SciTech Connect

    Ishihara, Seiichiro; Haga, Hisashi; Yasuda, Motoaki; Mizutani, Takeomi; Kawabata, Kazushige; Shirato, Hiroki; Nishioka, Takeshi

    2010-06-04

    Radiotherapy is one of the effective therapies used for treating various malignant tumors. However, the emergence of tolerant cells after irradiation remains problematic due to their high metastatic ability, sometimes indicative of poor prognosis. In this study, we showed that subcloned human lung adenocarcinoma cells (A549P-3) that are irradiation-tolerant indicate high invasive activity in vitro, and exhibit an integrin {beta}1 activity-dependent migratory pattern. In collagen gel overlay assay, majority of the A549P-3 cells displayed round morphology and low migration activity, whereas a considerable number of A549P-3IR cells surviving irradiation displayed a spindle morphology and high migration rate. Blocking integrin {beta}1 activity reduced the migration rate of A549P-3IR cells and altered the cell morphology allowing them to assume a round shape. These results suggest that the A549P-3 cells surviving irradiation acquire a highly invasive integrin {beta}1-dependent phenotype, and integrin {beta}1 might be a potentially effective therapeutic target in combination with radiotherapy.

  19. Chromosome damage evolution after low and high LET irradiation

    NASA Astrophysics Data System (ADS)

    Andreev, Sergey; Eidelman, Yuri

    Ionizing radiation induces DNA and chromatin lesions which are converted to chromosome lesions detected in the first post-irradiation mitosis by classic cytogenetic techniques as chromosomal aberrations (CAs). These techniques allow to monitor also delayed aberrations observed after many cell generations post-irradiation - the manifestation of chromosomal instability phenotype (CIN). The problem discussed is how to predict time evolution from initial to delayed DNA/chromosome damage. To address this question, in the present work a mechanistic model of CIN is elaborated which integrates pathways of (*) DNA damage induction and its conversion to chromosome lesions (aberrations), (**) lesion transmission and generation through cell cycles. Delayed aberrations in subsequent cycles are formed in the model owing to two pathways, DNA damage generation de novo as well as CA transmission from previous cycles. DNA damage generation rate is assumed to consist of bystander and non-bystander components. Bystander signals impact all cells roughly equally, whereas non-bystander DSB generation rate differs for the descendants of unirradiated and irradiated cells. Monte Carlo simulation of processes underlying CIN allows to predict the time evolution of initial radiation-induced damage - kinetics curve for delayed unstable aberrations (dicentrics) together with dose response and RBE as a function of time after high vs low LET irradiation. The experimental data for radiation-induced CIN in TK6 lymphoblastoid cells and human lymphocytes irradiated with low (gamma) and high (Fe, C) LET radiation are analyzed on the basis of the proposed model. One of the conclusions is that without bystander signaling, just taking into account the initial DNA damage and non-bystander DSB generation, it is impossible to describe the available experimental data for high-LET-induced CIN. The exact contribution of bystander effects for high vs low LET remains unknown, but the relative contribution may be

  20. Loss of structural water and carbonate of Nd:YAG laser-irradiated human enamel.

    PubMed

    Corrêa-Afonso, Alessandra Marques; Bachmann, Luciano; de Almeida, Cíntia Guimarães; Dibb, Regina Guenka Palma; Borsatto, Maria Cristina

    2015-05-01

    The objective of this study was to use Fourier Transform Infrared (FTIR) spectroscopy and Scanning Electron Microscopy (SEM) to assess whether Nd:YAG laser irradiation associated with a dye or not alters the chemical constitution of the enamel. Fourteen enamel sections were randomly divided into two groups: (1) Nd:YAG and (2) dye + Nd:YAG. First, the untreated enamel surfaces were analyzed by FTIR to acquire the control absorption spectrum. Next, Group 2 received a layer of inactivated coal diluted in deionized water before laser treatment. Enamel samples belonging to groups 1 and 2 were then irradiated with a 1,064-nm Nd:YAG laser (80 mJ, 10 Hz) in the contact mode; the carbonate absorption band and the water absorption band were measured in each sample after irradiation. The water band was measured again 24 h, 48 h, and 7 days after irradiation. Group 1 had statistically similar water and carbonate contents before and after irradiation. Group 2 displayed significantly lower (p < 0.05) water content after irradiation, which remained constant along time at 24 and 48 h. After 7 days, the water content increased slightly, being statistically higher than in the other experimental periods, except for the control. The carbonate/phosphate ratio was measured only at the beginning, and after irradiation, it decreased only in Group 2 indicating carbonate loss (p < 0.05). Irradiation with 1,064-nm Nd:YAG laser associated with a dye reduces the carbonate and structural water content in the enamel.

  1. Pretreatment of Ferulic Acid Protects Human Dermal Fibroblasts against Ultraviolet A Irradiation

    PubMed Central

    Hahn, Hyung Jin; Kim, Ki Bbeum; Bae, Seunghee; Choi, Byung Gon; An, Sungkwan

    2016-01-01

    Background Approximately 90%~99% of ultraviolet A (UVA) ray reaches the Earth's surface. The deeply penetrating UVA rays induce the formation of reactive oxygen species (ROS), which results in oxidative stress such as photoproducts, senescence, and cell death. Thus, UVA is considered a primary factor that promotes skin aging. Objective Researchers investigated whether pretreatment with ferulic acid protects human dermal fibroblasts (HDFs) against UVA-induced cell damages. Methods HDF proliferation was analyzed using the water-soluble tetrazolium salt assay. Cell cycle distribution and intracellular ROS levels were assessed by flow cytometric analysis. Senescence was evaluated using a senescence-associated β-galactosidase assay, while Gadd45α promoter activity was analyzed through a luciferase assay. The expression levels of superoxide dismutase 1 (SOD1), catalase (CAT), xeroderma pigmentosum complementation group A and C, matrix metalloproteinase 1 and 3, as well as p21 and p16 were measured using quantitative real-time polymerase chain reaction. Results Inhibition of proliferation and cell cycle arrest were detected in cells that were irradiated with UVA only. Pretreatment with ferulic acid significantly increased the proliferation and cell cycle progression in HDFs. Moreover, ferulic acid pretreatment produced antioxidant effects such as reduced DCF intensity, and affected SOD1 and CAT mRNA expression. These effects were also demonstrated in the analysis of cell senescence, promoter activity, expression of senescent markers, and DNA repair. Conclusion These results demonstrate that ferulic acid exerts protective effects on UVA-induced cell damages via anti-oxidant and stress-inducible cellular mechanisms in HDFs. PMID:27904274

  2. Induction and processing of oxidative clustered DNA lesions in 56Fe-ion-irradiated human monocytes.

    PubMed

    Tsao, Doug; Kalogerinis, Peter; Tabrizi, Isla; Dingfelder, Michael; Stewart, Robert D; Georgakilas, Alexandros G

    2007-07-01

    Space and cosmic radiation is characterized by energetic heavy ions of high linear energy transfer (LET). Although both low- and high-LET radiations can create oxidative clustered DNA lesions and double-strand breaks (DSBs), the local complexity of oxidative clustered DNA lesions tends to increase with increasing LET. We irradiated 28SC human monocytes with doses from 0-10 Gy of (56)Fe ions (1.046 GeV/ nucleon, LET = 148 keV/microm) and determined the induction and processing of prompt DSBs and oxidative clustered DNA lesions using pulsed-field gel electrophoresis (PFGE) and Number Average Length Analysis (NALA). The (56)Fe ions produced decreased yields of DSBs (10.9 DSB Gy(-1) Gbp(-1)) and clusters (1 DSB: approximately 0.8 Fpg clusters: approximately 0.7 Endo III clusters: approximately 0.5 Endo IV clusters) compared to previous results with (137)Cs gamma rays. The difference in the relative biological effectiveness (RBE) of the measured and predicted DSB yields may be due to the formation of spatially correlated DSBs (regionally multiply damaged sites) which result in small DNA fragments that are difficult to detect with the PFGE assay. The processing data suggest enhanced difficulty compared with gamma rays in the processing of DSBs but not clusters. At the same time, apoptosis is increased compared to that seen with gamma rays. The enhanced levels of apoptosis observed after exposure to (56)Fe ions may be due to the elimination of cells carrying high levels of persistent DNA clusters that are removed only by cell death and/or "splitting" during DNA replication.

  3. Therapeutic Electromagnetic Field (TEMF) and gamma irradiation on human breast cancer xenograft growth, angiogenesis and metastasis

    PubMed Central

    Cameron, Ivan L; Sun, Lu-Zhe; Short, Nicholas; Hardman, W Elaine; Williams, C Douglas

    2005-01-01

    Background The effects of a rectified semi-sinewave signal (15 mT amplitude, 120 pulses per second, EMF Therapeutics, Inc.) (TEMF) alone and in combination with gamma irradiation (IR) therapy in nude mice bearing a human MDA MB231 breast cancer xenograft were tested. Green fluorescence protein transfected cancer cells were injected into the mammary fat pad of young female mice. Six weeks later, mice were randomly divided into four treatment groups: untreated controls; 10 minute daily TEMF; 200 cGy of IR every other day (total 800 cGy); IR plus daily TEMF. Some mice in each group were euthanized 24 hours after the end of IR. TEMF treatment continued for 3 additional weeks. Tumor sections were stained for: endothelial cells with CD31 and PAS or hypoxia inducible factor 1α (HIF). Results Most tumors <35 mm3 were white but tumors >35 mm3 were pink and had a vascularized capsule. The cortex within 100 microns of the capsule had little vascularization. Blood vessels, capillaries, and endothelial pseudopods were found at >100 microns from the capsule (subcortex). Tumors >35 mm3 treated with IR 24 hours previously or with TEMF had decreased blood vessels in the subcortex and more endothelial pseudopods projecting into hypoxic, HIF positive areas than tumors from the control group. Mice that received either IR or TEMF had significantly fewer lung metastatic sites and slower tumor growth than did untreated mice. No harmful side effects were attributed to TEMF. Conclusion TEMF therapy provided a safe means for retarding tumor vascularization, growth and metastasis. PMID:16045802

  4. Protective activity of C-geranylflavonoid analogs from Paulownia tomentosa against DNA damage in 137Cs irradiated AHH-1 cells.

    PubMed

    Moon, Hyung-In; Jeong, Min Ho; Jo, Wol Soon

    2014-09-01

    Radiotherapy is an important form of treatment for a wide range of cancers, but it can damage DNA and cause adverse effects. We investigated if the diplacone analogs of P. tomentosa were radio-protective in a human lymphoblastoid cell line (AHH-1). Four geranylated flavonoids, diplacone, 3'-O-methyl-5'-hydroxydiplacone, 3'-O-methyl-5'-O-methyldiplacone and 3'-O-methyldiplacol, were tested for their antioxidant and radio-protective effects. Diplacone analogs effectively scavenged free radicals and inhibited radiation-induced DNA strand breaks in vitro. They significantly decreased levels of reactive oxygen species and cellular DNA damage in 2 Gy-irradiated AHH-1 cells. Glutathione levels and superoxide dismutase activity in irradiated AHH-1 cells increased significantly after treatment with these analogs. The enhanced biological anti-oxidant activity and radioprotective activity of diplacone analogs maintained the survival of irradiated AHH-1 cells in a clonogenic assay. These data suggest that diplacone analogs may protect healthy tissue surrounding tumor cells during radiotherapy to ensure better control of radiotherapy and allow higher doses of radiotherapy to be employed.

  5. Cyclobutane pyrimidine dimer formation and p53 production in human skin after repeated UV irradiation.

    PubMed

    Yamaguchi, Yuji; Coelho, Sergio G; Zmudzka, Barbara Z; Takahashi, Kaoruko; Beer, Janusz Z; Hearing, Vincent J; Miller, Sharon A

    2008-11-01

    Substantial differences in DNA damage caused by a single UV irradiation were found in our previous study on skin with different levels of constitutive pigmentation. In this study, we assessed whether facultative pigmentation induced by repeated UV irradiation is photoprotective. Three sites on the backs of 21 healthy subjects with type II-III skin were irradiated at 100-600 J/m(2) every 2-7 days over a 4- to 5-week period. The three sites received different cumulative doses of UV (1900, 2900 or 4200 J/m(2)) and were biopsied 1 day after the last irradiation. Biomarkers examined included pigment content assessed by Fontana-Masson staining, melanocyte function by expression of melanocyte-specific markers, DNA damage as cyclobutane pyrimidine dimers (CPD), nuclear accumulation of p53, apoptosis determined by TUNEL assay, and levels of p21 and Ser46-phosphorylated p53. Increases in melanocyte function and density, and in levels of apoptosis were similar among the 3 study sites irradiated with different cumulative UV doses. Levels of CPD decreased while the number of p53-positive cells increased as the cumulative dose of UV increased. These results suggest that pigmentation induced in skin by repeated UV irradiation protects against subsequent UV-induced DNA damage but not as effectively as constitutive pigmentation.

  6. Reduction of interleukin-6 expression in human synoviocytes and rheumatoid arthritis rat joints by linear polarized near infrared light (Superlizer) irradiation.

    PubMed

    Araki, Hidefumi; Imaoka, Asayo; Kuboyama, Noboru; Abiko, Yoshimitsu

    2011-01-01

    Rheumatoid arthritis (RA) is a systemic autoimmune disorder that involves inflammation and pain of joints. Low-level laser irradiation is being evaluated for treating RA, however, the effectiveness of linear polarized near infrared light (SuperLizer; SL) irradiation is unclear. It has been reported that interleukin 6 (IL-6) plays a key role in the progression of RA. In our previous study, using DNA microarray analysis, we examined the gene expression profiling of human rheumatoid fibroblast-like synoviocyte MH7A in response to IL-1ß administration and SL irradiation. As a result, IL-6 was listed in altered gene as increased by IL-1ß and decreased by SL irradiation. The reduction of IL-6 gene expression in MH7A by SL irradiation was confirmed by reverse transcription polymerase chain reaction (RT-PCR) and real-time PCR. Effect of SL irradiation on the RA inflammation in the collagen induced arthritis (CIA) rats was also studied by measuring temperature. IL-6 production in knee joint of rats was analyzed by immunohisto-chemistry. Scatter plot analysis demonstrated that an increase in IL-6 gene expression by IL-1ß was reduced by SL irradiation. The reduction of IL-6 mRNA level by SL irradiation was successfully confirmed by RT-PCR and real-time PCR. SL irradiation treated CIA rat decreased the temperature of knee joints. The immunohistochemical analysis demonstrated a strong IL-6 staining in synovial membrane tissue of CIA rat joint, and SL irradiation significantly reduced the staining. Since IL-6 has been identified to be an important proinflarnmatory cytokine in the pathogenesis of RA, the reduction of IL-6 expression is one of mechanisms in reduction of inflammation in RA joints by SL irradiation suggesting that SL irradiation may be useful for RA therapy. SL irradiation reduced IL-6 gene expression in MH7A, and reduced inflammation and IL-6 protein expression in knee joint of CIA rats.

  7. Ultraviolet irradiation induces CYR61/CCN1, a mediator of collagen homeostasis, through activation of transcription factor AP-1 in human skin fibroblasts.

    PubMed

    Quan, Taihao; Qin, Zhaoping; Xu, Yiru; He, Tianyuan; Kang, Sewon; Voorhees, John J; Fisher, Gary J

    2010-06-01

    UV irradiation from the sun elevates the production of collagen-degrading matrix metalloproteinases (MMPs) and reduces the production of new collagen. This imbalance of collagen homeostasis impairs the structure and function of the dermal collagenous extracellular matrix (ECM), thereby promoting premature skin aging (photoaging). We report here that aberrant dermal collagen homeostasis in UV-irradiated human skin is mediated in part by a CCN-family member, cysteine-rich protein-61 (CYR61/CCN1). CYR61 is significantly elevated in acutely UV-irradiated human skin in vivo, and UV-irradiated human skin fibroblasts. Knockdown of CYR61 significantly attenuates UV irradiation-induced inhibition of type-I procollagen and upregulation of MMP-1. Determination of CYR61 mRNA and protein indicates that the primary mechanism of CYR61 induction by UV irradiation is transcriptional. Analysis of CYR61 proximal promoter showed that a sequence conforming to the consensus binding site for transcription factor activator protein-1 (AP-1) is required for promoter activity. UV irradiation increased the binding of AP-1-family members c-Jun and c-Fos to this AP-1 site. Furthermore, functional blockade of c-Jun or knockdown of c-Jun significantly reduced the UV irradiation-induced activation of CYR61 promoter and CYR61 gene expression. These data show that CYR61 is transcriptionally regulated by UV irradiation through transcription factor AP-1, and mediates altered collagen homeostasis that occurs in response to UV irradiation in human skin fibroblasts.

  8. Three-dimensional Invasion of Human Glioblastoma Cells Remains Unchanged by X-ray and Carbon Ion Irradiation In Vitro

    SciTech Connect

    Eke, Iris; Storch, Katja; Kaestner, Ina; Vehlow, Anne; Faethe, Christina; Mueller-Klieser, Wolfgang; Taucher-Scholz, Gisela; Temme, Achim; Schackert, Gabriele

    2012-11-15

    Purpose: Cell invasion represents one of the major determinants that treatment has failed for patients suffering from glioblastoma. Contrary findings have been reported for cell migration upon exposure to ionizing radiation. Here, the migration and invasion capability of glioblastoma cells on and in collagen type I were evaluated upon irradiation with X-rays or carbon ions. Methods and Materials: Migration on and invasion in collagen type I were evaluated in four established human glioblastoma cell lines exposed to either X-rays or carbon ions. Furthermore, clonogenic radiation survival, proliferation (5-bromo-2-deoxyuridine positivity), DNA double-strand breaks ({gamma}H2AX/53BP1-positive foci), and expression of invasion-relevant proteins (eg, {beta}1 integrin, FAK, MMP2, and MMP9) were explored. Migration and invasion assays for primary glioblastoma cells also were carried out with X-ray irradiation. Results: Neither X-ray nor carbon ion irradiation affected glioblastoma cell migration and invasion, a finding similarly observed in primary glioblastoma cells. Intriguingly, irradiated cells migrated unhampered, despite DNA double-strand breaks and reduced proliferation. Clonogenic radiation survival was increased when cells had contact with extracellular matrix. Specific inhibition of the {beta}1 integrin or proliferation-associated signaling molecules revealed a critical function of JNK, PI3K, and p38 MAPK in glioblastoma cell invasion. Conclusions: These findings indicate that X-rays and carbon ion irradiation effectively reduce proliferation and clonogenic survival without modifying the migration and invasion ability of glioblastoma cells in a collagen type I environment. Addition of targeted agents against members of the MAPK and PI3K signaling axis to conventional chemoradiation therapy seems potentially useful to optimize glioblastoma therapy.

  9. DNA repair in modeled microgravity: double strand break rejoining activity in human lymphocytes irradiated with gamma-rays.

    PubMed

    Mognato, Maddalena; Girardi, Cristina; Fabris, Sonia; Celotti, Lucia

    2009-04-26

    Cell response to ionising radiation depends, besides on genetic and physiological features of the biological systems, on environmental conditions occurring during DNA repair. Many data showed that microgravity, experienced by astronauts during space flights or modeled on Earth, causes apoptosis, cytoskeletal alteration, cell growth inhibition, increased frequency of mutations and chromosome aberrations. In this study, we analysed the progression of the rejoining of double strand breaks (DSBs) in human peripheral blood lymphocytes (PBLs) irradiated with gamma-rays and incubated in static condition (1 g) or in modeled microgravity (MMG). gamma-H2AX foci formation and disappearance, monitored during the repair incubation, showed that the kinetics of DSBs rejoining was different in the two gravity conditions. The fraction of foci-positive cells decreased slower in MMG than in 1 g at 6 and 24 h after irradiation (P<0.01) and the mean number of gamma-H2AX foci per nucleus was significantly higher in MMG than in 1g at the same time-points (P<0.001). In the same samples we determined apoptotic level and the rate of DSB rejoining during post-irradiation incubation. A significant induction of apoptosis was observed in MMG at 24 h after irradiation (P<0.001), whereas at shorter times the level of apoptosis was slightly higher in MMG respect to 1 g. In accordance with the kinetics of gamma-H2AX foci, the slower rejoining of radiation-induced DSBs in MMG was observed by DNA fragmentation analyses during the repair incubation; the data of pulsed-field gel electrophoresis assay showed that the fraction of DNA released in the gel was significantly higher in PBL incubated in MMG after irradiation with respect to cells maintained in 1 g. Our results provide evidences that MMG incubation during DNA repair delayed the rate of radiation-induced DSB rejoining, and increased, as a consequence, the genotoxic effects of ionising radiation.

  10. Morphological and Structural Changes on Human Dental Enamel After Er:YAG Laser Irradiation: AFM, SEM, and EDS Evaluation

    PubMed Central

    Rodríguez-Vilchis, Laura Emma; Olea-Mejìa, Oscar Fernando; Sánchez-Flores, Ignacio; Centeno-Pedraza, Claudia

    2011-01-01

    Abstract Objective: The purpose of this study was to evaluate, using atomic force microscopy (AFM), scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS), the morphological and structural changes of the enamel after irradiation with the Er:YAG laser. Background data: A previous study showed that subablative Er:YAG laser irradiation produced undesirable morphological changes on the enamel surface, such as craters and cracks; however, the enamel acid resistance was not increased. Methods: Fifty-two samples of human enamel were divided into four groups (n = 13): Group I was the control (no laser irradiation), whereas Groups II, III, and IV were irradiated with the Er:YAG 100 mJ (12.7 J/cm2), 100 mJ (7.5 J/cm2), and 150 mJ (11 J/cm2), respectively, at 10 Hz with water spray. The morphological changes were observed by AFM and SEM. The weight percentages (wt%) of calcium (Ca), phosphorus (P), oxygen (O) and chlorine (Cl) were determined in the resultant craters and their periphery using EDS. Kruskal–Wallis and Mann–Whitney U tests were performed (p ≤ 0.05) to distinguish significant differences among the groups. Results: The AFM images showed cracks with depths between 250 nm and 750 nm for Groups II and IV, respectively, and the widths of these cracks were 5.37 μm and 2.58 μm. The interior of the cracks showed a rough surface. The SEM micrographs revealed morphological changes. Significant differences were detected in Ca, P, and Cl in the crater and its periphery. Conclusions: AFM observations showed triangular-shaped cracks, whereas craters and cracks were evident by SEM in all irradiated samples. It was not possible to establish a characteristic chemical pattern in the craters. PMID:21417912

  11. Morphological and structural changes on human dental enamel after Er:YAG laser irradiation: AFM, SEM, and EDS evaluation.

    PubMed

    Rodríguez-Vilchis, Laura Emma; Contreras-Bulnes, Rosalía; Olea-Mejìa, Oscar Fernando; Sánchez-Flores, Ignacio; Centeno-Pedraza, Claudia

    2011-07-01

    The purpose of this study was to evaluate, using atomic force microscopy (AFM), scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS), the morphological and structural changes of the enamel after irradiation with the Er:YAG laser. A previous study showed that subablative Er:YAG laser irradiation produced undesirable morphological changes on the enamel surface, such as craters and cracks; however, the enamel acid resistance was not increased. Fifty-two samples of human enamel were divided into four groups (n = 13): Group I was the control (no laser irradiation), whereas Groups II, III, and IV were irradiated with the Er:YAG 100 mJ (12.7 J/cm(2)), 100 mJ (7.5 J/cm(2)), and 150 mJ (11 J/cm(2)), respectively, at 10 Hz with water spray. The morphological changes were observed by AFM and SEM. The weight percentages (wt%) of calcium (Ca), phosphorus (P), oxygen (O) and chlorine (Cl) were determined in the resultant craters and their periphery using EDS. Kruskal-Wallis and Mann-Whitney U tests were performed (p ≤ 0.05) to distinguish significant differences among the groups. The AFM images showed cracks with depths between 250 nm and 750 nm for Groups II and IV, respectively, and the widths of these cracks were 5.37 μm and 2.58 μm. The interior of the cracks showed a rough surface. The SEM micrographs revealed morphological changes. Significant differences were detected in Ca, P, and Cl in the crater and its periphery. AFM observations showed triangular-shaped cracks, whereas craters and cracks were evident by SEM in all irradiated samples. It was not possible to establish a characteristic chemical pattern in the craters.

  12. Three-dimensional invasion of human glioblastoma cells remains unchanged by X-ray and carbon ion irradiation in vitro.

    PubMed

    Eke, Iris; Storch, Katja; Kästner, Ina; Vehlow, Anne; Faethe, Christina; Mueller-Klieser, Wolfgang; Taucher-Scholz, Gisela; Temme, Achim; Schackert, Gabriele; Cordes, Nils

    2012-11-15

    Cell invasion represents one of the major determinants that treatment has failed for patients suffering from glioblastoma. Contrary findings have been reported for cell migration upon exposure to ionizing radiation. Here, the migration and invasion capability of glioblastoma cells on and in collagen type I were evaluated upon irradiation with X-rays or carbon ions. Migration on and invasion in collagen type I were evaluated in four established human glioblastoma cell lines exposed to either X-rays or carbon ions. Furthermore, clonogenic radiation survival, proliferation (5-bromo-2-deoxyuridine positivity), DNA double-strand breaks (γH2AX/53BP1-positive foci), and expression of invasion-relevant proteins (eg, β1 integrin, FAK, MMP2, and MMP9) were explored. Migration and invasion assays for primary glioblastoma cells also were carried out with X-ray irradiation. Neither X-ray nor carbon ion irradiation affected glioblastoma cell migration and invasion, a finding similarly observed in primary glioblastoma cells. Intriguingly, irradiated cells migrated unhampered, despite DNA double-strand breaks and reduced proliferation. Clonogenic radiation survival was increased when cells had contact with extracellular matrix. Specific inhibition of the β1 integrin or proliferation-associated signaling molecules revealed a critical function of JNK, PI3K, and p38 MAPK in glioblastoma cell invasion. These findings indicate that X-rays and carbon ion irradiation effectively reduce proliferation and clonogenic survival without modifying the migration and invasion ability of glioblastoma cells in a collagen type I environment. Addition of targeted agents against members of the MAPK and PI3K signaling axis to conventional chemoradiation therapy seems potentially useful to optimize glioblastoma therapy. Copyright © 2012 Elsevier Inc. All rights reserved.

  13. Pentosidine in advanced glycation end-products (AGEs) during UVA irradiation generates active oxygen species and impairs human dermal fibroblasts.

    PubMed

    Okano, Y; Masaki, H; Sakurai, H

    2001-08-01

    Our previous study reported that advanced glycation end-products (AGE)-modified BSA produced active oxygen species, *O2-, H2O2, and *OH under UVA irradiation and enhanced the cytotoxicity of UVA light. We examined whether pentosidine in AGE-modified BSA was involved in one of the mechanisms generating the active oxygen species. In biological investigations, fibroblasts exposed to UVA (20 J/cm2) in the presence of pentosidine-rich compounds (PRCs), which were prepared with L-arginine, L-lysine and glucose, showed a time-dependent leakage of the cytosolic enzyme LDH. In addition, release of LDH was suppressed by addition of DMSO and deferoxamine under UVA irradiation. From these results, it was determined that PRCs exposed to UVA damaged the plasma membrane of human dermal fibroblasts due to the conversion of *OH from H2O2 via a Fenton-like reaction. These features of PRCs exposed to UVA were consistent with those of AGE-modified BSA. In an ESR study, PRCs under UVA irradiation yielded DMPO-OH (DMPO-OH adduct) using DMPO as a spin-trapping reagent. *O2- generation from UVA-irradiated PRCs was also indicated by the combination of NBT reduction and SOD. When PRCs were exposed to UVA light controlled with a long-pass filter, WG-360, it was found that their production of *O2- was prohibited less than 50% in the NBT reduction assay. The *O2- production profile of PRCs depending on the wavelength of UVA light was similar to that of AGE-modified BSA. Furthermore, it was found that the H2O2 level was increased by PRCs exposed to UVA. These results indicated that pentosidine is an important factor of AGE-modified BSA in active oxygen generation under UVA irradiation.

  14. Human neural stem cell transplantation provides long-term restoration of neuronal plasticity in the irradiated hippocampus.

    PubMed

    Acharya, Munjal M; Rosi, Susanna; Jopson, Timothy; Limoli, Charles L

    2015-01-01

    For the majority of CNS malignancies, radiotherapy provides the best option for forestalling tumor growth, but is frequently associated with debilitating and progressive cognitive dysfunction. Despite the recognition of this serious side effect, satisfactory long-term solutions are not currently available and have prompted our efforts to explore the potential therapeutic efficacy of cranial stem cell transplants. We have demonstrated that intrahippocampal transplantation of human neural stem cells (hNSCs) can provide long-lasting cognitive benefits using an athymic rat model subjected to cranial irradiation. To explore the possible mechanisms underlying the capability of engrafted cells to ameliorate radiation-induced cognitive dysfunction we analyzed the expression patterns of the behaviorally induced activity-regulated cytoskeleton-associated protein (Arc) in the hippocampus at 1 and 8 months postgrafting. While immunohistochemical analyses revealed a small fraction (4.5%) of surviving hNSCs in the irradiated brain that did not express neuronal or astroglial makers, hNSC transplantation impacted the irradiated microenvironment of the host brain by promoting the expression of Arc at both time points. Arc is known to play key roles in the neuronal mechanisms underlying long-term synaptic plasticity and memory and provides a reliable marker for detecting neurons that are actively engaged in spatial and contextual information processing associated with memory consolidation. Cranial irradiation significantly reduced the number of pyramidal (CA1) and granule neurons (DG) expressing behaviorally induced Arc at 1 and 8 months postirradiation. Transplantation of hNSCs restored the expression of plasticity-related Arc in the host brain to control levels. These findings suggest that hNSC transplantation promotes the long-term recovery of host hippocampal neurons and indicates that one mechanism promoting the preservation of cognition after irradiation involves trophic

  15. Spectral responses of the human circadian system depend on the irradiance and duration of exposure to light.

    PubMed

    Gooley, Joshua J; Rajaratnam, Shantha M W; Brainard, George C; Kronauer, Richard E; Czeisler, Charles A; Lockley, Steven W

    2010-05-12

    In humans, modulation of circadian rhythms by light is thought to be mediated primarily by melanopsin-containing retinal ganglion cells, not rods or cones. Melanopsin cells are intrinsically blue light-sensitive but also receive input from visual photoreceptors. We therefore tested in humans whether cone photoreceptors contribute to the regulation of circadian and neuroendocrine light responses. Dose-response curves for melatonin suppression and circadian phase resetting were constructed in subjects exposed to blue (460 nm) or green (555 nm) light near the onset of nocturnal melatonin secretion. At the beginning of the intervention, 555-nm light was equally effective as 460-nm light at suppressing melatonin, suggesting a significant contribution from the three-cone visual system (lambda(max) = 555 nm). During the light exposure, however, the spectral sensitivity to 555-nm light decayed exponentially relative to 460-nm light. For phase-resetting responses, the effects of exposure to low-irradiance 555-nm light were too large relative to 460-nm light to be explained solely by the activation of melanopsin. Our findings suggest that cone photoreceptors contribute substantially to nonvisual responses at the beginning of a light exposure and at low irradiances, whereas melanopsin appears to be the primary circadian photopigment in response to long-duration light exposure and at high irradiances. These results suggest that light therapy for sleep disorders and other indications might be optimized by stimulating both photoreceptor systems.

  16. Molecular Signatures of Cardiac Defects in Down Syndrome Lymphoblastoid Cell Lines Suggest Altered Ciliome and Hedgehog Pathways

    PubMed Central

    Ripoll, Clémentine; Rivals, Isabelle; Ait Yahya-Graison, Emilie; Dauphinot, Luce; Paly, Evelyne; Mircher, Clothilde; Ravel, Aimé; Grattau, Yann; Bléhaut, Henri; Mégarbane, André; Dembour, Guy; de Fréminville, Bénédicte; Touraine, Renaud; Créau, Nicole; Potier, Marie Claude; Delabar, Jean Maurice

    2012-01-01

    Forty percent of people with Down syndrome exhibit heart defects, most often an atrioventricular septal defect (AVSD) and less frequently a ventricular septal defect (VSD) or atrial septal defect (ASD). Lymphoblastoid cell lines (LCLs) were established from lymphocytes of individuals with trisomy 21, the chromosomal abnormality causing Down syndrome. Gene expression profiles generated from DNA microarrays of LCLs from individuals without heart defects (CHD−; n = 22) were compared with those of LCLs from patients with cardiac malformations (CHD+; n = 21). After quantile normalization, principal component analysis revealed that AVSD carriers could be distinguished from a combined group of ASD or VSD (ASD+VSD) carriers. From 9,758 expressed genes, we identified 889 and 1,016 genes differentially expressed between CHD− and AVSD and CHD− and ASD+VSD, respectively, with only 119 genes in common. A specific chromosomal enrichment was found in each group of affected genes. Among the differentially expressed genes, more than 65% are expressed in human or mouse fetal heart tissues (GEO dataset). Additional LCLs from new groups of AVSD and ASD+VSD patients were analyzed by quantitative PCR; observed expression ratios were similar to microarray results. Analysis of GO categories revealed enrichment of genes from pathways regulating clathrin-mediated endocytosis in patients with AVSD and of genes involved in semaphorin-plexin-driven cardiogenesis and the formation of cytoplasmic microtubules in patients with ASD-VSD. A pathway-oriented search revealed enrichment in the ciliome for both groups and a specific enrichment in Hedgehog and Jak-stat pathways among ASD+VSD patients. These genes or related pathways are therefore potentially involved in normal cardiogenesis as well as in cardiac malformations observed in individuals with trisomy 21. PMID:22912673

  17. Transcriptome profiling of UPF3B/NMD-deficient lymphoblastoid cells from patients with various forms of intellectual disability

    PubMed Central

    Nguyen, LS; Jolly, L; Shoubridge, C; Chan, WK; Huang, L; Laumonnier, F; Raynaud, M; Hackett, A; Field, M; Rodriguez, J; Srivastava, AK; Lee, Y; Long, R; Addington, AM; Rapoport, JL; Suren, S; Hahn, CN; Gamble, J; Wilkinson, MF; Corbett, MA; Gecz, J

    2014-01-01

    The nonsense-mediated mRNA decay (NMD) pathway was originally discovered by virtue of its ability to rapidly degrade aberrant mRNAs with premature termination codons. More recently, it was shown that NMD also directly regulates subsets of normal transcripts, suggesting that NMD has roles in normal biological processes. Indeed, several NMD factors have been shown to regulate neurological events (for example, neurogenesis and synaptic plasticity) in numerous vertebrate species. In man, mutations in the NMD factor gene UPF3B, which disrupts a branch of the NMD pathway, cause various forms of intellectual disability (ID). Using Epstein Barr virus—immortalized B cells, also known as lymphoblastoid cell lines (LCLs), from ID patients that have loss-of-function mutations in UPF3B, we investigated the genome-wide consequences of compromised NMD and the role of NMD in neuronal development and function. We found that ~5% of the human transcriptome is impacted in UPF3B patients. The UPF3B paralog, UPF3A, is stabilized in all UPF3B patients, and partially compensates for the loss of UPF3B function. Interestingly, UPF3A protein, but not mRNA, was stabilised in a quantitative manner that inversely correlated with the severity of patients’ phenotype. This suggested that the ability to stabilize the UPF3A protein is a crucial modifier of the neurological symptoms due to loss of UPF3B. We also identified ARHGAP24, which encodes a GTPase-activating protein, as a canonical target of NMD, and we provide evidence that deregulation of this gene inhibits axon and dendrite outgrowth and branching. Our results demonstrate that the UPF3B-dependent NMD pathway is a major regulator of the transcriptome and that its targets have important roles in neuronal cells. PMID:22182939

  18. All-trans Retinoic Acid Upregulates Reduced CD38 Transcription in Lymphoblastoid Cell Lines from Autism Spectrum Disorder

    PubMed Central

    Riebold, Mathias; Mankuta, David; Lerer, Elad; Israel, Salomon; Zhong, Songfa; Nemanov, Luba; Monakhov, Mikhail V; Levi, Shlomit; Yirmiya, Nurit; Yaari, Maya; Malavasi, Fabio; Ebstein, Richard P

    2011-01-01

    Deficits in social behavior in mice lacking the CD38 gene have been attributed to impaired secretion of oxytocin. In humans, similar deficits in social behavior are associated with autistic spectrum disorder (ASD), for which genetic variants of CD38 have been pinpointed as provisional risk factors. We sought to explore, in an in vitro model, the feasibility of the theory that restoring the level of CD38 in ASD patients could be of potential clinical benefit. CD38 transcription is highly sensitive to several cytokines and vitamins. One of these, all-trans retinoic acid (ATRA), a known inducer of CD38, was added during cell culture and tested on a large sample of N = 120 lymphoblastoid cell (LBC) lines from ASD patients and their parents. Analysis of CD38 mRNA levels shows that ATRA has an upmodulatory potential on LBC derived from ASD patients as well as from their parents. The next crucial issue addressed in our study was the relationship between levels of CD38 expression and psychological parameters. The results obtained indicate a positive correlation between CD38 expression levels and patient scores on the Vineland Adaptive Behavior Scale. In addition, analysis of the role of genetic polymorphisms in the dynamics of the molecule revealed that the genotype of a single-nucleotide polymorphism (rs6449182; C>G variation) in the CpG island of intron 1, harboring the retinoic-acid response element, exerts differential roles in CD38 expression in ASD and in parental LBC. In conclusion, our results provide an empirical basis for the development of a pharmacological ASD treatment strategy based on retinoids. PMID:21528155

  19. All-trans retinoic acid upregulates reduced CD38 transcription in lymphoblastoid cell lines from Autism spectrum disorder.

    PubMed

    Riebold, Mathias; Mankuta, David; Lerer, Elad; Israel, Salomon; Zhong, Songfa; Nemanov, Luba; Monakhov, Mikhail V; Levi, Shlomit; Yirmiya, Nurit; Yaari, Maya; Malavasi, Fabio; Ebstein, Richard P

    2011-01-01

    Deficits in social behavior in mice lacking the CD38 gene have been attributed to impaired secretion of oxytocin. In humans, similar deficits in social behavior are associated with autistic spectrum disorder (ASD), for which genetic variants of CD38 have been pinpointed as provisional risk factors. We sought to explore, in an in vitro model, the feasibility of the theory that restoring the level of CD38 in ASD patients could be of potential clinical benefit. CD38 transcription is highly sensitive to several cytokines and vitamins. One of these, all-trans retinoic acid (ATRA), a known inducer of CD38, was added during cell culture and tested on a large sample of N = 120 lymphoblastoid cell (LBC) lines from ASD patients and their parents. Analysis of CD38 mRNA levels shows that ATRA has an upmodulatory potential on LBC derived from ASD patients as well as from their parents. The next crucial issue addressed in our study was the relationship between levels of CD38 expression and psychological parameters. The results obtained indicate a positive correlation between CD38 expression levels and patient scores on the Vineland Adaptive Behavior Scale. In addition, analysis of the role of genetic polymorphisms in the dynamics of the molecule revealed that the genotype of a single-nucleotide polymorphism (rs6449182; C>G variation) in the CpG island of intron 1, harboring the retinoic-acid response element, exerts differential roles in CD38 expression in ASD and in parental LBC. In conclusion, our results provide an empirical basis for the development of a pharmacological ASD treatment strategy based on retinoids.

  20. Inhibiting the Aurora B Kinase Potently Suppresses Repopulation During Fractionated Irradiation of Human Lung Cancer Cell Lines

    SciTech Connect

    Sak, Ali; Stuschke, Martin; Groneberg, Michael; Kuebler, Dennis; Poettgen, Christoph; Eberhardt, Wilfried E.E.

    2012-10-01

    Purpose: The use of molecular-targeted agents during radiotherapy of non-small-cell lung cancer (NSCLC) is a promising strategy to inhibit repopulation, thereby improving therapeutic outcome. We assessed the combined effectiveness of inhibiting Aurora B kinase and irradiation on human NSCLC cell lines in vitro. Methods and Materials: NSCLC cell lines were exposed to concentrations of AZD1152-hydroxyquinazoline pyrazol anilide (AZD1152-HQPA) inhibiting colony formation by 50% (IC50{sub clone}) in combination with single dose irradiation or different fractionation schedules using multiple 2-Gy fractions per day up to total doses of 4-40 Gy. The total irradiation dose required to control growth of 50% of the plaque monolayers (TCD50) was determined. Apoptosis, G2/M progression, and polyploidization were also analyzed. Results: TCD50 values after single dose irradiation were similar for the H460 and H661 cell lines with 11.4 {+-} 0.2 Gy and 10.7 {+-} 0.3 Gy, respectively. Fractionated irradiation using 3 Multiplication-Sign 2 Gy/day, 2 Multiplication-Sign 2 Gy/day, and 1 Multiplication-Sign 2 Gy/day schedules significantly increased TCD50 values for both cell lines grown as plaque monolayers with increasing radiation treatment time. This could be explained by a repopulation effect per day that counteracts 75 {+-} 8% and 27 {+-} 6% of the effect of a 2-Gy fraction in H460 and H661 cells, respectively. AZD1152-HQPA treatment concomitant to radiotherapy significantly decreased the daily repopulation effect (H460: 28 {+-} 5%, H661: 10 {+-} 4% of a 2-Gy fraction per day). Treatment with IC50{sub clone} AZD1152-HPQA did not induce apoptosis, prolong radiation-induced G2 arrest, or delay cell cycle progression before the spindle check point. However, polyploidization was detected, especially in cell lines without functional p53. Conclusions: Inhibition of Aurora B kinase with low AZD1152-HQPA concentrations during irradiation of NSCLC cell lines affects repopulation during

  1. Quantitative measurement of DNA strand breaks and repair in. gamma. -irradiated human leukocytes from normal and ataxia telangiectasia donors

    SciTech Connect

    Thierry, D.; Rigaud, O.; Duranton, I.; Moustacchi, E.; Magdelenat, H.

    1985-06-01

    Fluorimetric analysis of DNA unwinding, which allows measurement of DNA strand breaks in human leukocytes, has been optimized by reducing the amount of cells required for the test and by modifying the DNA alkali unwinding conditions. The permitted measurement of DNA strand-break induction in cells irradiated with low (0.5-7 Gy) or high doses (5-20 Gy) of ..gamma.. rays. Linear dose-response curves were obtained for both dose ranges. Presence of cysteamine during irradiation caused a decrease in the extent of DNA strand breaks. The kinetics of the DNA standard-break rejoining process appeared to be biphasic over the dose range of 2-20 Gy when plotted on a linear vs linear axis (percentage of damage as a function of time). No difference in the level of DNA strand breaks and the rate of repair of these breaks was observed between leukocytes from three ataxia telangiectasia patients and those from normal donors.

  2. Caspase-independent cell death without generation of reactive oxygen species in irradiated MOLT-4 human leukemia cells.

    PubMed

    Yoshida, Kengo; Kubo, Yoshiko; Kusunoki, Yoichiro; Morishita, Yukari; Nagamura, Hiroko; Hayashi, Ikue; Kyoizumi, Seishi; Seyama, Toshio; Nakachi, Kei; Hayashi, Tomonori

    2009-01-01

    To improve our understanding of ionizing radiation effects on immune cells, we investigated steps leading to radiation-induced cell death in MOLT-4, a thymus-derived human leukemia cell. After exposure of MOLT-4 cells to 4 Gy of X-rays, irradiated cells sequentially showed increase in intracellular reactive oxygen species (ROS), decrease in mitochondrial membrane potential, and eventually apoptotic cell death. In the presence of the caspase inhibitor z-VAD-fmk, irradiated cells exhibited necrotic characteristics such as mitochondrial swelling instead of apoptosis. ROS generation was not detected during this necrotic cell death process. These results indicate that radiation-induced apoptosis in MOLT-4 cells requires elevation of intracellular ROS as well as activation of a series of caspases, whereas the cryptic necrosis program--which is independent of intracellular ROS generation and caspase activation--is activated when the apoptosis pathway is blocked.

  3. Changes in S100A8 expression in UV-irradiated and aged human skin in vivo.

    PubMed

    Lee, Young Mee; Kim, Yeon Kyung; Eun, Hee Chul; Chung, Jin Ho

    2009-08-01

    S100A8, a calcium-binding protein, is associated with keratinocyte differentiation, inflammation and wound healing. S100A8 is induced by various skin stresses and diseases, which suggests that S100A8 plays a role in those processes. However, it has not been reported how the expression of S100A8 is affected during skin aging or whether S100A8 plays a role in the skin aging process. In this study, we investigated the changes in S100A8 mRNA and protein following acute UV irradiation to human buttock skin and by intrinsic aging and photoaging in human sun-protected (upper-inner arm) and sun-exposed (forearm) skin of elderly subjects. Real-time PCR, western blot and immunohistochemical staining analyses of UV-irradiated young buttock skin revealed that S100A8 protein expression was increased at 24 h (3.0-fold) and 48 h (4.4-fold) after UV irradiation. S100A8 mRNA and protein were more highly expressed by 2.3- and 4.0-fold, respectively, in the sun-protected skin of elderly people than in that of young people. In addition, the sun-exposed skin of elderly expressed more S100A8 mRNA and protein than the sun-protected skin of the same individuals. In immunohistochemical staining, facial (photoaged) skin > or = 72 years showed higher epidermal expression of S100A8 than that of the other age groups. Based on the above results, our data suggest that the expression of S100A8 is affected by acute UV irradiation, intrinsic aging and photoaging processes.

  4. Ultraviolet irradiation increases the sensitivity of cultured human skin cells to cadmium probably through the inhibition of metallothionein gene expression.

    PubMed

    Yamada, Hirotomo; Murata, Mie; Suzuki, Kaoru; Koizumi, Shinji

    2004-11-01

    We previously developed an apparatus that can irradiate cultured cells with monochromatic ultraviolet (UV) rays to exactly assess the biological effects of UV components on mammalian cells. Using this device, we studied the effects of UV in and near the UVB region on the general as well as specific protein synthesis of the human skin-derived NB1RGB cells. We found that Cd-induced synthesis of metallothioneins (MTs), which are the proteins involved in the protection against heavy metals and oxidative stress, is inhibited by UV at 280 nm more extensively than total protein synthesis. Such an inhibition was observed when MTs were induced by different inducers such as Cd, Zn, and dexamethasone in three human cell lines, indicating that it is not an event specific to a certain inducer or a certain cell type. By contrast, UV at 300 or 320 nm showed only a marginal effect. UV at 280 nm was likely to block MT gene transcription because Cd-induced increase of MT mRNA was strongly inhibited by irradiation. Cd induction of 70-kDa heat shock protein mRNA was also inhibited by UV irradiation, suggesting that the expression of inducible genes are commonly sensitive to UV. Furthermore, we observed that the irradiation of UV at 280 nm renders NB1RGB cells extremely susceptible to Cd, probably due to the reduced MT synthesis. These observations strongly suggest that UV at 280 nm severely damages cellular inducible protective functions, warning us of a new risk of UV exposure.

  5. On the effect of X-ray irradiation on the deformation and fracture behavior of human cortical bone.

    PubMed

    Barth, Holly D; Launey, Maximilien E; Macdowell, Alastair A; Ager, Joel W; Ritchie, Robert O

    2010-06-01

    In situ mechanical testing coupled with imaging using high-energy synchrotron X-ray diffraction or tomography is gaining in popularity as a technique to investigate micrometer and even sub-micrometer deformation and fracture mechanisms in mineralized tissues, such as bone and teeth. However, the role of the irradiation in affecting the nature and properties of the tissue is not always taken into account. Accordingly, we examine here the effect of X-ray synchrotron-source irradiation on the mechanistic aspects of deformation and fracture in human cortical bone. Specifically, the strength, ductility and fracture resistance (both work-of-fracture and resistance-curve fracture toughness) of human femoral bone in the transverse (breaking) orientation were evaluated following exposures to 0.05, 70, 210 and 630 kGrays (kGy) irradiation. Our results show that the radiation typically used in tomography imaging can have a major and deleterious impact on the strength, post-yield behavior and fracture toughness of cortical bone, with the severity of the effect progressively increasing with higher doses of radiation. Plasticity was essentially suppressed after as little as 70 kGy of radiation; the fracture toughness was decreased by a factor of five after 210 kGy of radiation. Mechanistically, the irradiation was found to alter the salient toughening mechanisms, manifest by the progressive elimination of the bone's capacity for plastic deformation which restricts the intrinsic toughening from the formation "plastic zones" around crack-like defects. Deep-ultraviolet Raman spectroscopy indicated that this behavior could be related to degradation in the collagen integrity.

  6. Post-irradiation viability and cytotoxicity of natural killer cells isolated from human peripheral blood using different methods.

    PubMed

    Hietanen, Tenho; Pitkänen, Maunu; Kapanen, Mika; Kellokumpu-Lehtinen, Pirkko-Liisa

    2016-01-01

    Purpose We compared the pre- and post-irradiation viability and cytotoxicity of human peripheral natural killer cell (NK) populations obtained using different isolation methods. Material and methods Three methods were used to enrich total NK cells from buffy coats: (I) a Ficoll-Paque gradient, plastic adherence and a nylon wool column; (II) a discontinuous Percoll gradient; or (III) the Dynal NK cell isolation kit. Subsequently, CD16(+) and CD56(+) NK cell subsets were collected using (IV) flow cytometry or (V) magnetic-activated cell sorting (MACS) NK cell isolation kits. The yield, viability, purity and cytotoxicity of the NK cell populations were measured using trypan blue exclusion, flow cytometry using propidium iodide and (51)Cr release assays after enrichments as well as viability and cytotoxicity after a single radiation dose. Results The purity of the preparations, as measured by the CD16(+) and CD56(+) cell content, was equally good between methods I-III (p = 0.323), but the content of CD16(+) and CD56(+) cells using these methods was significantly lower than that using methods IV and V (p = 0.005). The viability of the cell population enriched via flow cytometry (85.5%) was significantly lower than that enriched via other methods (99.4-98.0%, p = 0.003). The cytotoxicity of NK cells enriched using methods I-III was significantly higher than that of NK cells enriched using methods IV and V (p = 0.000). In vitro the NK cells did not recover cytotoxic activity following irradiation. In addition, we detected considerable inter-individual variation in yield, cytotoxicity and radiation sensitivity between the NK cells collected from different human donors. Conclusions The selection of the appropriate NK cell enrichment method is very important for NK cell irradiation studies. According to our results, the Dynal and MACS NK isolation kits best retained the killing capacity and the viability of irradiated NK cells.

  7. On the effect of x-ray irradiation on the deformation and fracture behavior of human cortical bone

    SciTech Connect

    Barth, Holly D.; Launey, Maximilien E.; McDowell, Alastair A.; Ager III, Joel W.; Ritchie, Robert O.

    2010-01-10

    In situ mechanical testing coupled with imaging using high-energy synchrotron x-ray diffraction or tomography imaging is gaining in popularity as a technique to investigate micrometer and even sub-micrometer deformation and fracture mechanisms in mineralized tissues, such as bone and teeth. However, the role of the irradiation in affecting the nature and properties of the tissue is not always taken into account. Accordingly, we examine here the effect of x-ray synchrotron-source irradiation on the mechanistic aspects of deformation and fracture in human cortical bone. Specifically, the strength, ductility and fracture resistance (both work-of-fracture and resistance-curve fracture toughness) of human femoral bone in the transverse (breaking) orientation were evaluated following exposures to 0.05, 70, 210 and 630 kGy irradiation. Our results show that the radiation typically used in tomography imaging can have a major and deleterious impact on the strength, post-yield behavior and fracture toughness of cortical bone, with the severity of the effect progressively increasing with higher doses of radiation. Plasticity was essentially suppressed after as little as 70 kGy of radiation; the fracture toughness was decreased by a factor of five after 210 kGy of radiation. Mechanistically, the irradiation was found to alter the salient toughening mechanisms, manifest by the progressive elimination of the bone's capacity for plastic deformation which restricts the intrinsic toughening from the formation 'plastic zones' around crack-like defects. Deep-ultraviolet Raman spectroscopy indicated that this behavior could be related to degradation in the collagen integrity.

  8. Effects of Smads and BMPs induced by Ga-Al-As laser irradiation on calcification ability of human dental pulp cells.

    PubMed

    Matsui, Satoshi; Takeuchi, Hitomi; Tsujimoto, Yasuhisa; Matsushima, Kiyoshi

    2008-03-01

    We investigated the effects of Ga-Al-As laser irradiation on the mineralization ability of human dental pulp (HDP) cells and on Smads and bone morphogenetic protein (BMP) production as one mechanism for the transmission of laser photochemical energy to cells. HDP cells in vitro were irradiated once with a Ga-Al-As laser at 1.0 W for 500 s, and calcified nodule formation was assessed by Alizarin red S staining. The laser irradiation was greater in the laser-irradiated group than in the non-irradiated group. Both calcium production and alkaline phosphatase (ALP) activity were higher after laser irradiation. Expression of mRNAs for Smad1, Smad7, BMPs, ALP, and osteocalcin was greater after laser irradiation, whereas expression of Smad6 mRNA was inhibited. Production of BMP-2 and BMP-4 in conditioned medium was also higher after laser irradiation. These results suggest that Smads and BMPs play important roles in ALP activity and calcification upon laser irradiation of HDP cells.

  9. Costimulatory signal provided by a B-lymphoblastoid cell line and its Ia-negative variant.

    PubMed Central

    Reiser, H; Benacerraf, B

    1989-01-01

    We have analyzed the requirements of highly purified, resting murine CD4+ T lymphocytes for activation mediated by the lectin Con A and by monoclonal antibodies against the CD3 and Thy-1 molecules. Our results indicate that both the Ia-positive B-lymphoblastoid cell line M12 and its Ia-negative variant M12.C3 can provide the costimulatory activity necessary for these activation pathways. The costimulatory function is preserved upon fixation with paraformaldehyde, indicating that the costimulatory molecule(s) is (are) constitutively expressed on the cell surface. Our experiments also point to interesting differences between the M12 cell line and syngeneic Ia-positive antigen-presenting cells in generating a syngeneic mixed lymphocyte reaction. Finally, we show that the CD4+ T cell-M12.C3 cell interaction can be used to screen for interesting monoclonal antibodies that affect cell function. Images PMID:2532358

  10. Randomised controlled trial of lymphoblastoid interferon for chronic active hepatitis B.

    PubMed Central

    Anderson, M G; Harrison, T J; Alexander, G; Zuckerman, A J; Murray-Lyon, I M

    1987-01-01

    Thirty male patients (27 homosexual) with biopsy proven chronic active hepatitis B were randomised to receive lymphoblastoid interferon (Wellferon) or no treatment. All patients were HBeAg positive and had continuing viral replication. Patients receiving treatment were given a single daily intramuscular injection of interferon for 28 days at a starting dose of 2.5 MU/m2 increasing to a maximum of 7.5 MU/m2/day. Transient side effects of malaise and influenza like symptoms occurred in all patients and resolved rapidly after treatment. Hepatitis B viral replication was suppressed during interferon treatment in all patients but the effect was limited to the period of therapy. After one year there was no appreciable difference in viral markers between the two groups of patients and this treatment schedule appears less effective than the thrice weekly, three month regimes recently reported from other centres. PMID:3297940

  11. Olfactory sensations produced by high-energy photon irradiation of the olfactory receptor mucosa in humans

    SciTech Connect

    Sagar, S.M.; Thomas, R.J.; Loverock, L.T.; Spittle, M.F. )

    1991-04-01

    During irradiation of volumes that incorporate the olfactory system, a proportion of patients have complained of a pungent smell. A retrospective study was carried out to determine the prevalence of this side-effect. A questionnaire was sent to 40 patients whose treatment volumes included the olfactory region and also to a control group treated away from this region. The irradiated tumor volumes included the frontal lobe, whole brain, nasopharynx, pituitary fossa, and maxillary antrum. Of the 25 patients who replied, 60% experienced odorous symptoms during irradiation. They described the odor as unpleasant and consistent with ozone. Stimulation of olfactory receptors is considered to be caused by the radiochemical formation of ozone and free radicals in the mucus overlying the olfactory mucosa.

  12. Transplantation of human renal cell carcinoma into NMRI nu/nu mice. III. Effect of irradiation on tumor acceptance and tumor growth

    SciTech Connect

    Otto, U.; Huland, H.; Baisch, H.; Kloeppel, G.

    1985-07-01

    Irradiation of human renal cell carcinoma before radical tumor nephrectomy resulted in a significantly lower acceptance rate (1 of 7) in nude mice than for nonirradiated tumors (all of 13). The tumor tissue was transplanted into NMRI nu/nu mice immediately after nephrectomy. In this experimental system the authors demonstrated the reduced vitality of human tumor cells after irradiation. In a second series of experiments, 3 morphologically different human renal cell carcinomas were irradiated at various doses after establishment in nude mice. The irradiated tumor tissue was transplanted to the next passage. The morphology, proliferation rate and growth of these tumors were compared with those of nonirradiated controls. Radiation effect was dose dependent in the responding tumor types. The characteristics correlated with radiosensitivity were high proliferation rate (measured by flow cytometry), low cytologic grading and fast growth rate in the nude mice.

  13. EXTRACTS OF IRRADIATED MATURE HUMAN TOOTH CROWNS CONTAIN MMP-20 PROTEIN AND ACTIVITY

    PubMed Central

    MCGUIRE, J.D.; MOUSA, A.A.; ZHANG, BO J.; TODOKI, L.S.; HUFFMAN, N.T.; CHANDRABABU, K.B.; MORADIAN-OLDAK, J.; KEIGHTLEY, A.; WANG, Y.; WALKER, M.P.; GORSKI, J.P.

    2014-01-01

    Objectives We recently demonstrated a significant correlation between enamel delamination and tooth-level radiation dose in oral cancer patients. Since radiation can induce the synthesis and activation of matrix metalloproteinases, we hypothesized that irradiated teeth may contain active matrix metalloproteinases. Materials and Methods Extracted teeth from oral cancer patients treated with radiotherapy and from healthy subjects were compared. Extracted mature third molars from healthy subjects were irradiated in vitro and/or incubated for 0 to 6 months at 37°C. All teeth were then pulverized, extracted, and extracts subjected to proteomic and enzymatic analyses. Results Screening of irradiated crown extracts using mass spectrometry identified MMP-20 (enamelysin) which is expressed developmentally in dentin and enamel but believed to be removed prior to tooth eruption. MMP-20 was composed of catalytically active forms at Mr=43, 41, 24 and 22 kDa and was immunolocalized predominantly to the morphological dentin enamel junction. The proportion of different sized MMP-20 forms changed with incubation and irradiation. While the pattern was not altered directly by irradiation of healthy teeth with 70 G, subsequent incubation at 37°C for 3–6 months with or without prior irradiation caused the proportion of Mr=24–22 kDa MMP-20 bands to increase dramatically. Extracts of teeth from oral cancer patients who received >70 Gy radiation also contained relatively more 24 and 22 kDa MMP-20 than those of healthy age-related teeth. Conclusion MMP-20 is a radiation-resistant component of mature tooth crowns enriched in the dentin-enamel. We speculate that MMP-20 catalyzed degradation of organic matrix at this site could lead to enamel delamination associated with oral cancer radiotherapy. PMID:24607847

  14. A comparison of four methods for determining viability in human dermal fibroblasts irradiated with blue light.

    PubMed

    Masson-Meyers, Daniela S; Bumah, Violet V; Enwemeka, Chukuka S

    2016-01-01

    Several tests are available for assessing the viability of cells; however, there is a dearth of studies comparing the results obtained with each test. We compared the capability of four viability assays (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT), neutral red, trypan blue and live/dead fluorescence), to detect potential toxicity in fibroblasts irradiated with 470nm blue light. Cells were irradiated at 3, 55, 110 and 220J/cm(2), incubated for 24h and viability assessed using each test. MTT assay showed significant decreases in viability when cells were irradiated with 110 and 220J/cm(2) energy fluence (dose) (89% and 57% viable cells, respectively; p<0.0001, compared to control); likewise the trypan blue assay showed 42% and 46% viable cells (p<0.0001). Neutral red assay revealed significant decrease in viability when cells were irradiated with 220J/cm(2) (84% viable cells; p=0.0008, compared to control). The live/dead fluorescence assay was less sensitive, evincing 91% and 95% viable cells after irradiation with 110 and 220J/cm(2) respectively. (1) The four assays differed in their levels of sensitivity to cell viability. (2) The adverse effect of increasing doses seems to manifest as alteration of mitochondrial metabolism, followed by lysosomal dysfunction, membrane disruption and finally loss of cell membrane integrity. (3) Overall, irradiation with 3J/cm(2) or 55J/cm(2) did not adversely affect cell viability. Thus, doses below 110J/cm(2) appear safe. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Expression profiles are different in carbon ion-irradiated normal human fibroblasts and their bystander cells.

    PubMed

    Iwakawa, Mayumi; Hamada, Nobuyuki; Imadome, Kaori; Funayama, Tomoo; Sakashita, Testuya; Kobayashi, Yasuhiko; Imai, Takashi

    2008-07-03

    Evidence has accumulated that ionizing radiation induces biological effects in non-irradiated bystander cells having received signals from directly irradiated cells; however, energetic heavy ion-induced bystander response is incompletely characterized. Here we performed microarray analysis of irradiated and bystander fibroblasts in confluent cultures. To see the effects in bystander cells, each of 1, 5 and 25 sites was targeted with 10 particles of carbon ions (18.3 MeV/u, 103 keV/microm) using microbeams, where particles traversed 0.00026, 0.0013 and 0.0066% of cells, respectively. diated cells, cultures were exposed to 10% survival dose (D), 0.1D and 0.01D of corresponding broadbeams (108 keV/microm). Irrespective of the target numbers (1, 5 or 25 sites) and the time (2 or 6h postirradiation), similar expression changes were observed in bystander cells. Among 874 probes that showed more than 1.5-fold changes in bystander cells, 25% were upregulated and the remainder downregulated. These included genes related to cell communication (PIK3C2A, GNA13, FN1, ANXA1 and IL1RAP), stress response (RAD23B, ATF4 and EIF2AK4) and cell cycle (MYCN, RBBP4 and NEUROG1). Pathway analysis revealed serial bystander activation of G protein/PI-3 kinase pathways. Instead, genes related to cell cycle or death (CDKN1A, GADD45A, NOTCH1 and BCL2L1), and cell communication (IL1B, TCF7 and ID1) were upregulated in irradiated cells, but not in bystander cells. Our results indicate different expression profiles in irradiated and bystander cells, and imply that intercellular signaling between irradiated and bystander cells activate intracellular signaling, leading to the transcriptional stress response in bystander cells.

  16. Comparison of human lung cancer cell radiosensitivity after irradiations with therapeutic protons and carbon ions.

    PubMed

    Keta, Otilija D; Todorović, Danijela V; Bulat, Tanja M; Cirrone, Pablo Ga; Romano, Francesco; Cuttone, Giacomo; Petrović, Ivan M; Ristić Fira, Aleksandra M

    2017-05-01

    The aim of this study was to investigate effects of irradiations with the therapeutic proton and carbon ion beams in two non-small cell lung cancers, CRL5876 adenocarcinoma and HTB177 large cell lung carcinoma. The DNA damage response dynamics, cell cycle regulation, and cell death pathway activation were followed. Viability of both cell lines was lower after carbon ions compared to the therapeutic proton irradiations. HTB177 cells showed higher recovery than CRL5876 cells seven days following the treatments, but the survival rates of both cell lines were lower after exposure to carbon ions with respect to therapeutic protons. When analyzing cell cycle distribution of both CRL5876 and HTB177 cells, it was noticed that therapeutic protons predominantly induced G1 arrest, while the cells after carbon ions were arrested in G2/M phase. The results illustrated that differences in the levels of phosphorylated H2AX, a double-strand break marker, exist after therapeutic proton and carbon ion irradiations. We also observed dose- and time-dependent increase in the p53 and p21 levels after applied irradiations. Carbon ions caused larger increase in the quantity of p53 and p21 compared to therapeutic protons. These results suggested that various repair mechanisms were induced in the treated cells. Considering the fact that we have not observed any distinct change in the Bax/Bcl-2 ratio following irradiations, it seemed that different types of cell death were involved in the response to the two types of irradiations that were applied.

  17. DNA fragmentation pattern in human fibroblasts after irradiation with iron ions

    NASA Astrophysics Data System (ADS)

    Campa, Alessandro

    In this work we studied the fragmentation pattern produced by the double stand breaks (DSB) induced in AG1522 primary human fibroblasts by two different iron beams, one of energy 414 MeV/u, and the other of energy 115 MeV/u (with dose-average LET in water equal to 202 keV/µm and 442 keV/µm, respectively). Irradiation with several doses up to 200 Gy was performed at the HIMAC facility of the National Institute of Radiological Sciences, Chiba, Japan. Experimental data, first obtained for fragments belonging to the size ranges 23-1000 kbp and 1000-5700 kbp (Belli et al., 2006), have successively been obtained also for fragments belonging to the size ranges 1-9 kbp and 9-23 kbp; the experimental analysis was performed with pulsed and constant field electrophoresis. The RBE for DSB production was evaluated in two different fragment size ranges (i.e., 23-5700 kbp and 1-5700 kbp), and it was found larger for the wider size range, especially for the beam with the higher LET. The experimental results have been compared to those computed on the basis of the Monte Carlo PARTRAC simulation code, following the line of research started in Campa et al. (2005), and exploiting the recent update of the PARTRAC code to ions heavier than helium (Friedland et al., 2006). Because the agreement has been found satisfactory for both radiation qualities, the spectra outside the experimentally observable fragment size range were also computed in order to evaluate the overall fragmentation pattern. The marked increases of the RBEs for DSB production, obtained when also the very small fragments (< 1 kbp) are included, makes them closer to the RBE values observed for the late cellular effects. This finding is a further indication for the biological significance of the spatial correlation of DSB at short distances. This work was partially supported by ASI (Italian Space Agency, "Mo-Ma/COUNT" project). References M. Belli, A. Campa, V. Dini, G. Esposito, Y. Furusawa, G. Simone, E. Sorrentino

  18. Phorbol diesters and transferrin modulate lymphoblastoid cell transferrin receptor expression by two different mechanisms

    SciTech Connect

    Alcantara, O.; Phillips, J.L.; Boldt, D.H.

    1986-12-01

    Expression of transferrin receptors (TfR) by activated lymphocytes is necessary for lymphocyte DNA synthesis and proliferation. Regulation of TfR expression, therefore, is a mechanism by which the lymphocyte's proliferative potential may be directed and controlled. The authors studied mechanisms by which lymphoblastoid cells modulate TfR expression during treatment with phorbol diesters or iron transferrin (FeTf), agents which cause downregulation of cell surface TfR. Phorbol diester-induced TfR downregulation occurred rapidly, being detectable at 2 min and reaching maximal decreases of 50% by 15 min. It was inhibited by cold but not by agents that destabilize cytoskeletal elements. Furthermore, this downregulation was reversed rapidly by washing or by treatment with the membrane interactive agent, chlorpromazine. In contrast, FeTf-induced TfR downregulation occurred slowly. Decreased expression of TfR was detectable only after 15 min and maximal downregulation was achieved after 60 min. Although FeTf-induced downregulation also was inhibited by cold, it was inhibited in addition by a group of microtubule destabilizing agents (colchicine, vinblastine, podophyllotoxin) or cytochalasin B, a microfilament inhibitor. Furthermore, FeTf-induced downregulation was not reversed readily by washing or by treatment with chlorpromazine. Phorbol diesters cause TfR downregulation by a cytoskeleton-independent mechanism. These data indicate that TfR expression is regulated by two independent mechanisms in lymphoblastoid cells, and they provide the possibility that downregulation of TfR by different mechanisms may result in different effects in these cells.

  19. Human fibroblast strain with normal survival but abnormal postreplication repair after ultraviolet light irradiation

    SciTech Connect

    Doniger, J.; Barrett, S.F.; Robbins, J.H.

    1980-08-01

    Postreplication repair has been studied in ultraviolet light (UV-irradiated) fibroblast strains derived from eight apparently normal control donors and seven xeroderma pigmentosum patients. One control donor strain had an intermediate defect in postreplication repair similar to that in excision-deficient xeroderma pigmentosum fibroblasts. However, unlike the xeroderma pigmentosum strains, this control donor strain had normal UV-induced unscheduled DNA synthesis and normal survival after irradiation with UV. This unique fibroblast strain should be useful in studies designed to elucidate the possible role of postreplication repair in UV-induced carcinogenesis and mutagenesis.

  20. Reactive oxygen species formation and bystander effects in gradient irradiation on human breast cancer cells

    PubMed Central

    Rong, Yi; Lee, Shin Hee; Wu, Shiyong; Zuo, Li

    2016-01-01

    Ionizing radiation (IR) in cancer radiotherapy can induce damage to neighboring cells via non-targeted effects by irradiated cells. These so-called bystander effects remain an area of interest as it may provide enhanced efficacy in killing carcinomas with minimal radiation. It is well known that reactive oxygen species (ROS) are ubiquitous among most biological activities. However, the role of ROS in bystander effects has not been thoroughly elucidated. We hypothesized that gradient irradiation (GI) has enhanced therapeutic effects via the ROS-mediated bystander pathways as compared to uniform irradiation (UI). We evaluated ROS generation, viability, and apoptosis in breast cancer cells (MCF-7) exposed to UI (5 Gy) or GI (8–2 Gy) in radiation fields at 2, 24 and 48 h after IR. We found that extracellular ROS release induced by GI was higher than that by UI at both 24 h (p < 0.001) and 48 h (p < 0.001). More apoptosis and less viability were observed in GI when compared to UI at either 24 h or 48 h after irradiation. The mean effective doses (ED) of GI were ~130% (24 h) and ~48% (48 h) higher than that of UI, respectively. Our results suggest that GI is superior to UI regarding redox mechanisms, ED, and toxic dosage to surrounding tissues. PMID:27223435

  1. Effects of proton beam irradiation on mitochondrial biogenesis in a human colorectal adenocarcinoma cell line.

    PubMed

    Ha, Byung Geun; Jung, Sung Suk; Shon, Yun Hee

    2017-09-01

    Proton beam therapy has recently been used to improve local control of tumor growth and reduce side-effects by decreasing the global dose to normal tissue. However, the regulatory mechanisms underlying the physiological role of proton beam radiation are not well understood, and many studies are still being conducted regarding these mechanisms. To determine the effects of proton beams on mitochondrial biogenesis, we investigated: mitochondrial DNA (mtDNA) mass; the gene expression of mitochondrial transcription factors, functional regulators, and dynamic-related regulators; and the phosphorylation of the signaling molecules that participate in mitochondrial biogenesis. Both the mtDNA/nuclear DNA (nDNA) ratio and the mitochondria staining assays showed that proton beam irradiation increases mitochondrial biogenesis in 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced aggressive HT-29 cells. Simultaneously, proton beam irradiation increases the gene expression of the mitochondrial transcription factors PGC-1α, NRF1, ERRα, and mtTFA, the dynamic regulators DRP1, OPA1, TIMM44, and TOM40, and the functional regulators CytC, ATP5B and CPT1-α. Furthermore, proton beam irradiation increases the phosphorylation of AMPK, an important molecule involved in mitochondrial biogenesis that is an energy sensor and is regulated by the AMP/ATP ratio. Based on these findings, we suggest that proton beam irradiation inhibits metastatic potential by increasing mitochondrial biogenesis and function in TPA-induced aggressive HT-29 cells.

  2. Irradiated homologous tarsal plate banking: a new alternative in eyelid reconstruction. Part II. Human data

    SciTech Connect

    Jordan, D.R.; Tse, D.T.; Anderson, R.L.; Hansen, S.O. )

    1990-01-01

    Reconstruction of full thickness eyelid defects requires the correction of both posterior lamella (tarsus, conjunctiva) and anterior lamella (skin, muscle). Irradiated homologous tarsal plate provides a structured framework for the lid reconstruction, and is incorporated nicely into the normal lid anatomy.

  3. DNA synthesis on UV irradiated model templates using human DNA polymerases alpha and beta: primer slippage to account for evident transdimer continuity in product.

    PubMed

    Philippe, M; Wang, T S; Hanawalt, P C; Korn, D

    1982-01-01

    We have studied the comparative behavior of human DNA polymerases alpha and beta on a polynucleotide template of dT100 with dA15 covalently attached at the 3' end to serve as primer when defined numbers of pyrimidine dimers are introduced by UV (254 nm) irradiation. We have obtained the surprising result that with both alpha and beta polymerases the incorporation of labelled dATP is enhanced when the template has been irradiated (maximum value at 1000 J/m2 UV incident dose). In the presence of Mn2+, DNA polymerase beta produces a product size corresponding to full copying of the template whether irradiated or not. In marked contrast DNA polymerase alpha produces only short products on unirradiated strands but full copying of irradiated templates. Evidently both polymerases utilize a much larger fraction of the template pool following UV irradiation.

  4. Inhibition of Autophagy Enhances Curcumin United light irradiation-induced Oxidative Stress and Tumor Growth Suppression in Human Melanoma Cells

    PubMed Central

    Niu, Tianhui; Tian, Yan; Mei, Zhusong; Guo, Guangjin

    2016-01-01

    Malignant melanoma is the most aggressive form of skin carcinoma, which possesses fast propagating and highly invasive characteristics. Curcumin is a natural phenol compound that has various biological activities, such as anti-proliferative and apoptosis-accelerating impacts on tumor cells. Unfortunately, the therapeutical activities of Cur are severely hindered due to its extremely low bioavailability. In this study, a cooperative therapy of low concentration Cur combined with red united blue light irradiation was performed to inspect the synergistic effects on the apoptosis, proliferation and autophagy in human melanoma A375 cell. The results showed that red united blue light irradiation efficaciously synergized with Cur to trigger oxidative stress-mediated cell death, induce apoptosis and inhibit cell proliferation. Meanwhile, Western blotting revealed that combined disposure induced the formation of autophagosomes. Conversely, inhibition of the autophagy enhanced apoptosis, obstructed cell cycle arrest and induced reversible proliferation arrest to senescence. These findings suggest that Cur combined with red united blue light irradiation could generate photochemo-preventive effects via enhancing apoptosis and triggering autophagy, and pharmacological inhibition of autophagy convert reversible arrested cells to senescence, therefore reducing the possibility that damaged cells might escape programmed death. PMID:27502897

  5. Procollagen-III in serum, plasminogen activation and fibronectin in bronchoalveolar lavage fluid during and following irradiation of human lung

    SciTech Connect

    Maasilta, P.; Salonen, E.M.; Vaheri, A.; Kivisaari, L.; Holsti, L.R.; Mattson, K. )

    1991-05-01

    In the search for predictors of late radiation-induced lung injury we studied procollagen type III peptide concentration (P-III-P) in serum as well as fibronectin and plasminogen activation in bronchoalveolar lavage (BAL) fluid during and following irradiation of human lung. The patients received either high-dose hemithorax irradiation for pleural mesothelioma (11 patients) or high-dose irradiation with individually shaped fields for non-small cell lung cancer (12 patients). The severity of radiation fibrosis was assessed clinically from CT scans 6 months and 12 months after treatment. Four scores were used: severe, moderate, mild, or normal. Radiological lung injury varied from 'severe' (9 patients) to near absence of injury-'normal' (6 patients). Serum levels of P-III-P, when measured weekly during the 5-week period of radiotherapy or at several time-points after treatment, did not show consistent changes, nor did the levels correlate with the score for radiation fibrosis as assessed by CT scanning. Changes in fibronectin levels or in markers of plasminogen activation in BAL fluid did not correlate with the development of late lung injury. The levels of BAL fluid plasmin and plasminogen activator as assessed zymographically, but not the free net enzyme values, showed a tendency to be elevated in patients with severe radiation-induced lung injury, suggesting a possible role for inhibitors of the plasminogen activation cascade in the process of radiation-induced lung injury.

  6. Matrix metalloproteinase-1 inhibitory activities of Morinda citrifolia seed extract and its constituents in UVA-irradiated human dermal fibroblasts.

    PubMed

    Masuda, Megumi; Murata, Kazuya; Naruto, Shunsuke; Uwaya, Akemi; Isami, Fumiyuki; Matsuda, Hideaki

    2012-01-01

    The objective of this study was to examine whether a 50% ethanolic extract (MCS-ext) of the seeds of Morinda citrifolia (noni) and its constituents have matrix metalloproteinase-1 (MMP-1) inhibitory activity in UVA-irradiated normal human dermal fibroblasts (NHDFs). The MCS-ext (10 μg/mL) inhibited MMP-1 secretion from UVA-irradiated NHDFs, without cytotoxic effects, at 48 h after UV exposure. The ethyl acetate-soluble fraction of MCS-ext was the most potent inhibitor of MMP-1 secretion. Among the constituents of the fraction, a lignan, 3,3'-bisdemethylpinoresinol (1), inhibited the MMP-1 secretion at a concentration of 0.3 μM without cytotoxic effects. Furthermore, 1 (0.3 μM) reduced the level of intracellular MMP-1 expression. Other constituents, namely americanin A (2), quercetin (3) and ursolic acid (4), were inactive. To elucidate inhibition mechanisms of MMP-1 expression and secretion, the effect of 1 on mitogen-activated protein kinases (MAPKs) phosphorylation was examined. Western blot analysis revealed that 1 (0.3 μM) reduced the phosphorylations of p38 and c-Jun-N-terminal kinase (JNK). These results suggested that 1 suppresses intracellular MMP-1 expression, and consequent secretion from UVA-irradiated NHDFs, by down-regulation of MAPKs phosphorylation.

  7. Glycyrrhizic acid (GA), a triterpenoid saponin glycoside alleviates ultraviolet-B irradiation-induced photoaging in human dermal fibroblasts.

    PubMed

    Afnan, Quadri; Adil, Mushtaq Dar; Nissar-Ul, Ashraf; Rafiq, Ahmad Rather; Amir, Hussian Faridi; Kaiser, Peerzada; Gupta, Vijay Kumar; Vishwakarma, Ram; Tasduq, Sheikh Abdullah

    2012-05-15

    Glycyrrhizic acid (GA), a triterpenoid saponin glycoside from the roots and rhizomes of licorice is used in traditional and modern medicine for the treatment of numerous medical conditions including skin diseases and beauty care product. In the present study, we investigated the effect of GA against ultraviolet B (UVB) irradiation-induced photoaging in human dermal fibroblasts (HDFs) and its possible mechanism of action. HDFs were subjected to photoaging by sub-toxic dose of UVB (10 mj/cm(2)) irradiation. Cell viability, matrix metalloproteinase 1 (MMP1), pro-collagen 1, cellular and nuclear morphology, cell cycle, intracellular reactive oxygen species (ROS), caspase 3 and hyaluronidase inhibition assays were performed. Western blotting was used to evaluate the expression of NF-kappa B (NF-κB) and cytochrome-C proteins. GA treatment significantly inhibited photoaging. It achieved this by reducing ROS, NF-κB, cytochrome c, caspase 3 levels and inhibiting hyaluronidase enzyme. The main mechanism seems to be, most likely by blocking MMP1 activation by modulating NF-κB signaling. These findings may be useful for development of natural and safe photoprotective agents against UVB irradiation. Copyright © 2012 Elsevier GmbH. All rights reserved.

  8. Photoprotective Potential of Anthocyanins Isolated from Acanthopanax divaricatus Var. albeofructus Fruits against UV Irradiation in Human Dermal Fibroblast Cells.

    PubMed

    Lyu, Su-Yun; Park, Won-Bong

    2012-03-01

    Ultraviolet (UV) A penetrates deeply into the skin and induces the generation of reactive oxygen species (ROS) causing damage to fibroblasts, which leads to aging of the skin. However, the body has developed an antioxidant defence system against the harmful effects of ROS. Enzymes such as superoxide dismutase (SOD) and catalase (CAT) play critical roles on the removal of excess ROS in living organisms. In this study, the antioxidant activities of anthocyanins (cyanidin 3-galactoside and cyanidin 3-lathyroside) from Acanthopanax divaricatus var. albeofructus (ADA) fruits were investigated by xylenol orange, thiobarbituric acid reactive substances (TBARS), and antioxidant enzyme assay. As a result, generation of H2O2 and lipid peroxide induced by UVA-irradiation in human dermal fibroblast (HDF-N) cells was reduced by treatment of anthocyanins. Also, augmented enzyme (SOD and CAT) activities were observed in UVA-irradiated cells when treated with anthocyanin. In conclusion, the results obtained show that anthocyanins from ADA fruits are potential candidates for the protection of fibroblast against the damaging effects of UVA irradiation. Furthermore, anthocyanin may be a good candidate for antioxidant agent development.

  9. The irradiation action on human dental tissue by X-rays and electrons--a nanoindenter study.

    PubMed

    Fränzel, Wolfgang; Gerlach, Reinhard

    2009-01-01

    It is known that ionizing radiation is used in medicine for Roentgen diagnostics and for radiation therapy. The radiation interacts with matter, in particular with biological one, essentially by scattering, photoelectric effect, Compton effect and pair production. To what extent the biological material is changed thereby, depends on the type and the amount of radiation energy, on the dose and on the tissue constitution. In modern radiation therapy two different kinds of radiation are used: high energy X-rays and electron radiation. In the case of head-neck tumors the general practice is an irradiation with high energy X-rays with absorbed dose to water up to 70 Gy. Teeth destruction has been identified as a side effect during irradiation. In addition, damage to the salivary glands is often observed which leads to a decrease or even the complete loss of the salivary secretion (xerostomia). This study shows how the different energy and radiation types damage the tooth tissue. The effects of both, high X-ray energy and high energy electrons, on the mechanical properties hardness and elasticity of the human dental tissue are measured by the nanoindentation technique. We compare these results with the effect of the irradiation of low X-ray energy on the dental tissue.

  10. Spectral responses of the human circadian system depend on the irradiance and duration of exposure to light

    PubMed Central

    Gooley, Joshua J; Rajaratnam, Shantha M; Brainard, George C; Kronauer, Richard E; Czeisler, Charles A; Lockley, Steven W

    2013-01-01

    In humans, circadian responses to light are thought to be mediated primarily by melanopsin-containing retinal ganglion cells, not rods or cones. Melanopsin cells are intrinsically blue-light sensitive, but also receive input from visual photoreceptors. We therefore tested in humans whether cone photoreceptors contribute to the regulation of circadian and neuroendocrine light responses. Dose-response curves for melatonin suppression and circadian phase resetting were constructed in subjects exposed to blue (460 nm) or green (555 nm) light near the onset of nocturnal melatonin secretion. At the beginning of the intervention, 555 nm light was just as effective as 460 nm light at suppressing melatonin, suggesting a significant contribution from the three-cone visual system (lambdamax 555 nm). During light exposure, however, the spectral sensitivity to 555 nm light decayed exponentially relative to 460 nm light. For phase-resetting responses, the effects of exposure to low irradiance 555 nm light were too large relative to 460 nm light to be explained solely by the activation of melanopsin. Our findings suggest that cone photoreceptors contribute substantially to non-visual responses at the beginning of a light exposure and at low irradiances, whereas melanopsin appears to be the primary circadian photopigment in response to long-duration light exposure and at high irradiances. These results are consistent with a non-redundant role for visual photoreceptors and melanopsin in mediating human non-visual photoreception and suggest that light therapy for circadian rhythm sleep disorders and other indications might be optimized by stimulating both the melanopsin- and cone-driven photoreceptor systems. PMID:20463367

  11. Evaluation of Potential Ionizing Irradiation Protectors and Mitigators Using Clonogenic Survival of Human Umbilical Cord Blood Hematopoietic Progenitor Cells

    PubMed Central

    Goff, Julie P.; Shields, Donna S.; Wang, Hong; Skoda, Erin M.; Sprachman, Melissa M.; Wipf, Peter; Garapati, Venkata Krishna; Atkinson, Jeffrey; London, Barry; Lazo, John S.; Kagan, Valerian; Epperly, Michael W.; Greenberger, Joel S.

    2013-01-01

    We evaluated the use of colony formation (CFU-GM, BFU-E, and CFU-GEMM) by human umbilical cord blood (CB) hematopoietic progenitor cells for testing novel small molecule ionizing irradiation protectors and mitigators. Each of 11 compounds was added before (protection) or after (mitigation) ionizing irradiation including: GS-nitroxides (JP4-039 and XJB-5-131), the bifunctional sulfoxide MMS-350, the phosphoinositol-3-kinase inhibitor (LY294002), TPP-imidazole fatty acid, (TPP-IOA), the nitric oxide synthase inhibitor (MCF-201-89), the p53/mdm2/mdm4 inhibitor (BEB55), methoxamine, isoproterenol, propanolol, and the ATP sensitive potassium channel blocker (glyburide). The drugs XJB-5-131, JP4-039, and MMS-350 were radiation protectors for CFU-GM. JP4-039 was also a radiation protector for CFU-GEMM. The drugs, XJB-5-131, JP4-039, and MMS-350 were radiation mitigators for BFU-E, MMS-350 and JP4-039 were mitigators for CFU-GM, and MMS350 was a mitigator for CFU-GEMM. In contrast, other drugs that were effective in murine assays: TTP-IOA, LY294002, MCF201-89, BEB55, propranolol, isoproterenol, methoxamine, and glyburide showed no significant protection or mitigation in human CB assays. These data support testing of new candidate clinical radiation protectors and mitigators using human CB clonogenic assays early in the drug discovery process, reducing the need for animal experiments. PMID:23933481

  12. Efficacy of low-power laser irradiation in the prevention of D-galactose-induced senescence in human dermal fibroblasts

    NASA Astrophysics Data System (ADS)

    Meng, Chengbo; Wu, Shengnan; Xing, Da

    2011-03-01

    Low-power laser (He-Ne) irradiation (LPLI) has been found to modulate various biological effects, especially those involved in promoting cell proliferation and metabolic regulation. However, the underlying mechanisms that LPLI prevents human cell senescence remain undefined. Herein, we devised a model enabling cell senescence using D-galactose for two days then treat with or without LPLI(< 15J/cm2), and investigated whether LPLI delays cell senescent in human dermal fibroblasts cells (HDF-a). First in this study, using SA-β-gal staining, compared with control cell we detected a lower frequency of SA-β-gal staining under the treatment of LPLI. Moreover, we found the growth rates of cell with LPLI was higher using CCK-8 analysis. Additionally, we also found LPLI induced HDF-a entered the irreversible G1 arrest measured by flow cytometry system. Therefore, LPLI may promote cell proliferation by stimulating cell-cycle progression and delay human cell senescence. Taken together, Low-power laser irradiation delay HDF-a cells senescence provides new information for the mechanisms of biological effects of LPLI.

  13. Recommendations to mitigate against human health risks incurred due to energetic particle irradiation beyond low earth orbit/BLEO

    NASA Astrophysics Data System (ADS)

    McKenna-Lawlor, Susan; Bhardwaj, Anil; Ferrari, Franco; Kuznetsov, Nikolay; Lal, Ajay K.; Li, Yinghui; Nagamatsu, Aiko; Nymmik, Rikho; Panasyuk, Michael; Petrov, Vladislav; Reitz, Günther; Pinsky, Lawrence; Shukor, Muszaphar (Sheikh); Singhvi, Ashok K.; Straube, Ulrich; Tomi, Leena; Lawrence, Townsend

    2015-04-01

    An account is provided of the main sources of energetic particle radiation in interplanetary space (Galactic Cosmic Radiation and Solar Energetic Particles) and career dose limits presently utilized by NASA to mitigate against the cancer and non-cancer effects potentially incurred by astronauts due to irradiation by these components are presented. Certain gaps in knowledge that presently militate against mounting viable human exploration in deep space due to the inherent health risks are identified and recommendations made as to how these gaps might be closed within a framework of global international cooperation.

  14. Radiosensitizing effect of misonidazole in acute and fractionated irradiation of a human osteosarcoma xenograft. [/sup 60/Co

    SciTech Connect

    Rofstad, E.K.; Brustad, T.

    1980-09-01

    The radiosensitizing effect of misonidazole (Ro-07-0582) in acute and fractionated irradiation of a human osteosarcoma grown in the athymic mutant nude mouse was studied. Tumor regrowth delay was used as a measure of response. The enhancement ratio of misonidazole was found to be 1.45 for an actue dose of 12.50 Gy and 1.25 for four fractions of 3.75 Gy, delivered over four consecutive days. It is concluded that the present osteosarcoma xenograft reoxygenated inadequately during the three day period which elapsed from the first to the fourth fraction of 3.75 Gy.

  15. Characterization of the effects of x-ray irradiation on the hierarchical structure and mechanical properties of human cortical bone

    SciTech Connect

    Barth, Holly; Zimmermann, Elizabeth; Schaible, Eric; Tang, Simon; Alliston, Tamara; Ritchie, Robert

    2011-08-19

    Bone comprises a complex structure of primarily collagen, hydroxyapatite and water, where each hierarchical structural level contributes to its strength, ductility and toughness. These properties, however, are degraded by irradiation, arising from medical therapy or bone-allograft sterilization. We provide here a mechanistic framework for how irradiation affects the nature and properties of human cortical bone over a range of characteristic (nano to macro) length-scales, following x-­ray exposures up to 630 kGy. Macroscopically, bone strength, ductility and fracture resistance are seen to be progressively degraded with increasing irradiation levels. At the micron-­scale, fracture properties, evaluated using in-situ scanning electron microscopy and synchrotron x-ray computed micro-tomography, provide mechanistic information on how cracks interact with the bone-matrix structure. At sub-micron scales, strength properties are evaluated with in-situ tensile tests in the synchrotron using small-/wide-angle x-ray scattering/diffraction, where strains are simultaneously measured in the macroscopic tissue, collagen fibrils and mineral. Compared to healthy bone, results show that the fibrillar strain is decreased by ~40% following 70 kGy exposures, consistent with significant stiffening and degradation of the collagen. We attribute the irradiation-­induced deterioration in mechanical properties to mechanisms at multiple length-scales, including changes in crack paths at micron-­scales, loss of plasticity from suppressed fibrillar sliding at sub-­micron scales, and the loss and damage of collagen at the nano-­scales, the latter being assessed using Raman and Fourier-Transform-Infrared spectroscopy and a fluorometric assay.

  16. Characterization of the effects of x-ray irradiation on the hierarchical structure and mechanical properties of human cortical bone.

    PubMed

    Barth, Holly D; Zimmermann, Elizabeth A; Schaible, Eric; Tang, Simon Y; Alliston, Tamara; Ritchie, Robert O

    2011-12-01

    Bone comprises a complex structure of primarily collagen, hydroxyapatite and water, where each hierarchical structural level contributes to its strength, ductility and toughness. These properties, however, are degraded by irradiation, arising from medical therapy or bone-allograft sterilization. We provide here a mechanistic framework for how irradiation affects the nature and properties of human cortical bone over a range of characteristic (nano to macro) length-scales, following x-ray exposures up to 630 kGy. Macroscopically, bone strength, ductility and fracture resistance are seen to be progressively degraded with increasing irradiation levels. At the micron-scale, fracture properties, evaluated using insitu scanning electron microscopy and synchrotron x-ray computed micro-tomography, provide mechanistic information on how cracks interact with the bone-matrix structure. At sub-micron scales, strength properties are evaluated with insitu tensile tests in the synchrotron using small-/wide-angle x-ray scattering/diffraction, where strains are simultaneously measured in the macroscopic tissue, collagen fibrils and mineral. Compared to healthy bone, results show that the fibrillar strain is decreased by ∼40% following 70 kGy exposures, consistent with significant stiffening and degradation of the collagen. We attribute the irradiation-induced deterioration in mechanical properties to mechanisms at multiple length-scales, including changes in crack paths at micron-scales, loss of plasticity from suppressed fibrillar sliding at sub-micron scales, and the loss and damage of collagen at the nano-scales, the latter being assessed using Raman and Fourier Transform Infrared spectroscopy and a fluorometric assay.

  17. Induction of expression of human immunodeficiency virus in a chronically infected promonocytic cell line by ultraviolet irradiation

    SciTech Connect

    Stanley, S.K.; Folks, T.M.; Fauci, A.S. )

    1989-08-01

    Infection with the human immunodeficiency virus (HIV) is often followed by a prolonged latent state, and mechanisms of maintaining latency or inducing expression from latency are active areas in AIDS research. It has been previously shown using a variety of viruses and cell systems that ultraviolet (UV) irradiation is capable of inducing the expression of latent viruses as well as augmenting the effects of acute viral infection. The ability of UV irradiation to affect HIV latency was investigated using a chronically HIV-infected, virus nonexpressing promonocytic cell line termed U1. After exposure to UV-C in doses ranging from 0.75 to 2.0 mJ/cm{sup 2}, U1 cells were induced to express virus as assessed by detection of elevated reverse transcriptase activity and p24 antigen levels in culture supernatants of treated cells compared with unstimulated controls. In addition, immunofluorescence on cytospin preparations of UV-irradiated cells revealed a time-dependent increase in viral antigen production after UV stimulation. A similar increase in RT levels was seen after exposure of U1 cells to UV-B, although somewhat higher doses of UV-B (mJ) were required compared with UV-C (mJ). Viral induction by UV irradiation was associated with a drop in viability and a static growth curve, suggesting that a certain level of cellular stress was most likely necessary to initiate viral expression. The potential role of UV-induced cell damage with activation of a cellular SOS repair response is a probable explanation of the enhanced viral production observed.

  18. Characterization of the effects of x-ray irradiation on the hierarchical structure and mechanical properties of human cortical bone

    PubMed Central

    Barth, Holly D.; Zimmermann, Elizabeth A.; Schaible, Eric; Tang, Simon Y.; Alliston, Tamara; Ritchie, Robert O.

    2012-01-01

    Bone comprises a complex structure of primarily collagen, hydroxyapatite and water, where each hierarchical structural level contributes to its strength, ductility and toughness. These properties, however, are degraded by irradiation, arising from medical therapy or bone-allograft sterilization. We provide here a mechanistic framework for how irradiation affects the nature and properties of human cortical bone over a range of characteristic (nano to macro) length-scales, following x-ray exposures up to 630 kGy. Macroscopically, bone strength, ductility and fracture resistance are seen to be progressively degraded with increasing irradiation levels. At the micron-scale, fracture properties, evaluated using insitu scanning electron microscopy and synchrotron x-ray computed micro-tomography, provide mechanistic information on how cracks interact with the bone-matrix structure. At sub-micron scales, strength properties are evaluated with insitu tensile tests in the synchrotron using small-/wide-angle x-ray scattering/diffraction, where strains are simultaneously measured in the macroscopic tissue, collagen fibrils and mineral. Compared to healthy bone, results show that the fibrillar strain is decreased by ~40% following 70 kGy exposures, consistent with significant stiffening and degradation of the collagen. We attribute the irradiation-induced deterioration in mechanical properties to mechanisms at multiple length-scales, including changes in crack paths at micron-scales, loss of plasticity from suppressed fibrillar sliding at sub-micron scales, and the loss and damage of collagen at the nano-scales, the latter being assessed using Raman and Fourier Transform Infrared spectroscopy and a fluorometric assay. PMID:21885114

  19. pH Dependant Binding and Irradiation of Protoporphyrin IX to Human Serum Albumin

    NASA Astrophysics Data System (ADS)

    Rozinek, Sarah; Brancaleon, Lorenzo

    2010-10-01

    Irradiation of the non-covalent complex, protoporphyrin IX (PPIX) bound to β-lactoglobulin (β-lg), causes a modest unfolding of the protein localized to Trp19. PPIX binds to β-lg at a site affected by the pH of the solution. At physiological pH, PPIX is known to bind HSA in hydrophobic binding sites located in subdomain IIA and IIIA. However, no evidence is presented for the binding behavior of PPIX to HSA in non-physological pH confirmations, nor on the effects of irradiation on the bound system at any pH. The combination of spectroscopic data and molecular simulations suggests that distinct PPIX-compatible binding sites become available at each confirmation of HSA at pH 3, 7.4, and 9.

  20. Generation of breast cancer stem cells by steroid hormones in irradiated human mammary cell lines.

    PubMed

    Vares, Guillaume; Cui, Xing; Wang, Bing; Nakajima, Tetsuo; Nenoi, Mitsuru

    2013-01-01

    Exposure to ionizing radiation was shown to result in an increased risk of breast cancer. There is strong evidence that steroid hormones influence radiosensitivity and breast cancer risk. Tumors may be initiated by a small subpopulation of cancer stem cells (CSCs). In order to assess whether the modulation of radiation-induced breast cancer risk by steroid hormones could involve CSCs, we measured by flow cytometry the proportion of CSCs in irradiated breast cancer cell lines after progesterone and estrogen treatment. Progesterone stimulated the expansion of the CSC compartment both in progesterone receptor (PR)-positive breast cancer cells and in PR-negative normal cells. In MCF10A normal epithelial PR-negative cells, progesterone-treatment and irradiation triggered cancer and stemness-associated microRNA regulations (such as the downregulation of miR-22 and miR-29c expression), which resulted in increased proportions of radiation-resistant tumor-initiating CSCs.

  1. Preoperative irradiation for the prevention of heterotopic ossification induces local inflammation in humans.

    PubMed

    Hoff, Paula; Rakow, Anastasia; Gaber, Timo; Hahne, Martin; Sentürk, Ufuk; Strehl, Cindy; Fangradt, Monique; Schmidt-Bleek, Katharina; Huscher, Dörte; Winkler, Tobias; Matziolis, Dörte; Matziolis, Georg; Badakhshi, Harun; Burmester, Gerd-Rüdiger; Duda, Georg N; Perka, Carsten; Buttgereit, Frank

    2013-07-01

    Radiation of the hip is an established method to prevent heterotopic ossification (HO) following total hip arthroplasty (THA) but the precise mechanism is unclear. As inflammatory processes are suggested to be involved in the pathogenesis of HO, we hypothesized that the preoperative irradiation impacts local immune components. Therefore, we quantified immune cell populations and cytokines in hematomas resulting from the transection of the femur in two groups of patients receiving THA: patients irradiated preoperatively (THA-X-hematoma: THA-X-H group) in the hip region (7 Gy) in order to prevent HO and patients who were not irradiated (THA-H group) but were postoperatively treated with non-steroidal anti-inflammatory drugs (NSAIDs). Radiation resulted in significantly increased frequencies of T cells, cytotoxic T cells, NKT cells and CD25+CD127- Treg cells, whereas the number of naive CD45RA-expressing cytotoxic T cells was reduced. These results indicate differential immune cell activation, corroborated by our findings of significantly higher concentrations of pro-inflammatory cytokines (e.g., IL-6, IFNγ) and chemokines (e.g., MCP-1, RANTES) in the THA-X-H group as compared to THA-H group. In contrast, the concentration of the angiogenic VEGF was significantly suppressed in the THA-X-H group. We conclude that preoperative irradiation results in significant changes in immune cell composition and cytokine secretion in THA-hematomas, establishing a specific - rather proinflammatory - milieu. This increase of inflammatory activity together with the observed suppression in VEGF secretion may contribute to the prevention of HO.

  2. Profiling of genes central to human mitochondrial energy metabolism following low intensity laser irradiation

    NASA Astrophysics Data System (ADS)

    Houreld, Nicolette N.; Masha, Roland; Abrahamse, Heidi

    2012-09-01

    Background: Wound healing involves three overlapping phases: inflammation, granulation and tissue remodelling. If this process is disrupted, delayed wound healing ensues, a common complication seen in diabetic patients. Low intensity laser irradiation (LILI) has been found to promote healing in such patients. However, the exact mechanisms of action are poorly understood. Purpose: This study aimed to profile the expression of key genes involved in mitochondrial respiration. Materials and Methods: Diabetic wounded fibroblast cells were exposed to a wavelength of 660 nm and a fluence of 5 J/cm2 and incubated for 30 min. Total RNA was isolated and 1 μg reverse transcribed into cDNA which was used for real-time polymerase chain reaction (PCR) array analysis. The array contained genes important for each of the mitochondrial complexes involved in the electron transport chain (ETC). Adenosine triphosphate (ATP) levels were also determined post-irradiation by ATP luminescence. Results: Genes involved in complex IV (cytochrome c oxidase), COX6B2 and COX6C, and PPA1 which is involved in complex V (ATP synthase) were significantly up-regulated. There was a significant increase in ATP levels in diabetic wounded cells post-irradiation. Discussion and Conclusion: LILI stimulates the ETC at a transcriptional level, resulting in an increase in ATP. This study helps understand the mechanisms of LILI in diabetic wound healing, and gives information on activation of genes in response to LILI.

  3. The effect of gamma irradiation on the osteoinductivity of demineralized human bone allograft.

    PubMed

    Arjmand, Babak; Aghayan, Hamid Reza; Larijani, Bagher; Sahebjam, Mehrnaz; Ghaderi, Firoozeh; Goodarzi, Parisa

    2014-01-01

    The gamma irradiation has been used for end sterilization of allograft bones and its effects with a 25 kGy dosage on the osteoinductive properties of demineralized bone allograft powder was studied. This work carried out using an experimental method in an animal model. In this study the demineralized bone allograft powder which had been sterilized and prepared with gamma irradiation in a 25 kGy dosage in 18 hours, was used as a study group and the demineralized bone allograft powder which had been prepared aseptically was used as the reference group. 30 mg of bone powder from each group were implanted into right and left paravertebral muscles of eighteen rats, separately. After four weeks, the implanted samples were harvested with a 0.5 cm border and then the osteoinductivity of implants in two groups were compared with histopathologic studies. In 94.4% of the reference samples a new bone formation was observed. In the study group, this difference was observed only in 27.7% of samples (P<0.002). It appears that using gamma irradiation may lead to a reduction in osteoinduction properties of demineralized bone allograft powder.

  4. Fluorescence-guided surgery in combination with UVC irradiation cures metastatic human pancreatic cancer in orthotopic mouse models.

    PubMed

    Hiroshima, Yukihiko; Maawy, Ali; Zhang, Yong; Sato, Sho; Murakami, Takashi; Yamamoto, Mako; Uehara, Fuminari; Miwa, Shinji; Yano, Shuya; Momiyama, Masashi; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Endo, Itaru; Hoffman, Robert M

    2014-01-01

    The aim of this study is to determine if ultraviolet light (UVC) irradiation in combination with fluorescence-guided surgery (FGS) can eradicate metastatic human pancreatic cancer in orthotopic nude-mouse models. Two weeks after orthotopic implantation of human MiaPaCa-2 pancreatic cancer cells, expressing green fluorescent protein (GFP), in nude mice, bright-light surgery (BLS) was performed on all tumor-bearing mice (n = 24). After BLS, mice were randomized into 3 treatment groups; BLS-only (n = 8) or FGS (n = 8) or FGS-UVC (n = 8). The residual tumors were resected using a hand-held portable imaging system under fluorescence navigation in mice treated with FGS and FGS-UVC. The surgical resection bed was irradiated with 2700 J/m2 UVC (254 nm) in the mice treated with FGS-UVC. The average residual tumor area after FGS (n = 16) was significantly smaller than after BLS only (n = 24) (0.135±0.137 mm2 and 3.338±2.929 mm2, respectively; p = 0.007). The BLS treated mice had significantly reduced survival compared to FGS- and FGS-UVC-treated mice for both relapse-free survival (RFS) (p<0.001 and p<0.001, respectively) and overall survival (OS) (p<0.001 and p<0.001, respectively). FGS-UVC-treated mice had increased RFS and OS compared to FGS-only treated mice (p = 0.008 and p = 0.025, respectively); with RFS lasting at least 150 days indicating the animals were cured. The results of the present study suggest that UVC irradiation in combination with FGS has clinical potential to increase survival.

  5. Fluorescence-Guided Surgery in Combination with UVC Irradiation Cures Metastatic Human Pancreatic Cancer in Orthotopic Mouse Models

    PubMed Central

    Hiroshima, Yukihiko; Maawy, Ali; Zhang, Yong; Sato, Sho; Murakami, Takashi; Yamamoto, Mako; Uehara, Fuminari; Miwa, Shinji; Yano, Shuya; Momiyama, Masashi; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Endo, Itaru; Hoffman, Robert M.

    2014-01-01

    The aim of this study is to determine if ultraviolet light (UVC) irradiation in combination with fluorescence-guided surgery (FGS) can eradicate metastatic human pancreatic cancer in orthotopic nude–mouse models. Two weeks after orthotopic implantation of human MiaPaCa-2 pancreatic cancer cells, expressing green fluorescent protein (GFP), in nude mice, bright-light surgery (BLS) was performed on all tumor-bearing mice (n = 24). After BLS, mice were randomized into 3 treatment groups; BLS-only (n = 8) or FGS (n = 8) or FGS-UVC (n = 8). The residual tumors were resected using a hand-held portable imaging system under fluorescence navigation in mice treated with FGS and FGS-UVC. The surgical resection bed was irradiated with 2700 J/m2 UVC (254 nm) in the mice treated with FGS-UVC. The average residual tumor area after FGS (n = 16) was significantly smaller than after BLS only (n = 24) (0.135±0.137 mm2 and 3.338±2.929 mm2, respectively; p = 0.007). The BLS treated mice had significantly reduced survival compared to FGS- and FGS-UVC-treated mice for both relapse-free survival (RFS) (p<0.001 and p<0.001, respectively) and overall survival (OS) (p<0.001 and p<0.001, respectively). FGS-UVC-treated mice had increased RFS and OS compared to FGS-only treated mice (p = 0.008 and p = 0.025, respectively); with RFS lasting at least 150 days indicating the animals were cured. The results of the present study suggest that UVC irradiation in combination with FGS has clinical potential to increase survival. PMID:24924955

  6. [Comparative study of time-correlated temperature and back-scattered light intensity for human Hegu acupoint and non-acupoint tissue irradiated by near-infrared laser].

    PubMed

    Zhou, Fang; Wei, Hua-Jiang; Guo, Zhou-Yi; Li, Ang; Yang, Ning-Ning; Yang, Hong-Qin; Xie, Shu-Sen

    2012-09-01

    Characteristics and differences of temperature and back-scattered light intensity in different depths of 0.2, 0.4, 0.6, 0.8 and 1 mm for both human Hegu acupoint and non-acupoint tissue irradiated by 808 nm diode laser at the different power of 15, 25 and 35 mW were studied. The temperature and the back-scattered light intensity in different depths of 0.2, 0.4, 0.6, 0.8 and 1 mm for human Hegu acupoint and non-acupoint tissue were measured by using the infrared thermography and optical coherence tomography. The result shows few differences in the temperature and the back-scattered light intensity of human Hegu acupoint and non-acupoint tissue before irradiation. The temperature and back-scattered light intensity of Hegu acupoint and the non-acupoint after irradiation were significantly higher, and the temperature and back-scattered light intensity of Hegu acupoint significantly were higher than the non-acupoint areas. At 0-40 min after the irradiation, the temperature and back-scattered light intensity of Hegu acupoint and the non-acupoint area will fluctuate and gradually decrease with the passage of time. From the results above, it is clearly seen that Hegu acupoint is different from non-acupoint both in the back-scattered light intensity and temperature after irradiation, and Hegu acupoint is more sensitive to laser irradiation than non-acupoint tissue.

  7. Intra-bone marrow transplantation of human CD34(+) cells into NOD/LtSz-scid IL-2rgamma(null) mice permits multilineage engraftment without previous irradiation.

    PubMed

    Bueno, Clara; Montes, Rosa; de la Cueva, Teresa; Gutierrez-Aránda, Iván; Menendez, Pablo

    2010-01-01

    Non-irradiated immunodeficient recipients provide the best physiologic setting for revealing hematopoietic stem cell (HSC) functions after xenotransplantion. An approach that efficiently permits the detection of human hematopoietic repopulating cells in non-irradiated recipients is therefore highly desired. We compared side-by-side the ability to reconstitute hematopoiesis via intra-bone marrow transplantation (IBMT) in three commonly used mouse strains avoiding previous irradiation. Non-irradiated NOD/SCID and NOD/SCID (beta2m-/- mouse strains prevent engraftment even after IBMT. In contrast, combining the robustness of the NOD/SCID IL-2Rgamma-/- recipient with the sensitivity of IBMT facilitates the detection, without previous host irradiation, of human SCID-repopulating cells 10 weeks after transplantation. The level of chimerism averaged 14% and multilineage engraftment (lymphoid dominant) was observed consistently in all mice. Analysis of injected and non-injected bones, spleen and peripheral blood demonstrated that engrafting cells were capable of in vivo migration and expansion. Combining the robustness of the NOD/SCID IL-2Rgamma-/- mouse strain with the sensitivity of IBMT strongly facilitates long-term multilineage engraftment and migration for human CD34(+) cells without the need for previous irradiation.

  8. Effects of ultraviolet-visible irradiation in the presence of melanin isolated from human black or red hair upon Ehrlich ascites carcinoma cells

    SciTech Connect

    Menon, I.A.; Persad, S.; Ranadive, N.S.; Haberman, H.F.

    1983-07-01

    The present study is an attempt to investigate the possibility that ultraviolet irradiation in the presence of pheomelanin may be more harmful to cells than the irradiation in the presence of eumelanin. The effects of UV-visible irradiation upon Ehrlich ascites carcinoma cells in the presence of the melanin isolated from human black hair (eumelanin) or from red hair (pheomelanin) were investigated. Irradiation of these cells was found to produce cell lysis, as observed by leakage of 51Cr from labeled cells and intracellular lactic dehydrogenase from the cells and decrease in cell viability demonstrated by the trypan blue exclusion test. The three parameters were quantitatively parallel to one another under various experimental conditions, namely different periods of irradiation and irradiation in the presence of different concentrations of melanin. The above effects were more pronounced when the irradiation was carried out in the presence of melanin from red hair than in the presence of black-hair melanin. In the absence of either melanin, the irradiation did not produce any significant effect in cell viability or cell lysis. Irradiation of the cells in the presence of red-hair melanin also decreased the transplantability of these cells. These observations clearly show that irradiation of cells in the presence of pheomelanin could produce cytotoxic effects. The present experimental design may have application in the development of in vitro models for the study of UV radiation-induced cutaneous carcinogenesis. The reactions of pheomelanin may be related to the susceptibility of ''Celtic'' skin to UV radiation-induced skin damage and carcinogenesis.

  9. Leakage of potassium from red blood cells following gamma ray irradiation in the presence of dipyridamole, trolox, human plasma or mannitol.

    PubMed

    Hirayama, Junichi; Abe, Hideki; Azuma, Hiroshi; Ikeda, Hisami

    2005-07-01

    Transfusion-associated graft-versus-host disease (TA-GVHD) is a fatal complication of blood transfusion resulting from the contamination of blood products by leukocytes. In order to prevent this disease, gamma or X-ray irradiation of blood components,which can inactivate leukocytes, is currently used. However, the minimal doses needed to destroy lymphocytes promote the leakage of potassium from red blood cells (RBCs), which can induce other side effects, such as hyperpotassemia and cardiac arrest. The reactive oxygen species (ROS) generated by the irradiation of aqueous solutions may accelerate the leakage through oxidation of the RBC membrane. Here we studied the effect of dipyridamole, Trolox, human plasma or mannitol on the leakage of potassium from RBCs following irradiation. RBC preparations (hematocrit; 30%) containing antioxidants were irradiated at 30 Gy and stored at 4 degrees C for 7 d. The leakage of potassium from the RBCs caused by the irradiation was significantly suppressed by dipyridamole (more than 50 microM), Trolox (more than 5 mM) or human plasma (50%). Mannitol (80 mM) is used to inhibit hemolysis as a constituent of MAP solution, which is a solution used for the storage of RBC products in Japan. Here it was clarified that the leakage of potassium from not only irradiated but also non-irradiated RBCs was unexpectedly promoted by mannitol. The amount of mannitol in MAP solution may have to be reconsidered. The osmotic pressure of the RBC preparation increased in a manner dependent on the concentration of mannitol. The elevated osmotic pressure may promote the leakage. In conclusion, although antioxidants have the potential to suppress the leakage of potassium ascribed to the irradiation, the extent of the suppression (10-20%) by dipyridamole (DPM), Trolox or human plasma seems insufficient for the clinical use of these agents as an additive for MAP solution.

  10. Human amnion-derived mesenchymal stem cells protect against UVA irradiation-induced human dermal fibroblast senescence, in vitro.

    PubMed

    Zhang, Chunli; Yuchi, Haishen; Sun, Lu; Zhou, Xiaoli; Lin, Jinde

    2017-08-01

    The aim of the present study was to determine if human amnion‑derived mesenchymal stem cells (HAMSCs) exert a protective effect on ultraviolet A (UVA) irradiation-induced human dermal fibroblast (HDF) senescence. A senescence model was constructed as follows: HDFs (104‑106 cells/well) were cultured in a six‑well plate in vitro and then exposed to UVA irradiation at 9 J/cm2 for 30 min. Following the irradiation period, HDFs were co‑cultured with HAMSCs, which were seeded on transwells. A total of 72 h following the co‑culturing, senescence‑associated β‑galactosidase staining was performed and reactive oxygen species (ROS) content and mitochondrial membrane potential (Δψm) were detected in the HDFs via flow cytometric analysis. The results demonstrated that the percentage of HDFs, detected via staining with X‑gal, were markedly decreased when co‑cultured with human HAMSCs, compared with the group that were not co‑cultured. The ROS content was decreased and the mitochondrial membrane potential (Δψm) recovered in cells treated with UVA and HAMSCs, compared with that of cells treated with UVA alone. Reverse transcription‑quantitative polymerase chain reaction revealed the significant effects of HAMSCs on the HDF senescence marker genes p53 and matrix metalloproteinase‑1 mRNA expression. In addition to this, western blot analysis verified the effects of HAMSCs on UVA induced senescence, providing a foundation for novel regenerative therapeutic methods. Furthermore, the results suggested that activation of the extracellular‑signal regulated kinase 1/2 mitogen activated protein kinase signal transduction pathway, is essential for the HAMSC‑mediated UVA protective effects. The decrease in ROS content additionally indicated that HAMSCs may exhibit the potential to treat oxidative stress‑mediated UVA skin senescence in the future.

  11. Stabilization of Quercetin Paradoxically Reduces Its Proapoptotic Effect on UVB-Irradiated Human Keratinocytes

    PubMed Central

    Olson, Erik R.; Melton, Tania; Dong, Zigang; Bowden, G. Tim

    2008-01-01

    UVB light promotes survival of initiated keratinocytes, in part, by the direct activation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway. Novel chemopreventative agents targeting UVB-induced signaling pathways are needed to reduce the incidence of nonmelanoma skin cancer. Quercetin (Qu) is a dietary flavonoid and a known inhibitor of PI3K. We determined that Qu degrades rapidly when diluted in DMEM and incubated under normal cell culture conditions. Degradation was delayed by supplementing the medium with 1 mmol/L ascorbic acid (AA), and as expected, stabilization actually increased the effectiveness of Qu as a PI3K inhibitor because basal and UVB-induced Akt phosphorylation were reduced compared with Qu treatment in the absence of AA. Although AA stabilization increased Qu-induced apoptosis in mock-irradiated HaCaT cells, consistent with it acting as a PI3K inhibitor (13.4% Annexin V–positive cells for AA-stabilized Qu versus 6.3% for Qu), AA stabilization of Qu actually reduced the ability of the compound to induce apoptosis of UVB-irradiated HaCaTs (29.7% of Qu-treated cells versus 15.5% of AA + Qu–treated cells). Similar trends were seen in the analysis of caspase-3 and poly(ADP-ribose) polymerase cleavage. Qu is known to oxidize to form reactive products, and we found that dihydroethidium is oxidized by Qu regardless of whether or not it was stabilized. Although redox cycling occurs even in the presence of AA, stabilization reduces the accumulation of reactive Qu products that contribute to the proapoptotic effect of the compound, and thus reduces the ability of the compound to induce apoptosis of UVB-irradiated HaCaT cells. PMID:19138980

  12. Delayed persistence of giant-nucleated cells induced by X-ray and proton irradiation in the progeny of replicating normal human f ibroblast cells

    NASA Astrophysics Data System (ADS)

    Almahwasi, A. A.; Jeynes, J. C.; Merchant, M. J.; Bradley, D. A.; Regan, P. H.

    2017-08-01

    Ionising radiation can induce giant-nucleated cells (GCs) in the progeny of irradiated populations, as demonstrated in various cellular systems. Most in vitro studies have utilised quiescent cancerous or normal cell lines but it is not clear whether radiation-induced GCs persist in the progeny of normal replicated cells. In the current work we show persistent induction of GCs in the progeny of normal human-diploid skin fibroblasts (AG1522). These cells were originally irradiated with a single equivalent clinical dose of 0.2, 1 or 2 Gy of either X-ray or proton irradiation and maintained in an active state for various post-irradiation incubation interval times before they were replated for GC analysis. The results demonstrate that the formation of GCs in the progeny of X-ray or proton irradiated cells was increased in a dose-dependent manner when measured 7 days after irradiation and this finding is in agreement with that reported for the AG1522 cells using other radiation qualities. For the 1 Gy X-ray doses it was found that the GC yield increased continually with time up to 21 days post-irradiation. These results can act as benchmark data for such work and may have important implications for studies aimed at evaluating the efficacy of radiation therapy and in determining the risk of delayed effects particularly when applying protons.

  13. The effect of uranyl acetate on human lymphoblastoid cells (RPMI 6410) and HeLa cells.

    PubMed Central

    Ghadially, F. N.; Yang-Steppuhn, S. E.; Lalonde, J. M.

    1982-01-01

    RPMI 6410 cells and HeLa cells were exposed to uranyl acetate. In RPMI 6410 cell cultures this produced single-membrane-bound presumably lysosomal bodies (called "uraniosomes") containing electron-dense crystals in the cultured cells and crystalline deposits in extracellular locations. Neither uraniosomes nor extracellular uranium deposits were found in HeLa cell cultures. All uraniosomes and extracellular uranium deposits analysed by electron-probed X-ray analysis were found to contain uranium, potassium and phosphorus. Traces of sulphur were detected in some but not all uraniosomes and extracellular uranium deposits. Traces of calcium were found in all extracellular uranium deposits and in some uraniosomes also. Images Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:7093141

  14. A comparative study of temperature elevation on human teeth root surfaces during Nd:YAG laser irradiation in root canals.

    PubMed

    Strakas, D; Franzen, R; Kallis, A; Vanweersch, L; Gutknecht, N

    2013-11-01

    The purpose of this study was to evaluate the temperatures on the root surfaces during Nd:YAG laser irradiation in root canals using pulse durations of 180 and 320 μs. Thirty extracted human teeth were used in this study. The teeth were enlarged up to ISO 40 (multi-rooted) or up to ISO 60 (single-rooted) by conventional technique using K-files. Then the teeth were placed into a water bath with a constant temperature of 37 °C and then irradiated with an Nd:YAG laser having an output power of 1.5 W, a frequency of 15 Hz, using an optic fiber of 200 μm diameter. The temperature on the root surface was measured by means of attaching thermocouples in three areas (coronal, mesial, and apical regions) of the root canals. The thermographic study showed that the average temperature elevation for both pulse durations on the root surfaces was less than 9 °C. There was no significant difference in the observed temperatures in coronal and mesial areas. Though a higher increase of temperature was observed in the apical region when the pulse length of the Nd:YAG laser was 320 μs. The results of the study showed that the temperature rises during Nd:YAG laser irradiation with parameters used in this study minimal to cause damage on bone and periodontal tissues. Moreover, it was suggested that in order to have lower temperature in the apical region, an Nd:YAG laser with a pulse length of 180 μs is preferred than one with a pulse length of 320 μs.

  15. Human skin auto-fluorescence decay as a function of irradiance and skin type

    NASA Astrophysics Data System (ADS)

    Debreczeny, Martin P.; Bates, Rebecca; Fitch, Rick M.; Galen, Karen P.; Ge, Jiajia; Dorshow, Richard B.

    2011-03-01

    The aim of this work was to establish measurement conditions under which endogenous skin fluorescence ("auto-fluorescence") is relatively invariant, so that changes in exogenous agents can be accurately determined. Fluorescence emission was measured on the volar forearm of 36 subjects, chosen to be equally representative of all 6 Fitzpatrick skin types. All subjects were exposed to approximately 40 minutes of optical excitation at 450 and 500 nm with 4 irradiances between 0.3 and 9 mW/cm2. Both non-optically-induced (e.g. tissue settling and fluctuation) and optically-induced variations were observed in the measured fluorescence and mechanisms explaining these effects are proposed. The optically-induced auto-fluorescence decay was independent of skin type when excited at 450 nm, but significantly dependent on skin type when excited at 500 nm. Further, the extent of decay over time was linearly related to irradiance at 500 nm, but at 450 nm was non-linear, with the extent of decay rolling off between 2 and 9 mW/cm2. In order to maintain the auto-fluorescence signal within 95% of its original value over a 30 minute period, the excitation at 450 nm would need to be limited to 1.5 mW/cm2, while excitation at 500 nm should be limited to 5 mW/cm2.

  16. Keratin immunoreactivity as an aid to the diagnosis of persistent adenocarcinoma in irradiated human prostates

    SciTech Connect

    Brawer, M.K.; Nagle, R.B.; Pitts, W.; Freiha, F.; Gamble, S.L.

    1989-02-01

    Postirradiation prostatic biopsy is believed by many to be the best measure of radiation effectiveness in prostatic cancer. Therapeutic irradiation may induce prostatic glandular atypia, which in its severe form can be confused with persistent adenocarcinoma on prostatic biopsies. In the current study, 37 postirradiation prostate biopsy specimens were evaluated by immunohistochemistry using a specific monoclonal anticytokeratin antibody (KA1) that reacts with the basal cells of normal or hyperplastic glands, but is nonreactive with the lumenal cells or with prostatic carcinoma cells. Persistent carcinoma was observed in 19 cases in which antibody staining was absent. The noncarcinomatous glands retained reactivity, but this reactivity appeared in a new and previously undescribed pattern. The irradiated lesion was characterized by cellular pleomorphisism, with enlargement of nuclei and loss of polarity. The immunoreactivity was seen in the enlarged basal cells and was seen to focally extend to involve the lumenal cell layer. In five of 37 cases, glands were seen that were so atypical on the routinely stained sections that a distinction from cancer could not be made. These same glands in the adjacent section reacted with KA1 in each case allowing us to conclude that the changes were benign. We conclude that the interpretation of postirradiation prostatic biopsy specimens may be aided by immunohistochemistry with this anticytokeratin antibody.

  17. Porcine circovirus type 2 morphogenesis in a clone derived from the l35 lymphoblastoid cell line.

    PubMed

    Rodríguez-Cariño, C; Duffy, C; Sánchez-Chardi, A; McNeilly, F; Allan, G M; Segalés, J

    2011-01-01

    Porcine circovirus type 2 (PCV2) is the essential infectious agent of post-weaning multisystemic wasting syndrome (PMWS), one of the most important diseases of swine. Although several studies have described different biological properties of the virus, some aspects of its replication cycle, including ultrastructural alterations, remain unknown. The aim of the present study was to describe for the first time a complete morphogenesis study of PCV2 in a clone of the lymphoblastoid L35 cell line at the ultrastructural level using electron microscopy techniques. Cells were infected with PCV2 at a multiplicity of infection of 10 and examined at 0, 6, 12, 24, 48, 60 and 72h post-infection. PCV2 was internalized by endocytosis, after which the virus aggregated in intracytoplasmic inclusion bodies (ICIs). Subsequently, PCV2 was closely associated with mitochondria, completing a first cytoplasmic phase. The virus entered the nucleus for replication and virus assembly and encapsidation occurred with the participation of the nuclear membrane. Immature virions left the nucleus and formed ICIs in a second cytoplasmic phase. The results suggest that at the end of the replication cycle (between 24 and 48h), PCV2 was released either by budding of mature virion clusters or by lysis of apoptotic or dead cells. In conclusion, the L35-derived clone represents a suitable in-vitro model for PCV2 morphogenesis studies and characterization of the PCV2 replication cycle.

  18. Lithium-induced Clock Gene Expression in Lymphoblastoid Cells of Bipolar Affective Patients.

    PubMed

    Kittel-Schneider, S; Schreck, S; Ziegler, C; Weißflog, L; Hilscher, M; Schwarz, R; Schnetzler, L; Neuner, M; Reif, A

    2015-07-01

    Disturbances of circadian rhythms occur in all episodes of bipolar disorder (BD). Lithium, as gold-standard in the maintenance treatment of BD, is known to influence circadian processes. In a pilot study lymphoblastoid cell lines (LCLs) were generated from 8 BD patients and 6 healthy controls. The LCLs were treated with lithiumchloride (LiCl) for 3 weeks. Cell cycles were then synchronized and expressional analysis by quantitative Real Time PCR was done. BD and controls differed in the period length regarding DBP (albumin D-box binding protein) expression and DBP expression was also influenced by lithium treatment. Furthermore, baseline DBP expression was significantly different between non-treated BD and healthy controls. None of the other analyzed circadian genes showed to be influenced by chronic lithium treatment or to be differentially regulated due to the diagnosis. We here show that chronic lithium treatment of LCLs leads to decreased expression of the clock gene DBP, rendering DBP a lithium-regulated gene. We could confirm the role of the circadian clock as well in lithium mode of action as in the pathomechanisms of BD although future studies with a greater number of participants and cell lines are needed. © Georg Thieme Verlag KG Stuttgart · New York.

  19. Epstein-Barr virus genetic variation in lymphoblastoid cell lines derived from Kenyan pediatric population.

    PubMed

    Simbiri, Kenneth O; Smith, Nicholas A; Otieno, Richard; Wohlford, Eric E M; Daud, Ibrahim I; Odada, Sumba P; Middleton, Frank; Rochford, Rosemary

    2015-01-01

    Epstein-Barr virus (EBV) is associated with Burkitt's lymphoma (BL), and in regions of sub-Saharan Africa where endemic BL is common, both the EBV Type 1 (EBV-1) and EBV Type 2 strains (EBV-2) are found. Little is known about genetic variation of EBV strains in areas of sub-Saharan Africa. In the present study, spontaneous lymphoblastoid cell lines (LCLs) were generated from samples obtained from Kenya. Polymerase chain reaction (PCR) amplification of the EBV genome was done using multiple primers and sequenced by next-generation sequencing (NGS). Phylogenetic analyses against the published EBV-1 and EBV-2 strains indicated that one sample, LCL10 was closely related to EBV-2, while the remaining 3 LCL samples were more closely related to EBV-1. Moreover, single nucleotide polymorphism (SNP) analyses showed clustering of LCL variants. We further show by analysis of EBNA-1, BLLF1, BPLF1, and BRRF2 that latent genes are less conserved than lytic genes in these LCLs from a single geographic region. In this study we have shown that NGS is highly useful for deciphering detailed inter and intra-variations in EBV genomes and that within a geographic region different EBV genetic variations can co-exist, the implications of which warrant further investigation. The findings will enhance our understanding of potential pathogenic variants critical to the development and maintenance of EBV-associated malignancies.

  20. Control of cell respiration by nitric oxide in Ataxia Telangiectasia lymphoblastoid cells.

    PubMed

    Masci, Alessandra; Mastronicola, Daniela; Arese, Marzia; Piane, Maria; De Amicis, Andrea; Blanck, Thomas J J; Chessa, Luciana; Sarti, Paolo

    2008-01-01

    Ataxia Telangiectasia (AT) patients are particularly sensitive to oxidative-nitrosative stress. Nitric oxide (NO) controls mitochondrial respiration via the reversible inhibition of complex IV. The mitochondrial response to NO of AT lymphoblastoid cells was investigated. Cells isolated from three patients and three intrafamilial healthy controls were selected showing within each group a normal diploid karyotype and homogeneous telomere length. Different complex IV NO-inhibition patterns were induced by varying the electron flux through the respiratory chain, using exogenous cell membrane permeable electron donors. Under conditions of high electron flux the mitochondrial NO inhibition of respiration was greater in AT than in control cells (P< or =0.05). This property appears peculiar to AT, and correlates well to the higher concentration of cytochrome c detected in the AT cells. This finding is discussed on the basis of the proposed mechanism of reaction of NO with complex IV. It is suggested that the peculiar response of AT mitochondria to NO stress may be relevant to the mitochondrial metabolism of AT patients.

  1. Characterization of an antigen associated with the Marek's disease lymphoblastoid cell line MSB-1.

    PubMed

    Ross, L J

    1982-06-01

    A Marek's disease lymphoblastoid cell line (MSB-1) has been analysed by immunoprecipitation for expression of tumour-associated antigen, Marek's disease virus (MDV)-specific antigens and antigens specific to avian leukosis-sarcoma viruses. Rabbit antisera raised against two independently derived cell lines after extensive absorption with normal chick cells reacted with a polypeptide of mol. wt. 40 000 (40K) in extracts of MSB-1 cells. The 40K polypeptide was not present in myeloblasts or in chick embryo fibroblasts (CEF) infected with MDV and did not react with antiserum raised against normal chicken thymus antigens. The possibility that the 40K polypeptide is a tumour-associated antigen is discussed. Seven MDV-specific antigens were noted in infected CEF (mol. wt. 110K, 100K, 80K, 70K, 50K, 35K and 32K) but none of these was detected in MSB-1 cells. The avian leukosis-sarcoma group-specific antigen P27gag and its precursor Pr76gag were not found in MSB-1 cells, confirming that expression of mature gag protein is not required for transformation by MDV. However, two polypeptides of unknown origin and function (mol. wt. 180K and 110K) were precipitated from MSB-1 cells with a rabbit anti-Rous sarcoma (Schmidt-Rupin, subgroup D) antiserum.

  2. Proteomic analysis of lymphoblastoid cells derived from monozygotic twins discordant for bipolar disorder: a preliminary study.

    PubMed

    Kazuno, An-a; Ohtawa, Kenji; Otsuki, Kaori; Usui, Masaya; Sugawara, Hiroko; Okazaki, Yuji; Kato, Tadafumi

    2013-01-01

    Bipolar disorder is a severe mental illness characterized by recurrent manic and depressive episodes. In bipolar disorder, family and twin studies suggest contributions from genetic and environmental factors; however, the detailed molecular pathogenesis is yet unknown. Thus, identification of biomarkers may contribute to the clinical diagnosis of bipolar disorder. Monozygotic twins discordant for bipolar disorder are relatively rare but have been reported. Here we performed a comparative proteomic analysis of whole cell lysate derived from lymphoblastoid cells of monozygotic twins discordant for bipolar disorder by using two-dimensional differential in-gel electrophoresis (2D-DIGE). We found approximately 200 protein spots to be significantly differentially expressed between the patient and the co-twin (t test, p<0.05). Some of the proteins were subsequently identified by liquid chromatography tandem mass spectrometry and included proteins involved in cell death and glycolysis. To examine whether these proteins could serve as biomarkers of bipolar disorder, we performed Western blot analysis using case-control samples. Expression of phosphoglycerate mutase 1 (PGAM1), which is involved in glycolysis, was significantly up-regulated in patients with bipolar disorder (t test, p<0.05). Although PGAM1 cannot be regarded as a qualified biomarker of bipolar disorder from this preliminary finding, it could be one of the candidates for further study to identify biomarkers of bipolar disorder.

  3. NKG2A-Expressing Natural Killer Cells Dominate the Response to Autologous Lymphoblastoid Cells Infected with Epstein–Barr Virus

    PubMed Central

    Hatton, Olivia; Strauss-Albee, Dara Marie; Zhao, Nancy Q.; Haggadone, Mikel D.; Pelpola, Judith Shanika; Krams, Sheri M.; Martinez, Olivia M.; Blish, Catherine A.

    2016-01-01

    Epstein–Barr virus (EBV) is a human γ-herpesvirus that establishes latency and lifelong infection in host B cells while achieving a balance with the host immune response. When the immune system is perturbed through immunosuppression or immunodeficiency, however, these latently infected B cells can give rise to aggressive B cell lymphomas. Natural killer (NK) cells are regarded as critical in the early immune response to viral infection, but their role in controlling expansion of infected B cells is not understood. Here, we report that NK cells from healthy human donors display increased killing of autologous B lymphoblastoid cell lines (LCLs) harboring latent EBV compared to primary B cells. Coculture of NK cells with autologous EBV+ LCL identifies an NK cell population that produces IFNγ and mobilizes the cytotoxic granule protein CD107a. Multi-parameter flow cytometry and Boolean analysis reveal that these functional cells are enriched for expression of the NK cell receptor NKG2A. Further, NKG2A+ NK cells more efficiently lyse autologous LCL than do NKG2A− NK cells. More specifically, NKG2A+2B4+CD16−CD57−NKG2C−NKG2D+ cells constitute the predominant NK cell population that responds to latently infected autologous EBV+ B cells. Thus, a subset of NK cells is enhanced for the ability to recognize and eliminate autologous, EBV-infected transformed cells, laying the groundwork for harnessing this subset for therapeutic use in EBV+ malignancies. PMID:28018364

  4. Bystander effect in human hepatoma HepG2 cells caused by medium transfers at different times after high-LET carbon ion irradiation

    NASA Astrophysics Data System (ADS)

    Wu, Qingfeng; Li, Qiang; Jin, Xiaodong; Liu, Xinguo; Dai, Zhongying

    2011-01-01

    Although radiation-induced bystander effects have been well documented in a variety of biological systems, whether irradiated cells have the ability to generate bystander signaling persistently is still unclear and the clinical relevance of bystander effects in radiotherapy remains to be elucidated. This study examines tumor cellular bystander response to autologous medium from cell culture irradiated with high-linear energy transfer (LET) heavy ions at a therapeutically relevant dose in terms of clonogenic cell survival. In vitro experiments were performed using human hepatoma HepG2 cell line exposed to 100 keV/μm carbon ions at a dose of 2 Gy. Two different periods (2 and 12 h) after irradiation, irradiated cell conditioned medium (ICCM) and replenished fresh medium were harvested and then transferred to unirradiated bystander cells. Cellular bystander responses were measured with the different medium transfer protocols. Significant higher survival fractions of unirradiated cells receiving the media from the irradiated cultures at the different times post-irradiation than those of the control were observed. Even replenishing fresh medium for unirradiated cells which had been exposed to the ICCM for 12 h could not prevent the bystander cells from the increased survival fraction. These results suggest that the irradiated cells could release unidentified signal factor(s), which induced the increase in survival fraction for the unirradiated bystander cells, into the media sustainedly and the carbon ions triggered a cascade of signaling events in the irradiated cells rather than secreting the soluble signal factor(s) just at a short period after irradiation. Based on the observations in this study, the importance of bystander effect in clinical radiotherapy was discussed and incorporating the bystander effect into the current radiobiological models, which are applicable to heavy ion radiotherapy, is needed urgently.

  5. Low-power laser irradiation promotes the proliferation and osteogenic differentiation of human periodontal ligament cells via cyclic adenosine monophosphate.

    PubMed

    Wu, Jyun-Yi; Chen, Chia-Hsin; Yeh, Li-Yin; Yeh, Ming-Long; Ting, Chun-Chan; Wang, Yan-Hsiung

    2013-06-01

    Retaining or improving periodontal ligament (PDL) function is crucial for restoring periodontal defects. The aim of this study was to evaluate the physiological effects of low-power laser irradiation (LPLI) on the proliferation and osteogenic differentiation of human PDL (hPDL) cells. Cultured hPDL cells were irradiated (660 nm) daily with doses of 0, 1, 2 or 4 J⋅cm(-2). Cell proliferation was evaluated by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, and the effect of LPLI on osteogenic differentiation was assessed by Alizarin Red S staining and alkaline phosphatase (ALP) activity. Additionally, osteogenic marker gene expression was confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR). Our data showed that LPLI at a dose of 2 J⋅cm(-2) significantly promoted hPDL cell proliferation at days 3 and 5. In addition, LPLI at energy doses of 2 and 4 J⋅cm(-2) showed potential osteogenic capacity, as it stimulated ALP activity, calcium deposition, and osteogenic gene expression. We also showed that cyclic adenosine monophosphate (cAMP) is a critical regulator of the LPLI-mediated effects on hPDL cells. This study shows that LPLI can promote the proliferation and osteogenic differentiation of hPDL cells. These results suggest the potential use of LPLI in clinical applications for periodontal tissue regeneration.

  6. Surface Treatment of Polymers by Ion Beam Irradiation to Control the Human Osteoblast Adhesion: Fluence and Current Density Study

    SciTech Connect

    Guibert, G.; Mikhailov, S.; Rossel, T.; Weder, G.; Betschart, B.; Meunier, C.

    2009-03-10

    In the biomaterial field, the modification of surfaces are used to create polymers with high performances, preserving their bulk properties and creating specific interactions between the designed surfaces and the cells or tissues. The polymers were irradiated with a 900 keV Helium beam to modify their surface properties. Cell cultivation on the samples was done using human osteoblasts cells (hFOB 1.19). For PTFE, PS and PEEK polymers, the cell adhesion occurs after reached some threshold values of fluences. For PET or PMMA polymers, the cells adhere on the non irradiated samples, however the fluence value modifies the cell density. For PMMA and PTFE both, the fluence and the current density influence the cell adhesion. By modifying the appropriate parameters on each material, the control of the cell adhesion is possible. Indeed the surface treatment must be selected and adapted according to the further application: for biosensors, tissue engineering, tissue regeneration, neural probes, drug delivery, bio-actuators etc.

  7. Protective Effect of Carvacrol on Oxidative Stress and Cellular DNA Damage Induced by UVB Irradiation in Human Peripheral Lymphocytes.

    PubMed

    Aristatile, Balakrishnan; Al-Numair, Khalid S; Al-Assaf, Abdullah H; Veeramani, Chinnadurai; Pugalendi, Kodukkur Viswanathan

    2015-11-01

    Exposure to ultraviolet B (UVB; 280-320 nm) radiation induces the formation of reactive oxygen species (ROS) in the biological system. In this study, we examined the protective effect of carvacrol on UVB-induced lipid peroxidation and oxidative DNA damage with reference to alterations in cellular an-tioxidant status in human lymphocytes. A series of in vitro assays (hydroxyl radical, superoxide, nitric oxide, DPPH (2,2-Diphenyl-1-picryl hydrazyl), and ABTS (2,2-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid) radical scavenging assays) demonstrate antioxidant property of carvacrol in our study. UVB exposure significantly increased thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LHPs), % tail DNA and tail moment; decreased % cell viability and antioxidant status in UVB-irradiated lymphocytes. Treatment with carvacrol 30 min prior to UVB-exposure resulted in a significant decline of TBARS, LHP, % tail DNA, and tail moment and increased % cell viability as carvacrol concentration increased. UVB irradiated lymphocytes with carvacrol alone (at 10 μg/mL) gave no significant change in cell viability, TBARS, LHP, % tail DNA, and tail moment when compared with normal lymphocytes. On the basis of our results, we conclude that carvacrol, a dietary antioxidant, mediates its protective effect through modulation of UVB-induced ROS.

  8. Effects of light emitting diode irradiation on neural differentiation of human umbilical cord-derived mesenchymal cells.

    PubMed

    Dehghani-Soltani, Samereh; Shojaee, Mohammad; Jalalkamali, Mahshid; Babaee, Abdolreza; Nematollahi-Mahani, Seyed Noureddin

    2017-08-30

    Recently, light emitting diodes (LEDs) have been introduced as a potential physical factor for proliferation and differentiation of various stem cells. Among the mesenchymal stem cells human umbilical cord matrix-derived mesenchymal (hUCM) cells are easily propagated in the laboratory and their low immunogenicity make them more appropriate for regenerative medicine procedures. We aimed at this study to evaluate the effect of red and green light emitted from LED on the neural lineage differentiation of hUCM cells in the presence or absence of retinoic acid (RA). Harvested hUCM cells exhibited mesenchymal and stemness properties. Irradiation of these cells by green and red LED with or without RA pre-treatment successfully differentiated them into neural lineage when the morphology of the induced cells, gene expression pattern (nestin, β-tubulin III and Olig2) and protein synthesis (anti-nestin, anti-β-tubulin III, anti-GFAP and anti-O4 antibodies) was evaluated. These data point for the first time to the fact that LED irradiation and optogenetic technology may be applied for neural differentiation and neuronal repair in regenerative medicine.

  9. Influence of flavonoids and vitamins on the MMP- and TIMP-expression of human dermal fibroblasts after UVA irradiation.

    PubMed

    Hantke, Bernd; Lahmann, Christine; Venzke, Kirsten; Fischer, Tim; Kocourek, Andreas; Windsor, L Jack; Bergemann, Jörg; Stäb, Franz; Tschesche, Harald

    2002-10-01

    UV irradiation leads to distinct changes in skin connective tissue by degradation of collagen, for example. Many of these alterations in the extracellular matrix are mediated by MMPs (matrix metalloproteinases) with reduced content of their antagonist TIMPs (tissue inhibitors of metalloproteinases). Potential candidates to reduce MMP activity in the skin after solar stimulation were examined. The influence of vitamin C, vitamin E and the flavonoids AGR (alpha-glucosylrutin) and 8-prenylnaringenine on the MMP and TIMP expression was investigated. Human dermal fibroblasts were incubated with these additives and irradiated with UVA [10 J cm(-2)]. The gene expression of MMP-1 (collagenase-1) and TIMP-1, the protein expression of MMP-1, MMP-2 (gelatinase-A), TIMP-1 and TIMP-2 as well as the enzyme activity of MMP-1 and MMP-2 were examined. AGR and vitamins C and E were shown to reduce MMP expression and activity, whereas 8-prenylnaringenine appeared to be responsible for the opposite effect. None of the substances considerably influenced the TIMP levels. AGR represented the most effective additive in reducing the collagenase protein expression to 60% and may be useful to level out the MMP activity in the skin after sun exposure. Furthermore, no protein expression of MMP-8, MMP-9, MMP-12 and MMP-13 could be detected.

  10. Survival responses of cell subpopulations isolated from a heterogeneous human colon tumour after combinations of hyperthermia and X-irradiation.

    PubMed

    Leith, J T; Heyman, P; Dewyngaert, J K; Glicksman, A S; Dexter, D L; Calabresi, P

    1983-03-01

    In summary, this research has investigated the effects of combined modality treatment (i.e., low linear energy transfer ionizing radiation and hyperthermia at 42.5 degrees C) on the survival responses of two tumour subpopulations (designated clones A and D) obtained from a heterogeneous human colon adenocarcinoma. A constant hyperthermic exposure (2 hours at 42.5 degrees C) was given either 3 min before or 3 min after graded exposure to X-rays. An isobologram analysis (Steel and Peckham 1979) of the clonogenic survival responses of the two tumour subpopulations showed that the clone A responses were within the envelope of additivity for either sequence of application. In contrast, the responses of the clone D tumour subpopulation exhibited a supra-additive response to the combined treatments with the sequence of heat followed by X-irradiation being somewhat more effective than the sequence of X-irradiation followed by heat. These data indicate that the responses of tumour subpopulations obtained from heterogeneous solid tumours to combined modality treatments may vary in an, at present, unpredictable manner.

  11. Delayed expression of apoptosis in X-irradiated human leukemic MOLT-4 cells transfected with mutant p53.

    PubMed

    Nakano, Hisako; Yonekawa, Hiromichi; Shinohara, Kunio

    2003-06-01

    The effects of X-rays on cell survival, apoptosis, and long-term response in the development of cell death as measured by the dye exclusion test were studied in human leukemic MOLT-4 cells (p53 wild-type) stably transfected with a mutant p53 cDNA expression vector. Cell survival, as determined from colony-forming ability, was increased in an expression level dependent manner, but the increase was partial even with the highest-expressing clone (B3). This contrasts with the prior observation that cell death and apoptosis in B3 are completely inhibited at 24 h after irradiation with 1.8 Gy of X-rays. The examination of B3 cells incubated for longer than 24 h after X-irradiation showed a delay in the induction of cell death and apoptosis. Western blot analysis revealed that the time required to reach the highest level of wild-type p53 protein in B3 was longer than the time in MOLT-4 and that the p53 may be stabilized by the phosphorylation at Ser-15. These results suggest that the introduction of mutant p53 into MOLT-4 merely delays the development of apoptosis, during which the cells could repair the damage induced by X-rays, and results in the partial increase in cell survival.

  12. ESA IBER-2 Molecular and Cellular Changes in Human Endothelial Cells in Response to Nickel Ion Irradiation (CORALS project)

    NASA Astrophysics Data System (ADS)

    Moreels, M.; Quintens, R.; De Vos, W.; Beck, M.; Tabury, K.; Suetens, A.; Abouelaradat, K.; Dieriks, B.; Ernst, E.; Lee, R.; Lambert, C.; Van Oostveldt, P.; Baatout, S.

    2013-02-01

    On Earth, most radiation exposures (medical and natural background) consist of low-linear energy transfer (LET) photons. In space, astronauts are exposed to higher doses and to more varied types of radiation. Cosmic radiation mainly consists of high-energy protons and high-Z and -energy (HZE) particles. These high-LET particles are predicted to account for most of the radiation induced health effects. In this regard, further analysis of the biological effects of HZE particles is essential. In the present study, endothelial cells were irradiated with different doses of nickel ions produced in the synchrotron at GSI (Darmstadt, Germany). After different time points, RNA was extracted for genome-wide analysis and supernatants were collected for multiplex cytokine assay. DNA double strand breaks were detected using γH2AX staining. Our results demonstrated that nickel irradiation induced molecular and cellular changes in human endothelial cells. Further analysis is ongoing to confirm the obtained data and to further explore the biological effects after nickel ion exposure.

  13. Effects of PBM in different energy densities and irradiance on maintaining cell viability and proliferation of pulp fibroblasts from human primary teeth.

    PubMed

    Marques, Nádia Carolina Teixeira; Neto, Natalino Lourenço; Prado, Mariel Tavares Oliveira; Vitor, Luciana Lourenço Ribeiro; Oliveira, Rodrigo Cardoso; Sakai, Vivien Thiemy; Santos, Carlos Ferreira; Machado, Maria Aparecida Andrade Moreira; Oliveira, Thais Marchini

    2017-08-11

    This study aimed to compare the effects of photobiomodulation (PBM) in different energy densities and irradiances on maintaining cell viability, and proliferation of pulp fibroblasts from human primary teeth (HPF) were cultured in DMEM and used between the fourth and eighth passages. Then, HPF were irradiated with the following different energy densities: 1.25 J/cm(2) (a), 2.50 J/cm(2) (b), 3.75 J/cm(2) (c), 5.00 J/cm(2) (d), and 6.25 J/cm(2) (e); but varying either the time of irradiation (groups 1a-1e) or the output power (groups 2a-2e). Positive (groups 1f and 2f) and negative controls (groups 1g and 2g), respectively, comprised non-irradiated cells grown in regular nutritional conditions (10% fetal bovine serum [FBS]) and under nutritional deficit (1% FBS). Cell viability and proliferation were respectively assessed through MTT and crystal violet (CV) assays at 24, 48, and 72 h after irradiation. Statistical analysis was performed by two-way ANOVA, followed by Tukey test (P < 0.05). The negative controls showed significantly lower viability in relation to most of the corresponding subgroups, both for MTT and CV assays. For both assays, the intragroup comparison showed that the periods of 24 h exhibited lower viability than the periods of 48 and 72 h for most of the subgroups, except the negative controls with lower viability. The different irradiation protocols (equal energy densities applied with different irradiances) showed no statistically significant differences on cell viability and proliferation at the evaluated periods. The proposed PBM in different energy densities and irradiance did not affect the viability and proliferation of pulp fibroblasts from human primary teeth.

  14. An in vitro cell irradiation protocol for testing photopharmaceuticals and the effect of blue, green, and red light on human cancer cell lines.

    PubMed

    Hopkins, S L; Siewert, B; Askes, S H C; Veldhuizen, P; Zwier, R; Heger, Michal; Bonnet, Sylvestre

    2016-05-11

    Traditionally, ultraviolet light (100-400 nm) is considered an exogenous carcinogen while visible light (400-780 nm) is deemed harmless. In this work, a LED irradiation system for in vitro photocytotoxicity testing is described. The LED irradiation system was developed for testing photopharmaceutical drugs, but was used here to determine the basal level response of human cancer cell lines to visible light of different wavelengths, without any photo(chemo)therapeutic. The effects of blue (455 nm, 10.5 mW cm(-2)), green (520 nm, 20.9 mW cm(-2)), and red light (630 nm, 34.4 mW cm(-2)) irradiation was measured for A375 (human malignant melanoma), A431 (human epidermoid carcinoma), A549 (human lung carcinoma), MCF7 (human mammary gland adenocarcinoma), MDA-MB-231 (human mammary gland adenocarcinoma), and U-87 MG (human glioblastoma-grade IV) cell lines. In response to a blue light dose of 19 J cm(-2), three cell lines exhibited a minimal (20%, MDA-MB-231) to moderate (30%, A549 and 60%, A375) reduction in cell viability, compared to dark controls. The other cell lines were not affected. Effective blue light doses that produce a therapeutic response in 50% of the cell population (ED50) compared to dark conditions were found to be 10.9 and 30.5 J cm(-2) for A375 and A549 cells, respectively. No adverse effects were observed in any of the six cell lines irradiated with a 19 J cm(-2) dose of 520 nm (green) or 630 nm (red) light. The results demonstrate that blue light irradiation can have an effect on the viability of certain human cancer cell types and controls should be used in photopharmaceutical testing, which uses high-energy (blue or violet) visible light activation.