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Sample records for radiation-induced apoptotic cell

  1. Anti-apoptotic peptides protect against radiation-induced cell death.

    PubMed

    McConnell, Kevin W; Muenzer, Jared T; Chang, Kathy C; Davis, Chris G; McDunn, Jonathan E; Coopersmith, Craig M; Hilliard, Carolyn A; Hotchkiss, Richard S; Grigsby, Perry W; Hunt, Clayton R

    2007-04-06

    The risk of terrorist attacks utilizing either nuclear or radiological weapons has raised concerns about the current lack of effective radioprotectants. Here it is demonstrated that the BH4 peptide domain of the anti-apoptotic protein Bcl-xL can be delivered to cells by covalent attachment to the TAT peptide transduction domain (TAT-BH4) and provide protection in vitro and in vivo from radiation-induced apoptotic cell death. Isolated human lymphocytes treated with TAT-BH4 were protected against apoptosis following exposure to 15Gy radiation. In mice exposed to 5Gy radiation, TAT-BH4 treatment protected splenocytes and thymocytes from radiation-induced apoptotic cell death. Most importantly, in vivo radiation protection was observed in mice whether TAT-BH4 treatment was given prior to or after irradiation. Thus, by targeting steps within the apoptosis signaling pathway it is possible to develop post-exposure treatments to protect radio-sensitive tissues.

  2. SIGN-R1 and complement factors are involved in the systemic clearance of radiation-induced apoptotic cells in whole-body irradiated mice

    SciTech Connect

    Park, Jin-Yeon; Loh, SoHee; Cho, Eun-hee; Choi, Hyeong-Jwa; Na, Tae-Young; Nemeno, Judee Grace E.; Lee, Jeong Ik; Yoon, Taek Joon; Choi, In-Soo; Lee, Minyoung; Lee, Jae-Seon; Kang, Young-Sun

    2015-08-07

    Although SIGN-R1-mediated complement activation pathway has been shown to enhance the systemic clearance of apoptotic cells, the role of SIGN-R1 in the clearance of radiation-induced apoptotic cells has not been characterized and was investigated in this study. Our data indicated that whole-body γ-irradiation of mice increased caspase-3{sup +} apoptotic lymphocyte numbers in secondary lymphoid organs. Following γ-irradiation, SIGN-R1 and complements (C4 and C3) were simultaneously increased only in the mice spleen tissue among the assessed tissues. In particular, C3 was exclusively activated in the spleen. The delayed clearance of apoptotic cells was markedly prevalent in the spleen and liver of SIGN-R1 KO mice, followed by a significant increase of CD11b{sup +} cells. These results indicate that SIGN-R1 and complement factors play an important role in the systemic clearance of radiation-induced apoptotic innate immune cells to maintain tissue homeostasis after γ-irradiation. - Highlights: • Splenic SIGN-R1{sup +} macrophages are activated after γ-irradiation. • C3 and C4 levels increased and C3 was activated in the spleen after γ-irradiation. • SIGN-R1 mediated the systemic clearance of radiation-induced apoptotic cells in spleen and liver.

  3. Podophyllotoxin and Rutin Modulates Ionizing Radiation-Induced Oxidative Stress and Apoptotic Cell Death in Mice Bone Marrow and Spleen

    PubMed Central

    Singh, Abhinav; Yashavarddhan, M. H.; Kalita, Bhargab; Ranjan, Rajiv; Bajaj, Sania; Prakash, Hridayesh; Gupta, Manju Lata

    2017-01-01

    The present study is aimed to investigate the radioprotective efficacy of G-003M (combination of podophyllotoxin and rutin) against gamma radiation-induced oxidative stress and subsequent cell death in mice bone marrow and spleen. Prophylactic administration of G-003M (−1 h) rendered more than 85% survival in mice exposed to 9 Gy (lethal dose) with dose reduction factor of 1.26. G-003M pretreated mice demonstrated significantly reduced level of reactive oxygen species, membrane lipid peroxidation, and retained glutathione level. In the same group, we obtained increased expression of master redox regulator, nuclear factor erythroid-derived like-2 factor (Nrf-2), and its downstream targets (heme oxygenase-1, Nqo-1, glutathione S-transferase, and thioredoxin reductase-1). In addition, G-003M preadministration has also shown a significant reduction in Keap-1 level (Nrf-2 inhibitor). Radiation-induced lethality was significantly amended in combination-treated (G-003M) mice as demonstrated by reduced 8-OHdG, annexin V FITC+ cells, and restored mitochondrial membrane potential. Expression of antiapoptotic protein Bcl-2 and Bcl-xL was restored in G-003M pretreated group. However, proapoptotic proteins (Puma, Bax, Bak, Caspase-3, and Caspase-7) were significantly declined in this group. Further analysis of immune cells revealed G-003M-mediated restoration of CD3 and CD19 receptor, which was found decreased to significant level following irradiation. Similarly, Gr-1, a marker of granulocytes, was also retained by G-003M administration prior to radiation. Modulatory potential of this formulation (G-003M) can be exploited as a safe and effective countermeasure against radiation-induced lymphohemopoietic injury. PMID:28289414

  4. Podophyllotoxin and Rutin Modulates Ionizing Radiation-Induced Oxidative Stress and Apoptotic Cell Death in Mice Bone Marrow and Spleen.

    PubMed

    Singh, Abhinav; Yashavarddhan, M H; Kalita, Bhargab; Ranjan, Rajiv; Bajaj, Sania; Prakash, Hridayesh; Gupta, Manju Lata

    2017-01-01

    The present study is aimed to investigate the radioprotective efficacy of G-003M (combination of podophyllotoxin and rutin) against gamma radiation-induced oxidative stress and subsequent cell death in mice bone marrow and spleen. Prophylactic administration of G-003M (-1 h) rendered more than 85% survival in mice exposed to 9 Gy (lethal dose) with dose reduction factor of 1.26. G-003M pretreated mice demonstrated significantly reduced level of reactive oxygen species, membrane lipid peroxidation, and retained glutathione level. In the same group, we obtained increased expression of master redox regulator, nuclear factor erythroid-derived like-2 factor (Nrf-2), and its downstream targets (heme oxygenase-1, Nqo-1, glutathione S-transferase, and thioredoxin reductase-1). In addition, G-003M preadministration has also shown a significant reduction in Keap-1 level (Nrf-2 inhibitor). Radiation-induced lethality was significantly amended in combination-treated (G-003M) mice as demonstrated by reduced 8-OHdG, annexin V FITC(+) cells, and restored mitochondrial membrane potential. Expression of antiapoptotic protein Bcl-2 and Bcl-xL was restored in G-003M pretreated group. However, proapoptotic proteins (Puma, Bax, Bak, Caspase-3, and Caspase-7) were significantly declined in this group. Further analysis of immune cells revealed G-003M-mediated restoration of CD3 and CD19 receptor, which was found decreased to significant level following irradiation. Similarly, Gr-1, a marker of granulocytes, was also retained by G-003M administration prior to radiation. Modulatory potential of this formulation (G-003M) can be exploited as a safe and effective countermeasure against radiation-induced lymphohemopoietic injury.

  5. Radiation-induced transmissable chromosomal instability in haemopoietic stem cells

    NASA Astrophysics Data System (ADS)

    Kadhim, M. A.; Wright, E. G.

    Heritable radiation-induced genetic alterations have long been assumed to be ``fixed'' within the first cell division. However, there is a growing body of evidence that a considerable fraction of cells surviving radiation exposure appear normal, but a variety of mutational changes arise in their progeny due to a transmissible genomic instability. In our investigations of G-banded metaphases, non-clonal cytogenetic aberrations, predominantly chromatid-type aberrations, have been observed in the clonal descendants of murine and human haemopoietic stem cells surviving low doses (~1 track per cell) of alpha-particle irradiations. The data are consistent with a transmissible genetic instability induced in a stem cell resulting in a diversity of chromosomal aberrations in its clonal progeny many cell divisions later. Recent studies have demonstrated that the instability phenotype persists in vivo and that the expression of chromosomal instability has a strong dependence on the genetic characteristics of the irradiated cell. At the time when cytogenetic aberrations are detected, an increased incidence of hprt mutations and apoptotic cells have been observed in the clonal descendants of alpha-irradiated murine haemopoietic stem cells. Thus, delayed chromosomal abnormalities, delayed cell death by apoptosis and late-arising specific gene mutations may reflect diverse consequences of radiation-induced genomic instability. The relationship, if any, between these effects is not established. Current studies suggest that expression of these delayed heritable effects is determined by the type of radiation exposure, type of cell and a variety of genetic factors.

  6. The ethyl acetate fraction of Sargassum muticum attenuates ultraviolet B radiation-induced apoptotic cell death via regulation of MAPK- and caspase-dependent signaling pathways in human HaCaT keratinocytes.

    PubMed

    Piao, Mei Jing; Kim, Ki Cheon; Zheng, Jian; Yao, Cheng Wen; Cha, Ji Won; Boo, Sun Jin; Yoon, Weon Jong; Kang, Hee Kyoung; Yoo, Eun Sook; Koh, Young Sang; Ko, Mi Hee; Lee, Nam Ho; Hyun, Jin Won

    2014-09-01

    Our previous work demonstrated that an ethyl acetate extract derived from Sargassum muticum (Yendo) Fenshol (SME) protected human HaCaT keratinocytes against ultraviolet B (UVB)-induced oxidative stress by increasing antioxidant activity in the cells, thereby inhibiting apoptosis. The aim of the current study was to further elucidate the anti-apoptotic mechanism of SME against UVB-induced cell damage. The expression levels of several apoptotic-associated and mitogen-activated kinase (MAPK) signaling proteins were determined by western blot analysis of UVB-irradiated HaCaT cells with or without prior SME treatment. In addition, the loss of mitochondrial membrane potential (Δψm) was detected using flow cytometry or confocal microscopy and the mitochondria membrane-permeate dye, JC-1. Apoptosis was assessed by quantifying DNA fragmentation and apoptotic body formation. Furthermore, cell viability was evaluated using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. SME absorbed electromagnetic radiation in the UVB range (280-320 nm) of the UV/visible light spectrum. SME also increased Bcl-2 and Mcl-1 expression in UVB-irradiated cells and decreased the Bax expression. Moreover, SME inhibited the UVB-induced disruption of mitochondrial membrane potential and prevented UVB-mediated increases in activated caspase-9 and caspase-3 (an apoptotic initiator and executor, respectively) levels. Notably, treatment with a pan-caspase inhibitor enhanced the anti-apoptotic effects of SME in UVB-irradiated cells. Finally, SME reduced the UVB-mediated phosphorylation of p38 MAPK and JNK, and prevented the UVB-mediated dephosphorylation of Erk1/2 and Akt. The present results indicate that SME safeguards HaCaT keratinocytes from UVB-mediated apoptosis by inhibiting a caspase-dependent signaling pathway.

  7. Effects of ceramide inhibition on radiation-induced apoptosis in human leukemia MOLT-4 cells.

    PubMed

    Takahashi, Eriko; Inanami, Osamu; Asanuma, Taketoshi; Kuwabara, Mikinori

    2006-03-01

    In the present study, using inhibitors of ceramide synthase (fumonisin B1), ketosphinganine synthetase (L-cycloserine), acid sphingomyelinase (D609 and desipramine) and neutral sphingomyelinase (GW4869), the role of ceramide in X-ray-induced apoptosis was investigated in MOLT-4 cells. The diacylglycerol kinase (DGK) assay showed that the intracellular concentration of ceramide increased time-dependently after X irradiation of cells, and this radiation-induced accumulation of ceramide did not occur prior to the appearance of apoptotic cells. Treatment with D609 significantly inhibited radiation-induced apoptosis, but did not inhibit the increase of intracellular ceramide. Treatment with desipramine or GW4869 prevented neither radiation-induced apoptosis nor the induced increase of ceramide. On the other hand, fumonisin B1 and L-cycloserine had no effect on the radiation-induced induction of apoptosis, in spite of significant inhibition of the radiation-induced ceramide. From these results, it was suggested that the increase of the intracellular concentration of ceramide was not essential for radiation-induced apoptosis in MOLT-4 cells.

  8. Ionizing Radiation-Induced Endothelial Cell Senescence and Cardiovascular Diseases

    PubMed Central

    Wang, Yingying; Boerma, Marjan; Zhou, Daohong

    2016-01-01

    Exposure to ionizing radiation induces not only apoptosis but also senescence. While the role of endothelial cell apoptosis in mediating radiation-induced acute tissue injury has been extensively studied, little is known about the role of endothelial cell senescence in the pathogenesis of radiation-induced late effects. Senescent endothelial cells exhibit decreased production of nitric oxide and expression of thrombomodulin, increased expression of adhesion molecules, elevated production of reactive oxygen species and inflammatory cytokines and an inability to proliferate and form capillary-like structures in vitro. These findings suggest that endothelial cell senescence can lead to endothelial dysfunction by dysregulation of vasodilation and hemostasis, induction of oxidative stress and inflammation and inhibition of angiogenesis, which can potentially contribute to radiation-induced late effects such as cardiovascular diseases (CVDs). In this article, we discuss the mechanisms by which radiation induces endothelial cell senescence, the roles of endothelial cell senescence in radiation-induced CVDs and potential strategies to prevent, mitigate and treat radiation-induced CVDs by targeting senescent endothelial cells. PMID:27387862

  9. PAI-1-Dependent Endothelial Cell Death Determines Severity of Radiation-Induced Intestinal Injury

    PubMed Central

    Abderrahmani, Rym; François, Agnes; Buard, Valerie; Tarlet, Georges; Blirando, Karl; Hneino, Mohammad; Vaurijoux, Aurelie; Benderitter, Marc; Sabourin, Jean-Christophe; Milliat, Fabien

    2012-01-01

    Normal tissue toxicity still remains a dose-limiting factor in clinical radiation therapy. Recently, plasminogen activator inhibitor type 1 (SERPINE1/PAI-1) was reported as an essential mediator of late radiation-induced intestinal injury. However, it is not clear whether PAI-1 plays a role in acute radiation-induced intestinal damage and we hypothesized that PAI-1 may play a role in the endothelium radiosensitivity. In vivo, in a model of radiation enteropathy in PAI-1 −/− mice, apoptosis of radiosensitive compartments, epithelial and microvascular endothelium was quantified. In vitro, the role of PAI-1 in the radiation-induced endothelial cells (ECs) death was investigated. The level of apoptotic ECs is lower in PAI-1 −/− compared with Wt mice after irradiation. This is associated with a conserved microvascular density and consequently with a better mucosal integrity in PAI-1 −/− mice. In vitro, irradiation rapidly stimulates PAI-1 expression in ECs and radiation sensitivity is increased in ECs that stably overexpress PAI-1, whereas PAI-1 knockdown increases EC survival after irradiation. Moreover, ECs prepared from PAI-1 −/− mice are more resistant to radiation-induced cell death than Wt ECs and this is associated with activation of the Akt pathway. This study demonstrates that PAI-1 plays a key role in radiation-induced EC death in the intestine and suggests that this contributes strongly to the progression of radiation-induced intestinal injury. PMID:22563394

  10. Acidic polysaccharide of Panax ginseng regulates the mitochondria/caspase-dependent apoptotic pathway in radiation-induced damage to the jejunum in mice.

    PubMed

    Bing, So Jin; Kim, Min Ju; Ahn, Ginnae; Im, Jaehak; Kim, Dae Seung; Ha, Danbee; Cho, Jinhee; Kim, Areum; Jee, Youngheun

    2014-04-01

    Owing to its susceptibility to radiation, the small intestine of mice is valuable for studying radioprotective effects. When exposed to radiation, intestinal crypt cells immediately go through apoptosis, which impairs swift differentiation necessary for the regeneration of intestinal villi. Our previous studies have elucidated that acidic polysaccharide of Panax ginseng (APG) protects the mouse small intestine from radiation-induced damage by lengthening villi with proliferation and repopulation of crypt cells. In the present study, we identified the molecular mechanism involved. C57BL/6 mice were irradiated with gamma-rays with or without APG and the expression levels of apoptosis-related molecules in the jejunum were investigated using immunohistochemistry. APG pretreatment strongly decreased the radiation-induced apoptosis in the jejunum. It increased the expression levels of anti-apoptotic proteins (Bcl-2 and Bcl-XS/L) and dramatically reduced the expression levels of pro-apoptotic proteins (p53, BAX, cytochrome c and caspase-3). Therefore, APG attenuated the apoptosis through the intrinsic pathway, which is controlled by p53 and Bcl-2 family members. Results presented in this study suggest that APG protects the mouse small intestine from irradiation-induced apoptosis through inhibition of the p53-dependent pathway and the mitochondria/caspase pathway. Thus, APG may be a potential agent for preventing radiation induced injuries in intestinal cells during radio-therapy such as in cancer treatment.

  11. Radiation-Induced Bystander Effects in Cultured Human Stem Cells

    PubMed Central

    Sokolov, Mykyta V.; Neumann, Ronald D.

    2010-01-01

    Background The radiation-induced “bystander effect” (RIBE) was shown to occur in a number of experimental systems both in vitro and in vivo as a result of exposure to ionizing radiation (IR). RIBE manifests itself by intercellular communication from irradiated cells to non-irradiated cells which may cause DNA damage and eventual death in these bystander cells. It is known that human stem cells (hSC) are ultimately involved in numerous crucial biological processes such as embryologic development; maintenance of normal homeostasis; aging; and aging-related pathologies such as cancerogenesis and other diseases. However, very little is known about radiation-induced bystander effect in hSC. To mechanistically interrogate RIBE responses and to gain novel insights into RIBE specifically in hSC compartment, both medium transfer and cell co-culture bystander protocols were employed. Methodology/Principal Findings Human bone-marrow mesenchymal stem cells (hMSC) and embryonic stem cells (hESC) were irradiated with doses 0.2 Gy, 2 Gy and 10 Gy of X-rays, allowed to recover either for 1 hr or 24 hr. Then conditioned medium was collected and transferred to non-irradiated hSC for time course studies. In addition, irradiated hMSC were labeled with a vital CMRA dye and co-cultured with non-irradiated bystander hMSC. The medium transfer data showed no evidence for RIBE either in hMSC and hESC by the criteria of induction of DNA damage and for apoptotic cell death compared to non-irradiated cells (p>0.05). A lack of robust RIBE was also demonstrated in hMSC co-cultured with irradiated cells (p>0.05). Conclusions/Significance These data indicate that hSC might not be susceptible to damaging effects of RIBE signaling compared to differentiated adult human somatic cells as shown previously. This finding could have profound implications in a field of radiation biology/oncology, in evaluating radiation risk of IR exposures, and for the safety and efficacy of hSC regenerative

  12. Inactivation of Kupffer Cells by Gadolinium Chloride Protects Murine Liver From Radiation-Induced Apoptosis

    SciTech Connect

    Du Shisuo; Qiang Min; Zeng Zhaochong; Ke Aiwu; Ji Yuan; Zhang Zhengyu; Zeng Haiying; Liu Zhongshan

    2010-03-15

    Purpose: To determine whether the inhibition of Kupffer cells before radiotherapy (RT) would protect hepatocytes from radiation-induced apoptosis. Materials and Methods: A single 30-Gy fraction was administered to the upper abdomen of Sprague-Dawley rats. The Kupffer cell inhibitor gadolinium chloride (GdCl3; 10 mg/kg body weight) was intravenously injected 24 h before RT. The rats were divided into four groups: group 1, sham RT plus saline (control group); group 2, sham RT plus GdCl3; group 3, RT plus saline; and group 4, RT plus GdCl3. Liver tissue was collected for measurement of apoptotic cytokine expression and evaluation of radiation-induced liver toxicity by analysis of liver enzyme activities, hepatocyte micronucleus formation, apoptosis, and histologic staining. Results: The expression of interleukin-1beta, interleukin-6, and tumor necrosis factor-alpha was significantly attenuated in group 4 compared with group 3 at 2, 6, 24, and 48 h after injection (p <0.05). At early points after RT, the rats in group 4 exhibited significantly lower levels of liver enzyme activity, apoptotic response, and hepatocyte micronucleus formation compared with those in group 3. Conclusion: Selective inactivation of Kupffer cells with GdCl3 reduced radiation-induced cytokine production and protected the liver against acute radiation-induced damage.

  13. Radiation-induced bystander effect in non-irradiated glioblastoma spheroid cells.

    PubMed

    Faqihi, Fahime; Neshastehriz, Ali; Soleymanifard, Shokouhozaman; Shabani, Robabeh; Eivazzadeh, Nazila

    2015-09-01

    Radiation-induced bystander effects (RIBEs) are detected in cells that are not irradiated but receive signals from treated cells. The present study explored these bystander effects in a U87MG multicellular tumour spheroid model. A medium transfer technique was employed to induce the bystander effect, and colony formation assay was used to evaluate the effect. Relative changes in expression of BAX, BCL2, JNK and ERK genes were analysed using RT-PCR to investigate the RIBE mechanism. A significant decrease in plating efficiency was observed for both bystander and irradiated cells. The survival fraction was calculated for bystander cells to be 69.48% and for irradiated cells to be 34.68%. There was no change in pro-apoptotic BAX relative expression, but anti-apoptotic BCL2 showed downregulation in both irradiated and bystander cells. Pro-apoptotic JNK in bystander samples and ERK in irradiated samples were upregulated. The clonogenic survival data suggests that there was a classic RIBE in U87MG spheroids exposed to 4 Gy of X-rays, using a medium transfer technique. Changes in the expression of pro- and anti-apoptotic genes indicate involvement of both intrinsic apoptotic and MAPK pathways in inducing these effects.

  14. Radiation induced genomic instability in bystander cells

    NASA Astrophysics Data System (ADS)

    Zhou, H.; Gu, S.; Randers-Pehrson, G.; Hei, T.

    There is considerable evidence that exposure to ionizing radiation may induce a heritable genomic instability that leads to a persisting increased frequency of genetic and functional changes in the non-irradiated progeny of a wide variety of irradiated cells Genomic instability is measured as delayed expressions in chromosomal alterations micronucleus formation gene mutations and decreased plating efficiency During the last decade numerous studies have shown that radiation could induce bystander effect in non-irradiated neighboring cells similar endpoints have also been used in genomic instability studies Both genomic instability and the bystander effect are phenomena that result in a paradigm shift in our understanding of radiation biology In the past it seemed reasonable to assume that the production of single- and double-strand DNA breaks are due to direct energy deposition of energy by a charged particle to the nucleus It turns out that biology is not quite that simple Using the Columbia University charged particle microbeam and the highly sensitive human hamster hybrid AL cell mutagenic assay we irradiated 10 of the cells with a lethal dose of 30 alpha particles through the nucleus After overnight incubation the remaining viable bystander cells were replated in dishes for colony formation Clonal isolates were expanded and cultured for 6 consecutive weeks to assess plating efficiency and mutation frequency Preliminary results indicated that there was no significant decrease in plating efficiency among the bystander colonies when compared with

  15. Mensenchymal stem cells can delay radiation-induced crypt death: impact on intestinal CD44(+) fragments.

    PubMed

    Chang, Peng-Yu; Jin, Xing; Jiang, Yi-Yao; Wang, Li-Xian; Liu, Yong-Jun; Wang, Jin

    2016-05-01

    Intestinal stem cells are primitive cells found within the intestinal epithelium that play a central role in maintaining epithelial homeostasis through self-renewal and commitment into functional epithelial cells. Several markers are available to identify intestinal stem cells, such as Lgr5, CD24 and EphB2, which can be used to sort intestinal stem cells from mammalian gut. Here, we identify and isolate intestinal stem cells from C57BL/6 mice by using a cell surface antigen, CD44. In vitro, some CD44(+) crypt cells are capable of forming "villus-crypt"-like structures (organoids). A subset strongly positive for CD44 expresses high levels of intestinal stem-cell-related genes, including Lgr5, Bmi1, Hopx, Lrig1, Ascl2, Smoc2 and Rnf43. Cells from this subset are more capable of developing into organoids in vitro, compared with the subset weakly positive for CD44. However, the organoids are sensitive to ionizing irradiation. We investigate the specific roles of mesenchymal stem cells in protecting organoids against radiation-induced crypt death. When co-cultured with mesenchymal stem cells, the crypt domains of irradiated organoids possess more proliferative cells and fewer apoptotic cells than those not co-cultured with mesenchymal stem cells. Cd44v6 continues to be expressed in the crypt domains of irradiated organoids co-cultured with mesenchymal stem cells. Our results indicate specific roles of mesenchymal stem cells in delaying radiation-induced crypt death in vitro.

  16. Mesenchymal stem cell-conditioned medium prevents radiation-induced liver injury by inhibiting inflammation and protecting sinusoidal endothelial cells.

    PubMed

    Chen, Yi-Xing; Zeng, Zhao-Chong; Sun, Jing; Zeng, Hai-Ying; Huang, Yan-; Zhang, Zhen-Yu

    2015-07-01

    Current management of radiation-induced liver injury is limited. Sinusoidal endothelial cell (SEC) apoptosis and inflammation are considered to be initiating events in hepatic damage. We hypothesized that mesenchymal stem cells (MSCs) possess anti-apoptotic and anti-inflammatory actions during hepatic irradiation, acting via paracrine mechanisms. This study aims to examine whether MSC-derived bioactive components are protective against radiation-induced liver injury in rats. MSC-conditioned medium (MSC-CM) was generated from rat bone marrow-derived MSCs. The effect of MSC-CM on the viability of irradiated SECs was examined by flow cytometric analysis. Activation of the Akt and ERK pathways was analyzed by western blot. MSC-CM was also delivered to Sprague-Dawley rats immediately before receiving liver irradiation, followed by testing for pathological features, changes in serum hyaluronic acid, ALT, and inflammatory cytokine levels, and liver cell apoptosis. MSC-CM enhanced the viability of irradiated SECs in vitro and induced Akt and ERK phosphorylation in these cells. Infusion of MSC-CM immediately before liver irradiation provided a significant anti-apoptotic effect on SECs and improved the histopathological features of injury in the irradiated liver. MSC-CM also reduced the secretion and expression of inflammatory cytokines and increased the expression of anti-inflammatory cytokines. MSC-derived bioactive components could be a novel therapeutic approach for treating radiation-induced liver injury.

  17. Genetic variation in radiation-induced cell death.

    PubMed

    Smirnov, Denis A; Brady, Lauren; Halasa, Krzysztof; Morley, Michael; Solomon, Sonia; Cheung, Vivian G

    2012-02-01

    Radiation exposure through environmental, medical, and occupational settings is increasingly common. While radiation has harmful effects, it has utility in many applications such as radiotherapy for cancer. To increase the efficacy of radiation treatment and minimize its risks, a better understanding of the individual differences in radiosensitivity and the molecular basis of radiation response is needed. Here, we integrated human genetic and functional genomic approaches to study the response of human cells to radiation. We measured radiation-induced changes in gene expression and cell death in B cells from normal individuals. We found extensive individual variation in gene expression and cellular responses. To understand the genetic basis of this variation, we mapped the DNA sequence variants that influence expression response to radiation. We also identified radiation-responsive genes that regulate cell death; silencing of these genes by small interfering RNA led to an increase in radiation-induced cell death in human B cells, colorectal and prostate cancer cells. Together these results uncovered DNA variants that contribute to radiosensitivity and identified genes that can be targeted to increase the sensitivity of tumors to radiation.

  18. Immunosuppressive effects of apoptotic cells

    NASA Astrophysics Data System (ADS)

    Voll, Reinhard E.; Herrmann, Martin; Roth, Edith A.; Stach, Christian; Kalden, Joachim R.; Girkontaite, Irute

    1997-11-01

    Apoptotic cell death is important in the development and homeostasis of multicellular organisms and is a highly controlled means of eliminating dangerous, damaged or unnecessary cells without causing an inflammatory response or tissue damage,. We now show that the presence of apoptotic cells during monocyte activation increases their secretion of the anti-inflammatory and immunoregulatory cytokine interleukin 10 (IL-10) and decreases secretion of the proinflammatory cytokines tumour necrosis factor-α (TNF-α), IL-1 and IL-12. This may inhibit inflammation and contribute to impaired cell-mediated immunity in conditions associated with increased apoptosis, such as viral infections, pregnancy, cancer and exposure to radiation.

  19. DNA damage in cells exhibiting radiation-induced genomic instability

    SciTech Connect

    Keszenman, Deborah J.; Kolodiuk, Lucia; Baulch, Janet E.

    2015-02-22

    Cells exhibiting radiation induced genomic instability exhibit varied spectra of genetic and chromosomal aberrations. Even so, oxidative stress remains a common theme in the initiation and/or perpetuation of this phenomenon. Isolated oxidatively modified bases, abasic sites, DNA single strand breaks and clustered DNA damage are induced in normal mammalian cultured cells and tissues due to endogenous reactive oxygen species generated during normal cellular metabolism in an aerobic environment. While sparse DNA damage may be easily repaired, clustered DNA damage may lead to persistent cytotoxic or mutagenic events that can lead to genomic instability. In this study, we tested the hypothesis that DNA damage signatures characterised by altered levels of endogenous, potentially mutagenic, types of DNA damage and chromosomal breakage are related to radiation-induced genomic instability and persistent oxidative stress phenotypes observed in the chromosomally unstable progeny of irradiated cells. The measurement of oxypurine, oxypyrimidine and abasic site endogenous DNA damage showed differences in non-double-strand breaks (DSB) clusters among the three of the four unstable clones evaluated as compared to genomically stable clones and the parental cell line. These three unstable clones also had increased levels of DSB clusters. The results of this study demonstrate that each unstable cell line has a unique spectrum of persistent damage and lead us to speculate that alterations in DNA damage signaling and repair may be related to the perpetuation of genomic instability.

  20. DNA damage in cells exhibiting radiation-induced genomic instability

    DOE PAGES

    Keszenman, Deborah J.; Kolodiuk, Lucia; Baulch, Janet E.

    2015-02-22

    Cells exhibiting radiation induced genomic instability exhibit varied spectra of genetic and chromosomal aberrations. Even so, oxidative stress remains a common theme in the initiation and/or perpetuation of this phenomenon. Isolated oxidatively modified bases, abasic sites, DNA single strand breaks and clustered DNA damage are induced in normal mammalian cultured cells and tissues due to endogenous reactive oxygen species generated during normal cellular metabolism in an aerobic environment. While sparse DNA damage may be easily repaired, clustered DNA damage may lead to persistent cytotoxic or mutagenic events that can lead to genomic instability. In this study, we tested the hypothesismore » that DNA damage signatures characterised by altered levels of endogenous, potentially mutagenic, types of DNA damage and chromosomal breakage are related to radiation-induced genomic instability and persistent oxidative stress phenotypes observed in the chromosomally unstable progeny of irradiated cells. The measurement of oxypurine, oxypyrimidine and abasic site endogenous DNA damage showed differences in non-double-strand breaks (DSB) clusters among the three of the four unstable clones evaluated as compared to genomically stable clones and the parental cell line. These three unstable clones also had increased levels of DSB clusters. The results of this study demonstrate that each unstable cell line has a unique spectrum of persistent damage and lead us to speculate that alterations in DNA damage signaling and repair may be related to the perpetuation of genomic instability.« less

  1. Induction of Excess Centrosomes in Neural Progenitor Cells during the Development of Radiation-Induced Microcephaly

    PubMed Central

    Shimada, Mikio; Matsuzaki, Fumio; Kato, Akihiro; Kobayashi, Junya; Matsumoto, Tomohiro; Komatsu, Kenshi

    2016-01-01

    The embryonic brain is one of the tissues most vulnerable to ionizing radiation. In this study, we showed that ionizing radiation induces apoptosis in the neural progenitors of the mouse cerebral cortex, and that the surviving progenitor cells subsequently develop a considerable amount of supernumerary centrosomes. When mouse embryos at Day 13.5 were exposed to γ-rays, brains sizes were reduced markedly in a dose-dependent manner, and these size reductions persisted until birth. Immunostaining with caspase-3 antibodies showed that apoptosis occurred in 35% and 40% of neural progenitor cells at 4 h after exposure to 1 and 2 Gy, respectively, and this was accompanied by a disruption of the apical layer in which mitotic spindles were positioned in unirradiated mice. At 24 h after 1 Gy irradiation, the apoptotic cells were completely eliminated and proliferation was restored to a level similar to that of unirradiated cells, but numerous spindles were localized outside the apical layer. Similarly, abnormal cytokinesis, which included multipolar division and centrosome clustering, was observed in 19% and 24% of the surviving neural progenitor cells at 48 h after irradiation with 1 and 2 Gy, respectively. Because these cytokinesis aberrations derived from excess centrosomes result in growth delay and mitotic catastrophe-mediated cell elimination, our findings suggest that, in addition to apoptosis at an early stage of radiation exposure, radiation-induced centrosome overduplication could contribute to the depletion of neural progenitors and thereby lead to microcephaly. PMID:27367050

  2. Involvement of intracellular expression of FGF12 in radiation-induced apoptosis in mast cells.

    PubMed

    Nakayama, Fumiaki; Müller, Kerstin; Hagiwara, Akiko; Ridi, Roland; Akashi, Makoto; Meineke, Viktor

    2008-09-01

    Several fibroblast growth factors (FGFs) are able to reduce and improve radiation-induced tissue damage through the activation of surface fibroblast growth factor receptors (FGFRs). In contrast, some FGFs lack classical signal sequences, which play roles in the release of FGFs, and the intracellular function of these FGFs is not well clarified. In this study, we evaluated the transcript levels of 22 FGFs in a human mast cell line, HMC-1, using quantitative RT-PCR and found that FGF2 and FGF12 were expressed in HMC-1 cells. FGF12 not only lacks classical signal sequences but also fails to activate FGFRs. HMC-1 cells were transfected with an expression vector of FGF12 to clarify the intracellular function of FGF12 after irradiation. The overexpression of FGF12 in HMC-1 cells decreased ionizing radiation-induced apoptosis, and siRNA-mediated repression of FGF12 expression augmented apoptosis in HMC-1 cells. The overexpression of FGF12 strongly suppressed the marked augmentation of apoptosis induced by inhibition of the MEK/ERK pathway with PD98059. In contrast, the mitogen-activated protein kinase (MAPK) scaffold protein islet brain 2 (IB2), which was reported to bind to FGF12, did not interfere with the anti-apoptotic effect of FGF12. The expression of FGF12 transcripts was also detected in murine cultured mast cells derived from bone marrow or fetal skin. These findings suggest that FGF12 intracellularly suppresses radiation-induced apoptosis in mast cells independently of IB2.

  3. Original Research: Label-free detection for radiation-induced apoptosis in glioblastoma cells.

    PubMed

    Qi, Dandan; Feng, Jingwen; Yang, Chengwen; Jin, Changrong; Sa, Yu; Feng, Yuanming

    2016-10-01

    Current flow cytometry (FCM) requires fluorescent dyes labeling cells which make the procedure costly and time consuming. This manuscript reports a feasibility study of detecting the cell apoptosis with a label-free method in glioblastoma cells. A human glioma cell line M059K was exposed to 8 Gy dose of radiation, which enables the cells to undergo radiation-induced apoptosis. The rates of apoptosis were studied at different time points post-irradiation with two different methods: FCM in combination with Annexin V-FITC/PI staining and a newly developed technique named polarization diffraction imaging flow cytometry. Totally 1000 diffraction images were acquired for each sample and the gray level co-occurrence matrix (GLCM) algorithm was used in morphological characterization of the apoptotic cells. Among the feature parameters extracted from each image pair, we found that the two GLCM parameters of angular second moment (ASM) and sum entropy (SumEnt) exhibit high sensitivities and consistencies as the apoptotic rates (Pa) measured with FCM method. In addition, no significant difference exists between Pa and ASM_S, Pa and SumEnt_S, respectively (P > 0.05). These results demonstrated that the new label-free method can detect cell apoptosis effectively. Cells can be directly used in the subsequent biochemical experiments as the structure and function of cells and biomolecules are well-preserved with this new method.

  4. Mechanisms of radiation-induced neoplastic cell transformation

    SciTech Connect

    Yang, T.C.H.; Tobias, C.A.

    1984-04-01

    Studies with cultured mammalian cells demonstrated clearly that radiation can transform cells directly and can enhance the cell transformation by oncogenic DNA viruses. In general, high-LET heavy-ion radiation can be more effective than X and gamma rays in inducing neoplastic cell transformation. Various experimental results indicate that radiation-induced DNA damage, most likely double-strand breaks, is important for both the initiation of cell transformation and for the enhancement of viral transformation. Some of the transformation and enhancement lesions can be repaired properly in the cell, and the amount of irrepairable lesions produced by a given dose depends on the quality of radiation. An inhibition of repair processes with chemical agents can increase the transformation frequency of cells exposed to radiation and/or oncogenic viruses, suggesting that repair mechanisms may play an important role in the radiation transformation. The progression of radiation-transformed cells appears to be a long and complicated process that can be modulated by some nonmutagenic chemical agents, e.g., DMSO. Normal cells can inhibit the expression of transforming properties of tumorigenic cells through an as yet unknown mechanism. The progression and expression of transformation may involve some epigenetic changes in the irradiated cells. 38 references, 15 figures, 1 table.

  5. C-Myc regulates radiation-induced G2/M cell cycle arrest and cell death in human cervical cancer cells.

    PubMed

    Cui, Fengmei; Hou, Jun; Huang, Chengcheng; Sun, Xiujin; Zeng, Yanan; Cheng, Huiying; Wang, Hao; Li, Chao

    2017-04-01

    The study was conducted to investigate the role of c-Myc in the regulation of ionizing radiation-induced cell cycle arrest and cell death in human cervical cancer cells. Control and c-Myc-silenced Hela cells were collected at different time points after (60) Co γ-ray radiation. Flow cytometry was used to measure cell cycle distribution and apoptosis. Immunofluorescence was applied to determine the percentage of cells in M phase. Transmission electron microscopy and immunoblotting were used to detect the induction of autophagy after radiation. Immunoblotting was also used to measure the expression levels of apoptosis-related proteins. In c-Myc-silenced cells, radiation induced delayed but long-lasting G2/M arrest and an abnormal M phase compared with the control. In addition, c-Myc knockdown significantly inhibited apoptotic cell death induced by radiation. Meanwhile, radiation-induced autophagy appeared stronger in c-Myc-silenced cells. Mechanically, we found that Caspase 8 and survivin expression was decreased in c-Myc-silenced Hela-630 cells. These data showed that c-Myc serves as a co-regulator in radiation-induced G2/M cell cycle arrest and cell death in human cervical cancer cells. © 2017 Japan Society of Obstetrics and Gynecology.

  6. Radiation-induced chromosomal instability in human mammary epithelial cells

    NASA Technical Reports Server (NTRS)

    Durante, M.; Grossi, G. F.; Yang, T. C.

    1996-01-01

    Karyotypes of human cells surviving X- and alpha-irradiation have been studied. Human mammary epithelial cells of the immortal, non-tumorigenic cell line H184B5 F5-1 M/10 were irradiated and surviving clones isolated and expanded in culture. Cytogenetic analysis was performed using dedicated software with an image analyzer. We have found that both high- and low-LET radiation induced chromosomal instability in long-term cultures, but with different characteristics. Complex chromosomal rearrangements were observed after X-rays, while chromosome loss predominated after alpha-particles. Deletions were observed in both cases. In clones derived from cells exposed to alpha-particles, some cells showed extensive chromosome breaking and double minutes. Genomic instability was correlated to delayed reproductive death and neoplastic transformation. These results indicate that chromosomal instability is a radiation-quality-dependent effect which could determine late genetic effects, and should therefore be carefully considered in the evaluation of risk for space missions.

  7. Radiation-induced chromosomal instability in human mammary epithelial cells

    NASA Technical Reports Server (NTRS)

    Durante, M.; Grossi, G. F.; Yang, T. C.

    1996-01-01

    Karyotypes of human cells surviving X- and alpha-irradiation have been studied. Human mammary epithelial cells of the immortal, non-tumorigenic cell line H184B5 F5-1 M/10 were irradiated and surviving clones isolated and expanded in culture. Cytogenetic analysis was performed using dedicated software with an image analyzer. We have found that both high- and low-LET radiation induced chromosomal instability in long-term cultures, but with different characteristics. Complex chromosomal rearrangements were observed after X-rays, while chromosome loss predominated after alpha-particles. Deletions were observed in both cases. In clones derived from cells exposed to alpha-particles, some cells showed extensive chromosome breaking and double minutes. Genomic instability was correlated to delayed reproductive death and neoplastic transformation. These results indicate that chromosomal instability is a radiation-quality-dependent effect which could determine late genetic effects, and should therefore be carefully considered in the evaluation of risk for space missions.

  8. Radiation-induced chromosomal instability in human mammary epithelial cells

    NASA Astrophysics Data System (ADS)

    Durante, M.; Grossi, G. F.; Yang, T. C.

    Karyotypes of human cells surviving X- and alpha-irradiation have been studied. Human mammary epithelial cells of the immortal, non-tumorigenic cell line H184B5 F5-1 M/10 were irradiated and surviving clones isolated and expanded in culture. Cytogenetic analysis was performed using dedicated software with an image analyzer. We have found that both high- and low-LET radiation induced chromosomal instability in long-term cultures, but with different characteristics. Complex chromosomal rearrangements were observed after X-rays, while chromosome loss predominated after alpha-particles. Deletions were observed in both cases. In clones derived from cells exposed to alpha-particles, some cells showed extensive chromosome breaking and double minutes. Genomic instability was correlated to delayed reproductive death and neoplastic transformation. These results indicate that chromosomal instability is a radiation-quality-dependent effect which could determine late genetic effects, and should therefore be carefully considered in the evaluation of risk for space missions.

  9. Radiation-Induced Notch Signaling in Breast Cancer Stem Cells

    SciTech Connect

    Lagadec, Chann; Vlashi, Erina; Alhiyari, Yazeed; Phillips, Tiffany M.; Bochkur Dratver, Milana; Pajonk, Frank

    2013-11-01

    Purpose: To explore patterns of Notch receptor and ligand expression in response to radiation that could be crucial in defining optimal dosing schemes for γ-secretase inhibitors if combined with radiation. Methods and Materials: Using MCF-7 and T47D breast cancer cell lines, we used real-time reverse transcription–polymerase chain reaction to study the Notch pathway in response to radiation. Results: We show that Notch receptor and ligand expression during the first 48 hours after irradiation followed a complex radiation dose–dependent pattern and was most pronounced in mammospheres, enriched for breast cancer stem cells. Additionally, radiation activated the Notch pathway. Treatment with a γ-secretase inhibitor prevented radiation-induced Notch family gene expression and led to a significant reduction in the size of the breast cancer stem cell pool. Conclusions: Our results indicate that, if combined with radiation, γ-secretase inhibitors may prevent up-regulation of Notch receptor and ligand family members and thus reduce the number of surviving breast cancer stem cells.

  10. Cholecystokinin attenuates radiation-induced lung cancer cell apoptosis by modulating p53 gene transcription

    PubMed Central

    Han, Yi; Su, Chongyu; Yu, Daping; Zhou, Shijie; Song, Xiaoyun; Liu, Shuku; Qin, Ming; Li, Yunsong; Xiao, Ning; Cao, Xiaoqing; Shi, Kang; Cheng, Xu; Liu, Zhidong

    2017-01-01

    The deregulation of p53 in cancer cells is one of the important factors by which cancer cells escape from the immune surveillance. Cholecystokinin (CCK) has strong bioactivity in the regulation of a number of cell activities. This study tests a hypothesis that CCK interferes with p53 expression to affect the apoptotic process in lung cancer (tumor) cells. In this study, tumor-bearing mice and A549 cells (a tumor cell line) were irradiated. The expression of CCK and p53 in tumor cells was assessed with RT-qPCR and Western blotting. The binding of p300 to the promoter of p53 was evaluated by chromatin immunoprecipitation. We observed that, with a given amount and within a given period, small doses/more sessions of irradiation markedly increased the levels of CCK in the sera and tumor cells, which were positively correlated with the tumor growth in mice and negatively correlated with tumor cell apoptosis. CCK increased the levels of histone acetyltransferase p300 and repressed the levels of nuclear factor-kB at the p53 promoter locus in tumor cells, which suppressed the expression of p53. In conclusion, CCK plays an important role in attenuating the radiation-induced lung cancer cell apoptosis. CCK may be a novel therapeutic target in the treatment of lung cancers. PMID:28337291

  11. Low dose radiation-induced endothelial cell retraction.

    PubMed

    Kantak, S S; Diglio, C A; Onoda, J M

    1993-09-01

    We characterized in vitro the effects of gamma-radiation (12.5-100 cGy) on pulmonary microvascular endothelial cell (PMEC) morphology and F-actin organization. Cellular retraction was documented by phase-contrast microscopy and the organization of actin microfilaments was determined by immunofluorescence. Characterization included radiation dose effects, their temporal duration and reversibility of the effects. A dose-dependent relationship between the level of exposure (12.5-100 cGy) and the rate and extent of endothelial retraction was observed. Moreover, analysis of radiation-induced depolymerization of F-actin microfilament stress fibres correlated positively with the changes in PMEC morphology. The depolymerization of the stress fibre bundles was dependent on radiation dose and time. Cells recovered from exposure to reform contact inhibited monolayers > or = 24 h post-irradiation. Concomitantly, the depolymerized microfilaments reorganized to their preirradiated state as microfilament stress fibres arrayed parallel to the boundaries of adjacent contact-inhibited cells. The data presented here are representative of a series of studies designed to characterize low-dose radiation effects on pulmonary microvascular endothelium. Our data suggest that post-irradiation lung injuries (e.g. oedema) may be induced with only a single fraction of therapeutic radiation, and thus microscopic oedema may initiate prior to the lethal effects of radiation on the microvascular endothelium, and much earlier than would be suggested by the time course for clinically-detectable oedema.

  12. Mesenchymal stem cells stimulate intestinal stem cells to repair radiation-induced intestinal injury

    PubMed Central

    Gong, Wei; Guo, Mengzheng; Han, Zhibo; Wang, Yan; Yang, Ping; Xu, Chang; Wang, Qin; Du, Liqing; Li, Qian; Zhao, Hui; Fan, Feiyue; Liu, Qiang

    2016-01-01

    The loss of stem cells residing in the base of the intestinal crypt has a key role in radiation-induced intestinal injury. In particular, Lgr5+ intestinal stem cells (ISCs) are indispensable for intestinal regeneration following exposure to radiation. Mesenchymal stem cells (MSCs) have previously been shown to improve intestinal epithelial repair in a mouse model of radiation injury, and, therefore, it was hypothesized that this protective effect is related to Lgr5+ ISCs. In this study, it was found that, following exposure to radiation, transplantation of MSCs improved the survival of the mice, ameliorated intestinal injury and increased the number of regenerating crypts. Furthermore, there was a significant increase in Lgr5+ ISCs and their daughter cells, including Ki67+ transient amplifying cells, Vil1+ enterocytes and lysozyme+ Paneth cells, in response to treatment with MSCs. Crypts isolated from mice treated with MSCs formed a higher number of and larger enteroids than those from the PBS group. MSC transplantation also reduced the number of apoptotic cells within the small intestine at 6 h post-radiation. Interestingly, Wnt3a and active β-catenin protein levels were increased in the small intestines of MSC-treated mice. In addition, intravenous delivery of recombinant mouse Wnt3a after radiation reduced damage in the small intestine and was radioprotective, although not to the same degree as MSC treatment. Our results show that MSCs support the growth of endogenous Lgr5+ ISCs, thus promoting repair of the small intestine following exposure to radiation. The molecular mechanism of action mediating this was found to be related to increased activation of the Wnt/β-catenin signaling pathway. PMID:27685631

  13. Autophagy promotes radiation-induced senescence but inhibits bystander effects in human breast cancer cells.

    PubMed

    Huang, Yao-Huei; Yang, Pei-Ming; Chuah, Qiu-Yu; Lee, Yi-Jang; Hsieh, Yi-Fen; Peng, Chih-Wen; Chiu, Shu-Jun

    2014-07-01

    Ionizing radiation induces cellular senescence to suppress cancer cell proliferation. However, it also induces deleterious bystander effects in the unirradiated neighboring cells through the release of senescence-associated secretory phenotypes (SASPs) that promote tumor progression. Although autophagy has been reported to promote senescence, its role is still unclear. We previously showed that radiation induces senescence in PTTG1-depleted cancer cells. In this study, we found that autophagy was required for the radiation-induced senescence in PTTG1-depleted breast cancer cells. Inhibition of autophagy caused the cells to switch from radiation-induced senescence to apoptosis. Senescent cancer cells exerted bystander effects by promoting the invasion and migration of unirradiated cells through the release of CSF2 and the subsequently activation of the JAK2-STAT3 and AKT pathways. However, the radiation-induced bystander effects were correlated with the inhibition of endogenous autophagy in bystander cells, which also resulted from the activation of the CSF2-JAK2 pathway. The induction of autophagy by rapamycin reduced the radiation-induced bystander effects. This study reveals, for the first time, the dual role of autophagy in radiation-induced senescence and bystander effects.

  14. Rhubarb extract has a protective role against radiation-induced brain injury and neuronal cell apoptosis.

    PubMed

    Lu, Kui; Zhang, Cheng; Wu, Wenjun; Zhou, Min; Tang, Yamei; Peng, Ying

    2015-08-01

    Oxidative stress caused by ionizing radiation is involved in neuronal damage in a number of disorders, including trauma, stroke, Alzheimer's disease and amyotrophic lateral sclerosis. Ionizing radiation can lead to the formation of free radicals, which cause neuronal apoptosis and have important roles in the development of some types of chronic brain disease. The present study evaluated the effects of varying concentrations (2, 5 and 10 µg/ml) of ethanolic rhubarb extract on the neuronal damage caused by irradiation in primary neuronal cultures obtained from the cortices of rat embryos aged 20 days. Brain damage was induced with a single dose of γ-irradiation that induced DNA fragmentation, increased lactate dehydrogenase release in neuronal cells and acted as a trigger for microglial cell proliferation. Treatment with rhubarb extract significantly decreased radiation-induced lactate dehydrogenase release and DNA fragmentation, which are important in the process of cell apoptosis. The rhubarb extract exhibited dose-dependent inhibition of lactate dehydrogenase release and neuronal cell apoptosis that were induced by the administration of ionizing radiation. The effect of a 10 µg/ml dose of rhubarb extract on the generation of reactive oxygen species (ROS) induced by radiation was also investigated. This dose led to significant inhibition of ROS generation. In conclusion, the present study showed a protective role of rhubarb extract against irradiation-induced apoptotic neuronal cell death and ROS generation.

  15. Radiation induces autophagic cell death via the p53/DRAM signaling pathway in breast cancer cells.

    PubMed

    Cui, Li; Song, Zhiheng; Liang, Bing; Jia, Lili; Ma, Shumei; Liu, Xiaodong

    2016-06-01

    Autophagy is known to play a role in the response of breast cancer cells to radiation therapy. However, the mechanisms that mediate the process of autophagy and contribute to radiation-induced cell death and cell survival remain to be fully characterized. Therefore, in this study, the functional role of autophagy in radiation-induced cytotoxicity in breast cancer cells was investigated. After MCF-7 cells were exposed to various doses of radiation, increased monodansylcadaverine (MDC) staining and a greater deposition of LC3-positive puncta were observed. Expression of the autophagy-related proteins, Beclin 1 and LC3-II, were also found to be upregulated. Radiation-induced autophagic cell death was partially abrogated following the administration of 3-methyladenine (3-MA) and in knockdown experiments of Atg5 and Beclin 1. In the gene microarray analysis performed after irradiation, a number of differentially expressed genes were identified. In particular, upregulation of both the mRNA and protein levels of the autophagy-related genes, DRAM and TIGAR, were detected. However, inhibition of autophagy by 3-MA reduced the radiation-induced upregulation of LC3-II and DRAM. Conversely, silencing of p53 downregulated the expression of LC3-II and DRAM following radiation. Silencing of DRAM reversed the upregulation of LC3-II and DRAM following radiation, partially blocked radiation-induced cell death, and no significant change in p53 expression was detected. Based on these results, the p53/DRAM signaling pathway appears to contribute to radiation-induced autophagic cell death in MCF-7 breast cancer cells.

  16. Gene silencing of Tead3 abrogates radiation-induced adaptive response in cultured mouse limb bud cells.

    PubMed

    Vares, Guillaume; Wang, Bing; Tanaka, Kaoru; Shang, Yi; Taki, Keiko; Nakajima, Tetsuo; Nenoi, Mitsuru

    2011-01-01

    There is a crucial need to better understand the effects of low-doses of ionizing radiation in fetal models. Radiation-induced adaptive response (AR) was described in mouse embryos pre-exposed in utero to low-doses of X-rays, which exhibited lower apoptotic levels in the limb bud. We previously described AR-specific gene modulations in the mouse embryo. In this study, we evaluated the role of three candidate genes in the apoptotic AR in a micromass culture of limb bud cells: Csf1, Cacna1a and Tead3. Gene silencing of these three genes abrogated AR. Knowing that TEAD3 protein levels are significantly higher in adapted cells and that YAP/TAZ/TEAD are involved in the control of cell proliferation and apoptosis, we suggest that modulation of Tead3 could play a role in the induction of AR in our model, seen as a reduction of radiation-induced apoptosis and a stimulation of proliferation and differentiation in limb bud cells.

  17. Apoptotic cells enhance pathogenesis of Listeria monocytogenes.

    PubMed

    Pattabiraman, Goutham; Palasiewicz, Karol; Visvabharathy, Lavanya; Freitag, Nancy E; Ucker, David S

    2017-04-01

    Infections by pathogenic microorganisms elicit host immune responses, which crucially limit those infections. Pathogens employ various strategies to evade host immunity. We have identified the exploitation of the repertoire of potent immunosuppressive responses elicited normally by apoptotic cells ("Innate Apoptotic Immunity"; IAI) as one of these strategies. In the case of Listeria monocytogenes, an environmentally ubiquitous, foodborne bacterial pathogen capable of causing life-threatening invasive disease in immunocompromised and elderly individuals, the induction of host cell apoptosis appears to play an important role in pathogenesis. Previous studies have documented extensive lymphocyte apoptosis resulting from L. monocytogenes infection and demonstrated paradoxically that lymphocyte-deficient animals exhibit diminished susceptibility to listerial pathogenicity. We speculated that the triggering of IAI following the induction of host cell apoptosis was responsible for enhanced pathogenesis, and that the administration of exogenous apoptotic cells would serve to exert this effect. Importantly, apoptotic cells, which are not susceptible to L. monocytogenes infection, do not provide a niche for bacterial replication. Our experiments confirm that apoptotic cells, including exogenous apoptotic cells induced to die independently of the pathogen, specifically enhance pathogenesis. The recognition of a role of apoptotic cells and Innate Apoptotic Immunity in microbial pathogenesis provides an intriguing and novel insight for therapeutic approaches for the control of pathogenic infections.

  18. Radiation-induced glioblastoma signaling cascade regulates viability, apoptosis and differentiation of neural stem cells (NSC).

    PubMed

    Ivanov, Vladimir N; Hei, Tom K

    2014-12-01

    Ionizing radiation alone or in combination with chemotherapy is the main treatment modality for brain tumors including glioblastoma. Adult neurons and astrocytes demonstrate substantial radioresistance; in contrast, human neural stem cells (NSC) are highly sensitive to radiation via induction of apoptosis. Irradiation of tumor cells has the potential risk of affecting the viability and function of NSC. In this study, we have evaluated the effects of irradiated glioblastoma cells on viability, proliferation and differentiation potential of non-irradiated (bystander) NSC through radiation-induced signaling cascades. Using media transfer experiments, we demonstrated significant effects of the U87MG glioblastoma secretome after gamma-irradiation on apoptosis in non-irradiated NSC. Addition of anti-TRAIL antibody to the transferred media partially suppressed apoptosis in NSC. Furthermore, we observed a dramatic increase in the production and secretion of IL8, TGFβ1 and IL6 by irradiated glioblastoma cells, which could promote glioblastoma cell survival and modify the effects of death factors in bystander NSC. While differentiation of NSC into neurons and astrocytes occurred efficiently with the corresponding differentiation media, pretreatment of NSC for 8 h with medium from irradiated glioblastoma cells selectively suppressed the differentiation of NSC into neurons, but not into astrocytes. Exogenous IL8 and TGFβ1 increased NSC/NPC survival, but also suppressed neuronal differentiation. On the other hand, IL6 was known to positively affect survival and differentiation of astrocyte progenitors. We established a U87MG neurosphere culture that was substantially enriched by SOX2(+) and CD133(+) glioma stem-like cells (GSC). Gamma-irradiation up-regulated apoptotic death in GSC via the FasL/Fas pathway. Media transfer experiments from irradiated GSC to non-targeted NSC again demonstrated induction of apoptosis and suppression of neuronal differentiation of NSC. In

  19. Radiation-induced glioblastoma signaling cascade regulates viability, apoptosis and differentiation of neural stem cells (NSC)

    PubMed Central

    Ivanov, Vladimir N.; Hei, Tom K.

    2015-01-01

    Ionizing radiation alone or in combination with chemotherapy is the main treatment modality for brain tumors including glioblastoma. Adult neurons and astrocytes demonstrate substantial radioresistance; in contrast, human neural stem cells (NSC) are highly sensitive to radiation via induction of apoptosis. Irradiation of tumor cells has the potential risk of affecting the viability and function of NSC. In this study, we have evaluated the effects of irradiated glioblastoma cells on viability, proliferation and differentiation potential of non-irradiated (bystander) NSC through radiation-induced signaling cascades. Using media transfer experiments, we demonstrated significant effects of the U87MG glioblastoma secretome after gamma-irradiation on apoptosis in non-irradiated NSC. Addition of anti-TRAIL antibody to the transferred media partially suppressed apoptosis in NSC. Furthermore, we observed a dramatic increase in the production and secretion of IL8, TGFβ1 and IL6 by irradiated glioblastoma cells, which could promote glioblastoma cell survival and modify the effects of death factors in bystander NSC. While differentiation of NSC into neurons and astrocytes occurred efficiently with the corresponding differentiation media, pretreatment of NSC for 8 h with medium from irradiated glioblastoma cells selectively suppressed the differentiation of NSC into neurons, but not into astrocytes. Exogenous IL8 and TGFβ1 increased NSC/NPC survival, but also suppressed neuronal differentiation. On the other hand, IL6 was known to positively affect survival and differentiation of astrocyte progenitors. We established a U87MG neurosphere culture that was substantially enriched by SOX2+ and CD133+ glioma stem-like cells (GSC). Gamma-irradiation up-regulated apoptotic death in GSC via the FasL/Fas pathway. Media transfer experiments from irradiated GSC to non-targeted NSC again demonstrated induction of apoptosis and suppression of neuronal differentiation of NSC. In summary

  20. The influence of Trp53 in the dose response of radiation-induced apoptosis, DNA repair and genomic stability in murine haematopoietic cells.

    PubMed

    Lemon, Jennifer A; Taylor, Kristina; Verdecchia, Kyle; Phan, Nghi; Boreham, Douglas R

    2014-07-01

    Apoptotic and DNA damage endpoints are frequently used as surrogate markers of cancer risk, and have been well-studied in the Trp53+/- mouse model. We report the effect of differing Trp53 gene status on the dose response of ionizing radiation exposures (0.01-2 Gy), with the unique perspective of determining if effects of gene status remain at extended time points. Here we report no difference in the dose response for radiation-induced DNA double-strand breaks in bone marrow and genomic instability (MN-RET levels) in peripheral blood, between wild-type (Trp53+/+) and heterozygous (Trp53+/-) mice. The dose response for Trp53+/+ mice showed higher initial levels of radiation-induced lymphocyte apoptosis relative to Trp53+/- between 0 and 1 Gy. Although this trend was observed up to 12 hours post-irradiation, both genotypes ultimately reached the same level of apoptosis at 14 hours, suggesting the importance of late-onset p53-independent apoptotic responses in this mouse model. Expected radiation-induced G1 cell cycle delay was observed in Trp53+/+ but not Trp53+/-. Although p53 has an important role in cancer risk, we have shown its influence on radiation dose response can be temporally variable. This research highlights the importance of caution when using haematopoietic endpoints as surrogates to extrapolate radiation-induced cancer risk estimation.

  1. The influence of TRP53 in the dose response of radiation-induced apoptosis, DNA repair and genomic stability in murine haematopoietic cells

    DOE PAGES

    Lemon, Jennifer A.; Taylor, Kristina; Verdecchia, Kyle; ...

    2014-01-01

    Apoptotic and DNA damage endpoints are frequently used as surrogate markers of cancer risk, and have been well-studied in the Trp53+/- mouse model. We report the effect of differing Trp53 gene status on the dose response of ionizing radiation exposures (0.01-2 Gy), with the unique perspective of determining if effects of gene status remain at extended time points. Here we report no difference in the dose response for radiation-induced DNA double-strand breaks in bone marrow and genomic instability (MN-RET levels) in peripheral blood, between wild-type (Trp53+/+) and heterozygous (Trp53+/-) mice. The dose response for Trp53+/+ mice showed higher initial levelsmore » of radiation-induced lymphocyte apoptosis relative to Trp53+/- between 0 and 1 Gy. Although this trend was observed up to 12 hours post-irradiation, both genotypes ultimately reached the same level of apoptosis at 14 hours, suggesting the importance of late-onset p53-independent apoptotic responses in this mouse model. Expected radiation-induced G1 cell cycle delay was observed in Trp53+/+ but not Trp53+/-. Although p53 has an important role in cancer risk, we have shown its influence on radiation dose response can be temporally variable. This research highlights the importance of caution when using haematopoietic endpoints as surrogates to extrapolate radiation-induced cancer risk estimation.« less

  2. Stabilization of apoptotic cells: generation of zombie cells.

    PubMed

    Oropesa-Ávila, M; Andrade-Talavera, Y; Garrido-Maraver, J; Cordero, M D; de la Mata, M; Cotán, D; Paz, M V; Pavón, A D; Alcocer-Gómez, E; de Lavera, I; Lema, R; Zaderenko, A P; Rodríguez-Moreno, A; Sánchez-Alcázar, J A

    2014-08-14

    Apoptosis is characterized by degradation of cell components but plasma membrane remains intact. Apoptotic microtubule network (AMN) is organized during apoptosis forming a cortical structure beneath plasma membrane that maintains plasma membrane integrity. Apoptotic cells are also characterized by high reactive oxygen species (ROS) production that can be potentially harmful for the cell. The aim of this study was to develop a method that allows stabilizing apoptotic cells for diagnostic and therapeutic applications. By using a cocktail composed of taxol (a microtubule stabilizer), Zn(2+) (a caspase inhibitor) and coenzyme Q10 (a lipid antioxidant), we were able to stabilize H460 apoptotic cells in cell cultures for at least 72 h, preventing secondary necrosis. Stabilized apoptotic cells maintain many apoptotic cell characteristics such as the presence of apoptotic microtubules, plasma membrane integrity, low intracellular calcium levels and mitochondrial polarization. Apoptotic cell stabilization may open new avenues in apoptosis detection and therapy.

  3. Stabilization of apoptotic cells: generation of zombie cells

    PubMed Central

    Oropesa-Ávila, M; Andrade-Talavera, Y; Garrido-Maraver, J; Cordero, M D; de la Mata, M; Cotán, D; Paz, M V; Pavón, A D; Alcocer-Gómez, E; de Lavera, I; Lema, R; Zaderenko, A P; Rodríguez-Moreno, A; Sánchez-Alcázar, J A

    2014-01-01

    Apoptosis is characterized by degradation of cell components but plasma membrane remains intact. Apoptotic microtubule network (AMN) is organized during apoptosis forming a cortical structure beneath plasma membrane that maintains plasma membrane integrity. Apoptotic cells are also characterized by high reactive oxygen species (ROS) production that can be potentially harmful for the cell. The aim of this study was to develop a method that allows stabilizing apoptotic cells for diagnostic and therapeutic applications. By using a cocktail composed of taxol (a microtubule stabilizer), Zn2+ (a caspase inhibitor) and coenzyme Q10 (a lipid antioxidant), we were able to stabilize H460 apoptotic cells in cell cultures for at least 72 h, preventing secondary necrosis. Stabilized apoptotic cells maintain many apoptotic cell characteristics such as the presence of apoptotic microtubules, plasma membrane integrity, low intracellular calcium levels and mitochondrial polarization. Apoptotic cell stabilization may open new avenues in apoptosis detection and therapy. PMID:25118929

  4. cAMP signaling inhibits radiation-induced ATM phosphorylation leading to the augmentation of apoptosis in human lung cancer cells.

    PubMed

    Cho, Eun-Ah; Kim, Eui-Jun; Kwak, Sahng-June; Juhnn, Yong-Sung

    2014-02-24

    The ataxia-telangiectasia mutated (ATM) protein kinase plays a central role in coordinating the cellular response to radiation-induced DNA damage. cAMP signaling regulates various cellular responses including metabolism and gene expression. This study aimed to investigate the mechanism through which cAMP signaling regulates ATM activation and cellular responses to ionizing radiation in lung cancer cells. Lung cancer cells were transfected with constitutively active stimulatory G protein (GαsQL), and irradiated with γ-rays. The phosphorylation of ATM and protein phosphatase 2A was analyzed by western blotting, and apoptosis was assessed by western blotting, flow cytometry, and TUNNEL staining. The promoter activity of NF-κB was determined by dual luciferase reporter assay. BALB/c mice were treated with forskolin to assess the effect in the lung tissue. Transient expression of GαsQL significantly inhibited radiation-induced ATM phosphorylation in H1299 human lung cancer cells. Treatment with okadaic acid or knock down of PP2A B56δ subunit abolished the inhibitory effect of Gαs on radiation-induced ATM phosphorylation. Expression of GαsQL increased phosphorylation of the B56δ and PP2A activity, and inhibition of PKA blocked Gαs-induced PP2A activation. GαsQL enhanced radiation-induced cleavage of caspase-3 and PARP and increased the number of early apoptotic cells. The radiation-induced apoptosis was increased by inhibition of NF-κB using PDTC or inhibition of ATM using KU55933 or siRNA against ATM. Pretreatment of BALB/c mice with forskolin stimulated phosphorylation of PP2A B56δ, inhibited the activation of ATM and NF-κB, and augmented radiation-induced apoptosis in the lung tissue. GαsQL expression decreased the nuclear levels of the p50 and p65 subunits and NF-κB-dependent activity after γ-ray irradiation in H1299 cells. Pretreatment with prostaglandin E2 or isoproterenol increased B56δ phosphorylation, decreased radiation-induced ATM

  5. Apoptotic cell death and efferocytosis in atherosclerosis.

    PubMed

    Van Vré, Emily A; Ait-Oufella, Hafid; Tedgui, Alain; Mallat, Ziad

    2012-04-01

    Apoptotic cell death is an important feature of atherosclerotic plaques, and it seems to exert both beneficial and detrimental effects depending on the cell type and plaque stage. Because late apoptotic cells can launch proatherogenic inflammatory responses, adequate engulfment of apoptotic cells (efferocytosis) by macrophages is important to withstand atherosclerosis progression. Several efferocytosis systems, composed of different phagocytic receptors, apoptotic ligands, and bridging molecules, can be distinguished. Because phagocytes in atherosclerotic plaques are very much solicited, a fully operative efferocytosis system seems to be an absolute requisite. Indeed, recent studies demonstrate that deletion of just 1 of the efferocytosis pathways aggravates atherosclerosis. This review discusses the role of apoptosis in atherosclerosis and general mechanisms of efferocytosis, to end with indirect and direct indications of the significance of effective efferocytosis in atherosclerosis.

  6. Chemical chaperones reduce ionizing radiation-induced endoplasmic reticulum stress and cell death in IEC-6 cells.

    PubMed

    Lee, Eun Sang; Lee, Hae-June; Lee, Yoon-Jin; Jeong, Jae-Hoon; Kang, Seongman; Lim, Young-Bin

    2014-07-25

    Radiotherapy, which is one of the most effective approaches to the treatment of various cancers, plays an important role in malignant cell eradication in the pelvic area and abdomen. However, it also generates some degree of intestinal injury. Apoptosis in the intestinal epithelium is the primary pathological factor that initiates radiation-induced intestinal injury, but the mechanism by which ionizing radiation (IR) induces apoptosis in the intestinal epithelium is not clearly understood. Recently, IR has been shown to induce endoplasmic reticulum (ER) stress, thereby activating the unfolded protein response (UPR) signaling pathway in intestinal epithelial cells. However, the consequences of the IR-induced activation of the UPR signaling pathway on radiosensitivity in intestinal epithelial cells remain to be determined. In this study, we investigated the role of ER stress responses in IR-induced intestinal epithelial cell death. We show that chemical ER stress inducers, such as tunicamycin or thapsigargin, enhanced IR-induced caspase 3 activation and DNA fragmentation in intestinal epithelial cells. Knockdown of Xbp1 or Atf6 with small interfering RNA inhibited IR-induced caspase 3 activation. Treatment with chemical chaperones prevented ER stress and subsequent apoptosis in IR-exposed intestinal epithelial cells. Our results suggest a pro-apoptotic role of ER stress in IR-exposed intestinal epithelial cells. Furthermore, inhibiting ER stress may be an effective strategy to prevent IR-induced intestinal injury.

  7. Stabilization Of Apoptotic Cells: Generation Of Zombie Cells.

    PubMed

    Sánchez Alcázar, José A; Oropesa Ávila, Manuel; Andrade Talavera, Yuniesky; Garrido Maraver, Juan; de Lavera, Isabel; de la Mata, Mario; Cotán, David; Villanueva Paz, Marina; Delgado Pavón, Ana; Alcocer Gómez, Elisabet; Rodríguez Moreno, Antonio

    2015-08-01

    Apoptosis is characterized by degradation of cell components but plasma membrane remains intact. Apoptotic microtubule network (AMN) is organized during apoptosis forming a cortical structure beneath plasma membrane that maintains plasma membrane integrity. Apoptotic cells are also characterized by high reactive oxygen species (ROS) production that can be potentially harmful for the cell. The aim of this study was to develop a method that allows stabilizing apoptotic cells for diagnostic and therapeutic applications. We were able by using a cocktail composed of taxol (a microtubule stabilizer), Zn(2+) (a caspase inhibitor) and coenzyme Q10 (a lipid antioxidant) to stabilize H460 apoptotic cells in cell cultures for at least 72hours preventing secondary necrosis. Stabilized apoptotic cells maintain many apoptotic cells characteristics such as the presence of apoptotic microtubules, plasma membrane integrity, low intracellular calcium levels, plasma membrane potential, PS externalization and ability of being phagocytosed. Stabilized apoptotic cells can be considered as dying cells in which the cellular cortex and plasma membrane are maintained intact or alive. In a metaphorical sense, we can consider them as "living dead" or "zombie cells". Stabilization of apoptotic cells can be used for reliable detection and quantification of apoptosis in cultured cells and may allow a safer administration of apoptotic cells in clinical applications. Furthermore, it opens new avenues in the functional reconstruction of apoptotic cells for longer preservation.

  8. Detection of apoptotic cells using immunohistochemistry.

    PubMed

    Newbold, Andrea; Martin, Ben P; Cullinane, Carleen; Bots, Michael

    2014-11-03

    Immunohistochemistry is commonly used to show the presence of apoptotic cells in situ. In this protocol, B-cell lymphoma cells are injected into recipient mice and, on tumor formation, the mice are treated with the apoptosis inducer vorinostat (a histone deacetylase inhibitor). Tumor samples are fixed and sectioned, and fragmented DNA (a feature of apoptotic cells) is end-labeled by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL). Immunohistochemical methods are then used to detect the labeled DNA and identify B-cell lymphoma cells in the last stage of apoptosis. Because the assay can lead to false-positive results, it is advisable to carry out an additional assay (e.g., immunohistochemistry for active caspase-3) to confirm the presence of apoptotic cells.

  9. RhoA GTPase regulates radiation-induced alterations in endothelial cell adhesion and migration

    SciTech Connect

    Rousseau, Matthieu; Gaugler, Marie-Helene; Rodallec, Audrey; Bonnaud, Stephanie; Paris, Francois; Corre, Isabelle

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer We explore the role of RhoA in endothelial cell response to ionizing radiation. Black-Right-Pointing-Pointer RhoA is rapidly activated by single high-dose of radiation. Black-Right-Pointing-Pointer Radiation leads to RhoA/ROCK-dependent actin cytoskeleton remodeling. Black-Right-Pointing-Pointer Radiation-induced apoptosis does not require the RhoA/ROCK pathway. Black-Right-Pointing-Pointer Radiation-induced alteration of endothelial adhesion and migration requires RhoA/ROCK. -- Abstract: Endothelial cells of the microvasculature are major target of ionizing radiation, responsible of the radiation-induced vascular early dysfunctions. Molecular signaling pathways involved in endothelial responses to ionizing radiation, despite being increasingly investigated, still need precise characterization. Small GTPase RhoA and its effector ROCK are crucial signaling molecules involved in many endothelial cellular functions. Recent studies identified implication of RhoA/ROCK in radiation-induced increase in endothelial permeability but other endothelial functions altered by radiation might also require RhoA proteins. Human microvascular endothelial cells HMEC-1, either treated with Y-27632 (inhibitor of ROCK) or invalidated for RhoA by RNA interference were exposed to 15 Gy. We showed a rapid radiation-induced activation of RhoA, leading to a deep reorganisation of actin cytoskeleton with rapid formation of stress fibers. Endothelial early apoptosis induced by ionizing radiation was not affected by Y-27632 pre-treatment or RhoA depletion. Endothelial adhesion to fibronectin and formation of focal adhesions increased in response to radiation in a RhoA/ROCK-dependent manner. Consistent with its pro-adhesive role, ionizing radiation also decreased endothelial cells migration and RhoA was required for this inhibition. These results highlight the role of RhoA GTPase in ionizing radiation-induced deregulation of essential endothelial

  10. Early radiation-induced endothelial cell loss and blood-spinal cord barrier breakdown in the rat spinal cord.

    PubMed

    Li, Yu-Qing; Chen, Paul; Jain, Vipan; Reilly, Raymond M; Wong, C Shun

    2004-02-01

    Using a rat spinal cord model, this study was designed to characterize radiation-induced vascular endothelial cell loss and its relationship to early blood-brain barrier disruption in the central nervous system. Adult rats were given a single dose of 0, 2, 8, 19.5, 22, 30 or 50 Gy to the cervical spinal cord. At various times up to 2 weeks after irradiation, the spinal cord was processed for histological and immunohistochemical analysis. Radiation-induced apoptosis was assessed by morphology and TdT-mediated dUTP nick end labeling combined with immunohistochemical markers for endothelial and glial cells. Image analysis was performed to determine endothelial cell and microvessel density using immunohistochemistry with endothelial markers, namely endothelial barrier antigen, glucose transporter isoform 1, laminin and zonula occludens 1. Blood-spinal cord barrier permeability was assessed using immunohistochemistry for albumin and (99m)Tc-diethylenetriamine pentaacetic acid as a vascular tracer. Endothelial cell proliferation was assessed using in vivo BrdU labeling. During the first 24 h after irradiation, apoptotic endothelial cells were observed in the rat spinal cord. The decrease in endothelial cell density at 24 h after irradiation was associated with an increase in albumin immunostaining around microvessels. The decrease in the number of endothelial cells persisted for 7 days and recovery of endothelial density was apparent by day 14. A similar pattern of blood-spinal cord barrier disruption and recovery of permeability was observed over the 2 weeks, and an increase in BrdU-labeled endothelial cells was seen at day 3. These results are consistent with an association between endothelial cell death and acute blood-spinal cord barrier disruption in the rat spinal cord after irradiation.

  11. Increase of RhoB in gamma-radiation-induced apoptosis is regulated by c-Jun N-terminal kinase in Jurkat T cells.

    PubMed

    Kim, Chun-Ho; Won, Misun; Choi, Chung-Hae; Ahn, Jiwon; Kim, Bo-Kyung; Song, Kyung-Bin; Kang, Chang-Mo; Chung, Kyung-Sook

    2010-01-08

    The Ras-related small GTP-binding protein RhoB is known to be a pro-apoptotic protein and immediate-early inducible by genotoxic stresses. In addition, JNK activation is known to function in gamma-radiation-induced apoptosis. However, it is unclear how JNK activation and gamma-radiation-dependent RhoB induction are related. Here we verified the relationship between JNK activation and RhoB induction. RhoB induction by gamma-radiation occurred at the transcriptional level and transcriptional activation of RhoB was concomitant with an increase in RhoB protein. gamma-Radiation-induced RhoB expression was markedly attenuated by pretreatment with a JNK-specific inhibitor, SP600125, but not by a p38 MAPK inhibitor, SB203580. Inhibition of JNK caused a decrease in early apoptotic cell death that correlated with RhoB expression. However, PI3K inhibition had no significant effects, indicating that the AKT survival pathway was not involved. The siRNA knockdown of JNK resulted in a decrease in RhoB expression and the siRNA knockdown of RhoB restored cell growth even in the gamma-irradiated cells. These results suggest that RhoB regulation involves the JNK pathway and contributes to the early apoptotic response of Jurkat T cells to gamma-radiation. Copyright 2009 Elsevier Inc. All rights reserved.

  12. Chemical chaperones reduce ionizing radiation-induced endoplasmic reticulum stress and cell death in IEC-6 cells

    SciTech Connect

    Lee, Eun Sang; Lee, Hae-June; Lee, Yoon-Jin; Jeong, Jae-Hoon; Kang, Seongman; Lim, Young-Bin

    2014-07-25

    Highlights: • UPR activation precedes caspase activation in irradiated IEC-6 cells. • Chemical ER stress inducers radiosensitize IEC-6 cells. • siRNAs that targeted ER stress responses ameliorate IR-induced cell death. • Chemical chaperons prevent cell death in irradiated IEC-6 cells. - Abstract: Radiotherapy, which is one of the most effective approaches to the treatment of various cancers, plays an important role in malignant cell eradication in the pelvic area and abdomen. However, it also generates some degree of intestinal injury. Apoptosis in the intestinal epithelium is the primary pathological factor that initiates radiation-induced intestinal injury, but the mechanism by which ionizing radiation (IR) induces apoptosis in the intestinal epithelium is not clearly understood. Recently, IR has been shown to induce endoplasmic reticulum (ER) stress, thereby activating the unfolded protein response (UPR) signaling pathway in intestinal epithelial cells. However, the consequences of the IR-induced activation of the UPR signaling pathway on radiosensitivity in intestinal epithelial cells remain to be determined. In this study, we investigated the role of ER stress responses in IR-induced intestinal epithelial cell death. We show that chemical ER stress inducers, such as tunicamycin or thapsigargin, enhanced IR-induced caspase 3 activation and DNA fragmentation in intestinal epithelial cells. Knockdown of Xbp1 or Atf6 with small interfering RNA inhibited IR-induced caspase 3 activation. Treatment with chemical chaperones prevented ER stress and subsequent apoptosis in IR-exposed intestinal epithelial cells. Our results suggest a pro-apoptotic role of ER stress in IR-exposed intestinal epithelial cells. Furthermore, inhibiting ER stress may be an effective strategy to prevent IR-induced intestinal injury.

  13. Mesenchymal Stem Cell Therapy Protects Lungs from Radiation-Induced Endothelial Cell Loss by Restoring Superoxide Dismutase 1 Expression.

    PubMed

    Klein, Diana; Steens, Jennifer; Wiesemann, Alina; Schulz, Florian; Kaschani, Farnusch; Röck, Katharina; Yamaguchi, Masahiro; Wirsdörfer, Florian; Kaiser, Markus; Fischer, Jens W; Stuschke, Martin; Jendrossek, Verena

    2017-04-10

    Radiation-induced normal tissue toxicity is closely linked to endothelial cell (EC) damage and dysfunction (acute effects). However, the underlying mechanisms of radiation-induced adverse late effects with respect to the vascular compartment remain elusive, and no causative radioprotective treatment is available to date. The importance of injury to EC for radiation-induced late toxicity in lungs after whole thorax irradiation (WTI) was investigated using a mouse model of radiation-induced pneumopathy. We show that WTI induces EC loss as long-term complication, which is accompanied by the development of fibrosis. Adoptive transfer of mesenchymal stem cells (MSCs) either derived from bone marrow or aorta (vascular wall-resident MSCs) in the early phase after irradiation limited the radiation-induced EC loss and fibrosis progression. Furthermore, MSC-derived culture supernatants rescued the radiation-induced reduction in viability and long-term survival of cultured lung EC. We further identified the antioxidant enzyme superoxide dismutase 1 (SOD1) as a MSC-secreted factor. Importantly, MSC treatment restored the radiation-induced reduction of SOD1 levels after WTI. A similar protective effect was achieved by using the SOD-mimetic EUK134, suggesting that MSC-derived SOD1 is involved in the protective action of MSC, presumably through paracrine signaling. In this study, we explored the therapeutic potential of MSC therapy to prevent radiation-induced EC loss (late effect) and identified the protective mechanisms of MSC action. Adoptive transfer of MSCs early after irradiation counteracts radiation-induced vascular damage and EC loss as late adverse effects. The high activity of vascular wall-derived MSCs for radioprotection may be due to their tissue-specific action. Antioxid. Redox Signal. 26, 563-582.

  14. Mesenchymal Stem Cell Therapy Protects Lungs from Radiation-Induced Endothelial Cell Loss by Restoring Superoxide Dismutase 1 Expression

    PubMed Central

    Steens, Jennifer; Wiesemann, Alina; Schulz, Florian; Kaschani, Farnusch; Röck, Katharina; Yamaguchi, Masahiro; Wirsdörfer, Florian; Kaiser, Markus; Fischer, Jens W.; Stuschke, Martin; Jendrossek, Verena

    2017-01-01

    Abstract Aims: Radiation-induced normal tissue toxicity is closely linked to endothelial cell (EC) damage and dysfunction (acute effects). However, the underlying mechanisms of radiation-induced adverse late effects with respect to the vascular compartment remain elusive, and no causative radioprotective treatment is available to date. Results: The importance of injury to EC for radiation-induced late toxicity in lungs after whole thorax irradiation (WTI) was investigated using a mouse model of radiation-induced pneumopathy. We show that WTI induces EC loss as long-term complication, which is accompanied by the development of fibrosis. Adoptive transfer of mesenchymal stem cells (MSCs) either derived from bone marrow or aorta (vascular wall-resident MSCs) in the early phase after irradiation limited the radiation-induced EC loss and fibrosis progression. Furthermore, MSC-derived culture supernatants rescued the radiation-induced reduction in viability and long-term survival of cultured lung EC. We further identified the antioxidant enzyme superoxide dismutase 1 (SOD1) as a MSC-secreted factor. Importantly, MSC treatment restored the radiation-induced reduction of SOD1 levels after WTI. A similar protective effect was achieved by using the SOD-mimetic EUK134, suggesting that MSC-derived SOD1 is involved in the protective action of MSC, presumably through paracrine signaling. Innovation: In this study, we explored the therapeutic potential of MSC therapy to prevent radiation-induced EC loss (late effect) and identified the protective mechanisms of MSC action. Conclusions: Adoptive transfer of MSCs early after irradiation counteracts radiation-induced vascular damage and EC loss as late adverse effects. The high activity of vascular wall-derived MSCs for radioprotection may be due to their tissue-specific action. Antioxid. Redox Signal. 26, 563–582. PMID:27572073

  15. The Therapeutic Effect of Adipose-Derived Mesenchymal Stem Cells for Radiation-Induced Bladder Injury

    PubMed Central

    Qiu, Xuefeng; Zhang, Shiwei; Zhao, Xiaozhi; Fu, Kai; Guo, Hongqian

    2016-01-01

    This study was designed to investigate the protective effect of adipose derived mesenchymal stem cells (AdMSCs) against radiation-induced bladder injury (RIBI). Female rats were divided into 4 groups: (a) controls, consisting of nontreated rats; (b) radiation-treated rats; (c) radiation-treated rats receiving AdMSCs; and (d) radiation-treated rats receiving AdMSCs conditioned medium. AdMSCs or AdMSCs conditioned medium was injected into the muscular layer of bladder 24 h after radiation. Twelve weeks after radiation, urinary bladder tissue was collected for histological assessment and enzyme-linked immunosorbent assay (ELISA) after metabolic cage investigation. At the 1 w, 4 w, and 8 w time points following cells injection, 3 randomly selected rats in RC group and AdMSCs group were sacrificed to track injected AdMSCs. Metabolic cage investigation revealed that AdMSCs showed protective effect for radiation-induced bladder dysfunction. The histological and ELISA results indicated that the fibrosis and inflammation within the bladder were ameliorated by AdMSCs. AdMSCs conditioned medium showed similar effects in preventing radiation-induced bladder dysfunction. In addition, histological data indicated a time-dependent decrease in the number of AdMSCs in the bladder following injection. AdMSCs prevented radiation induced bladder dysfunction and histological changes. Paracrine effect might be involved in the protective effects of AdMSCs for RIBI. PMID:27051426

  16. The DNA-sensing AIM2 inflammasome controls radiation-induced cell death and tissue injury.

    PubMed

    Hu, Bo; Jin, Chengcheng; Li, Hua-Bing; Tong, Jiyu; Ouyang, Xinshou; Cetinbas, Naniye Malli; Zhu, Shu; Strowig, Till; Lam, Fred C; Zhao, Chen; Henao-Mejia, Jorge; Yilmaz, Omer; Fitzgerald, Katherine A; Eisenbarth, Stephanie C; Elinav, Eran; Flavell, Richard A

    2016-11-11

    Acute exposure to ionizing radiation induces massive cell death and severe damage to tissues containing actively proliferating cells, including bone marrow and the gastrointestinal tract. However, the cellular and molecular mechanisms underlying this pathology remain controversial. Here, we show that mice deficient in the double-stranded DNA sensor AIM2 are protected from both subtotal body irradiation-induced gastrointestinal syndrome and total body irradiation-induced hematopoietic failure. AIM2 mediates the caspase-1-dependent death of intestinal epithelial cells and bone marrow cells in response to double-strand DNA breaks caused by ionizing radiation and chemotherapeutic agents. Mechanistically, we found that AIM2 senses radiation-induced DNA damage in the nucleus to mediate inflammasome activation and cell death. Our results suggest that AIM2 may be a new therapeutic target for ionizing radiation exposure. Copyright © 2016, American Association for the Advancement of Science.

  17. Ionizing radiation induces tumor cell lysyl oxidase secretion

    PubMed Central

    2014-01-01

    Background Ionizing radiation (IR) is a mainstay of cancer therapy, but irradiation can at times also lead to stress responses, which counteract IR-induced cytotoxicity. IR also triggers cellular secretion of vascular endothelial growth factor, transforming growth factor β and matrix metalloproteinases, among others, to promote tumor progression. Lysyl oxidase is known to play an important role in hypoxia-dependent cancer cell dissemination and metastasis. Here, we investigated the effects of IR on the expression and secretion of lysyl oxidase (LOX) from tumor cells. Methods LOX-secretion along with enzymatic activity was investigated in multiple tumor cell lines in response to irradiation. Transwell migration assays were performed to evaluate invasive capacity of naïve tumor cells in response to IR-induced LOX. In vivo studies for confirming IR-enhanced LOX were performed employing immunohistochemistry of tumor tissues and ex vivo analysis of murine blood serum derived from locally irradiated A549-derived tumor xenografts. Results LOX was secreted in a dose dependent way from several tumor cell lines in response to irradiation. IR did not increase LOX-transcription but induced LOX-secretion. LOX-secretion could not be prevented by the microtubule stabilizing agent patupilone. In contrast, hypoxia induced LOX-transcription, and interestingly, hypoxia-dependent LOX-secretion could be counteracted by patupilone. Conditioned media from irradiated tumor cells promoted invasiveness of naïve tumor cells, while conditioned media from irradiated, LOX- siRNA-silenced cells did not stimulate their invasive capacity. Locally applied irradiation to tumor xenografts also increased LOX-secretion in vivo and resulted in enhanced LOX-levels in the murine blood serum. Conclusions These results indicate a differential regulation of LOX-expression and secretion in response to IR and hypoxia, and suggest that LOX may contribute towards an IR-induced migratory phenotype in

  18. Radiation-Induced Apoptosis in Breast Cancer Cells.

    DTIC Science & Technology

    1995-09-21

    agarose gel electrophoresis, where the fragments represent the 180-200 base pair nucleosome and multiples thereof. This specific DNA fragmentation in...cancer cell lines tested showed any DNA fragmentation to ladders after exposure to ionizing radiation, although the control HL-60 cells routinely...later days. Some circumstances have been identified where there is not a correlation between DNA fragmentation to oligonucleosomal sized pieces and

  19. Ionizing radiation induces heritable disruption of epithelial cell interactions

    NASA Technical Reports Server (NTRS)

    Park, Catherine C.; Henshall-Powell, Rhonda L.; Erickson, Anna C.; Talhouk, Rabih; Parvin, Bahram; Bissell, Mina J.; Barcellos-Hoff, Mary Helen; Chatterjee, A. (Principal Investigator)

    2003-01-01

    Ionizing radiation (IR) is a known human breast carcinogen. Although the mutagenic capacity of IR is widely acknowledged as the basis for its action as a carcinogen, we and others have shown that IR can also induce growth factors and extracellular matrix remodeling. As a consequence, we have proposed that an additional factor contributing to IR carcinogenesis is the potential disruption of critical constraints that are imposed by normal cell interactions. To test this hypothesis, we asked whether IR affected the ability of nonmalignant human mammary epithelial cells (HMEC) to undergo tissue-specific morphogenesis in culture by using confocal microscopy and imaging bioinformatics. We found that irradiated single HMEC gave rise to colonies exhibiting decreased localization of E-cadherin, beta-catenin, and connexin-43, proteins necessary for the establishment of polarity and communication. Severely compromised acinar organization was manifested by the majority of irradiated HMEC progeny as quantified by image analysis. Disrupted cell-cell communication, aberrant cell-extracellular matrix interactions, and loss of tissue-specific architecture observed in the daughters of irradiated HMEC are characteristic of neoplastic progression. These data point to a heritable, nonmutational mechanism whereby IR compromises cell polarity and multicellular organization.

  20. Ionizing radiation induces heritable disruption of epithelial cell interactions

    PubMed Central

    Park, Catherine C.; Henshall-Powell, Rhonda L.; Erickson, Anna C.; Talhouk, Rabih; Parvin, Bahram; Bissell, Mina J.; Barcellos-Hoff, Mary Helen

    2003-01-01

    Ionizing radiation (IR) is a known human breast carcinogen. Although the mutagenic capacity of IR is widely acknowledged as the basis for its action as a carcinogen, we and others have shown that IR can also induce growth factors and extracellular matrix remodeling. As a consequence, we have proposed that an additional factor contributing to IR carcinogenesis is the potential disruption of critical constraints that are imposed by normal cell interactions. To test this hypothesis, we asked whether IR affected the ability of nonmalignant human mammary epithelial cells (HMEC) to undergo tissue-specific morphogenesis in culture by using confocal microscopy and imaging bioinformatics. We found that irradiated single HMEC gave rise to colonies exhibiting decreased localization of E-cadherin, β-catenin, and connexin-43, proteins necessary for the establishment of polarity and communication. Severely compromised acinar organization was manifested by the majority of irradiated HMEC progeny as quantified by image analysis. Disrupted cell–cell communication, aberrant cell–extracellular matrix interactions, and loss of tissue-specific architecture observed in the daughters of irradiated HMEC are characteristic of neoplastic progression. These data point to a heritable, nonmutational mechanism whereby IR compromises cell polarity and multicellular organization. PMID:12960393

  1. Selenoprotein P Inhibits Radiation-Induced Late Reactive Oxygen Species Accumulation and Normal Cell Injury

    SciTech Connect

    Eckers, Jaimee C.; Kalen, Amanda L.; Xiao, Wusheng; Sarsour, Ehab H.; Goswami, Prabhat C.

    2013-11-01

    Purpose: Radiation is a common mode of cancer therapy whose outcome is often limited because of normal tissue toxicity. We have shown previously that the accumulation of radiation-induced late reactive oxygen species (ROS) precedes cell death, suggesting that metabolic oxidative stress could regulate cellular radiation response. The purpose of this study was to investigate whether selenoprotein P (SEPP1), a major supplier of selenium to tissues and an antioxidant, regulates late ROS accumulation and toxicity in irradiated normal human fibroblasts (NHFs). Methods and Materials: Flow cytometry analysis of cell viability, cell cycle phase distribution, and dihydroethidium oxidation, along with clonogenic assays, were used to measure oxidative stress and toxicity. Human antioxidant mechanisms array and quantitative real-time polymerase chain reaction assays were used to measure gene expression during late ROS accumulation in irradiated NHFs. Sodium selenite addition and SEPP1 overexpression were used to determine the causality of SEPP1 regulating late ROS accumulation and toxicity in irradiated NHFs. Results: Irradiated NHFs showed late ROS accumulation (4.5-fold increase from control; P<.05) that occurs after activation of the cell cycle checkpoint pathways and precedes cell death. The mRNA levels of CuZn- and Mn-superoxide dismutase, catalase, peroxiredoxin 3, and thioredoxin reductase 1 increased approximately 2- to 3-fold, whereas mRNA levels of cold shock domain containing E1 and SEPP1 increased more than 6-fold (P<.05). The addition of sodium selenite before the radiation treatment suppressed toxicity (45%; P<.05). SEPP1 overexpression suppressed radiation-induced late ROS accumulation (35%; P<.05) and protected NHFs from radiation-induced toxicity (58%; P<.05). Conclusion: SEPP1 mitigates radiation-induced late ROS accumulation and normal cell injury.

  2. Selenoprotein P inhibits radiation-induced late reactive oxygen species accumulation and normal cell injury.

    PubMed

    Eckers, Jaimee C; Kalen, Amanda L; Xiao, Wusheng; Sarsour, Ehab H; Goswami, Prabhat C

    2013-11-01

    Radiation is a common mode of cancer therapy whose outcome is often limited because of normal tissue toxicity. We have shown previously that the accumulation of radiation-induced late reactive oxygen species (ROS) precedes cell death, suggesting that metabolic oxidative stress could regulate cellular radiation response. The purpose of this study was to investigate whether selenoprotein P (SEPP1), a major supplier of selenium to tissues and an antioxidant, regulates late ROS accumulation and toxicity in irradiated normal human fibroblasts (NHFs). Flow cytometry analysis of cell viability, cell cycle phase distribution, and dihydroethidium oxidation, along with clonogenic assays, were used to measure oxidative stress and toxicity. Human antioxidant mechanisms array and quantitative real-time polymerase chain reaction assays were used to measure gene expression during late ROS accumulation in irradiated NHFs. Sodium selenite addition and SEPP1 overexpression were used to determine the causality of SEPP1 regulating late ROS accumulation and toxicity in irradiated NHFs. Irradiated NHFs showed late ROS accumulation (4.5-fold increase from control; P<.05) that occurs after activation of the cell cycle checkpoint pathways and precedes cell death. The mRNA levels of CuZn- and Mn-superoxide dismutase, catalase, peroxiredoxin 3, and thioredoxin reductase 1 increased approximately 2- to 3-fold, whereas mRNA levels of cold shock domain containing E1 and SEPP1 increased more than 6-fold (P<.05). The addition of sodium selenite before the radiation treatment suppressed toxicity (45%; P<.05). SEPP1 overexpression suppressed radiation-induced late ROS accumulation (35%; P<.05) and protected NHFs from radiation-induced toxicity (58%; P<.05). SEPP1 mitigates radiation-induced late ROS accumulation and normal cell injury. Copyright © 2013 Elsevier Inc. All rights reserved.

  3. The influence of TRP53 in the dose response of radiation-induced apoptosis, DNA repair and genomic stability in murine haematopoietic cells

    SciTech Connect

    Lemon, Jennifer A.; Taylor, Kristina; Verdecchia, Kyle; Phan, Nghi; Boreham, Douglas R.

    2014-01-01

    Apoptotic and DNA damage endpoints are frequently used as surrogate markers of cancer risk, and have been well-studied in the Trp53+/- mouse model. We report the effect of differing Trp53 gene status on the dose response of ionizing radiation exposures (0.01-2 Gy), with the unique perspective of determining if effects of gene status remain at extended time points. Here we report no difference in the dose response for radiation-induced DNA double-strand breaks in bone marrow and genomic instability (MN-RET levels) in peripheral blood, between wild-type (Trp53+/+) and heterozygous (Trp53+/-) mice. The dose response for Trp53+/+ mice showed higher initial levels of radiation-induced lymphocyte apoptosis relative to Trp53+/- between 0 and 1 Gy. Although this trend was observed up to 12 hours post-irradiation, both genotypes ultimately reached the same level of apoptosis at 14 hours, suggesting the importance of late-onset p53-independent apoptotic responses in this mouse model. Expected radiation-induced G1 cell cycle delay was observed in Trp53+/+ but not Trp53+/-. Although p53 has an important role in cancer risk, we have shown its influence on radiation dose response can be temporally variable. This research highlights the importance of caution when using haematopoietic endpoints as surrogates to extrapolate radiation-induced cancer risk estimation.

  4. Ferulic acid inhibits UVB-radiation induced photocarcinogenesis through modulating inflammatory and apoptotic signaling in Swiss albino mice.

    PubMed

    Ambothi, Kanagalakshmi; Prasad, N Rajendra; Balupillai, Agilan

    2015-08-01

    The aim of this study was to evaluate the photochemopreventive effects of ferulic acid (FA) against chronic ultraviolet-B (290-320 nm) induced oxidative stress, inflammation and angiogenesis in the skin of Swiss albino mice. Chronic UVB exposure (180 mJ/cm(2) for 30 weeks; thrice in a week) induced tumor formation in the mice skin that showed increased expression of carcinogenic and inflammatory markers when compared with the control animals. The intraperitoneal (FAIP) and topical (FAT) administration of FA significantly reduced the incidence of UVB-induced tumor volume and tumor weight in the mice skin. Histopathological studies revealed that both FAIP and FAT administration prevented the UVB-induced hyperplasia, squamous cell carcinoma (SCC) and dysplastic feature in the mice skin. Further, it has been observed that FA treatment reverted chronic UVB-induced oxidative damage (thiobarbituric acid reactive substances, superoxide dismutase, catalase, glutathione peroxidase) accompanied with modulation of vascular endothelial growth factor (VEGF), inducible nitric oxide synthase (iNOS), TNF-α and IL-6 in the mice skin tumor. FA treatment also modulates mutated p53, Bcl-2 and Bax expressions in the UVB-induced mice skin tumor. Thus, the results of the present study indicate ferulic acid has potential against UVB-induced carcinogenesis in the Swiss albino mice.

  5. Radiation-induced bystander signaling from somatic cells to germ cells in Caenorhabditis elegans.

    PubMed

    Guo, Xiaoying; Sun, Jie; Bian, Po; Chen, Lianyun; Zhan, Furu; Wang, Jun; Xu, An; Wang, Yugang; Hei, Tom K; Wu, Lijun

    2013-09-01

    Recently, radiation-induced bystander effects (RIBE) have been studied in mouse models in vivo, which clearly demonstrated bystander effects among somatic cells. However, there is currently no evidence for RIBE between somatic cells and germ cells in animal models in vivo. In the current study, the model animal Caenorhabditis elegans was used to investigate the bystander signaling from somatic cells to germ cells, as well as underlying mechanisms. C. elegans body size allows for precise microbeam irradiation and the abundant mutant strains for genetic dissection relative to currently adopted mouse models make it ideal for such analysis. Our results showed that irradiation of posterior pharynx bulbs and tails of C. elegans enhanced the level of germ cell apoptosis in bystander gonads. The irradiation of posterior pharynx bulbs also increased the level of DNA damage in bystander germ cells and genomic instability in the F1 progeny of irradiated worms, suggesting a potential carcinogenic risk in progeny even only somatic cells of parents are exposed to ionizing radiation (IR). It was also shown that DNA damage-induced germ cell death machinery and MAPK signaling pathways were both involved in the induction of germ cell apoptosis by microbeam induced bystander signaling, indicating a complex cooperation among multiple signaling pathways for bystander effects from somatic cells to germ cells.

  6. Effects of NOX1 on fibroblastic changes of endothelial cells in radiation-induced pulmonary fibrosis

    PubMed Central

    CHOI, SEO-HYUN; KIM, MISEON; LEE, HAE-JUNE; KIM, EUN-HO; KIM, CHUN-HO; LEE, YOON-JIN

    2016-01-01

    Lung fibrosis is a major complication in radiation-induced lung damage following thoracic radiotherapy, while the underlying mechanism has remained to be elucidated. The present study performed immunofluorescence and immunoblot assays on irradiated human pulmonary artery endothelial cells (HPAECs) with or without pre-treatment with VAS2870, a novel NADPH oxidase (NOX) inhibitor, or small hairpin (sh)RNA against NOX1, -2 or -4. VAS2870 reduced the cellular reactive oxygen species content induced by 5 Gy radiation in HPAECs and inhibited phenotypic changes in fibrotic cells, including increased alpha smooth muscle actin and vimentin, and decreased CD31 and vascular endothelial cadherin expression. These fibrotic changes were significantly inhibited by treatment with NOX1 shRNA, but not by NOX2 or NOX4 shRNA. Next, the role of NOX1 in pulmonary fibrosis development was assessed in the lung tissues of C57BL/6J mice following thoracic irradiation using trichrome staining. Administration of an NOX1-specific inhibitor suppressed radiation-induced collagen deposition and fibroblastic changes in the endothelial cells (ECs) of these mice. The results suggested that radiation-induced pulmonary fibrosis may be efficiently reduced by specific inhibition of NOX1, an effect mediated by reduction of fibrotic changes of ECs. PMID:27053172

  7. Modulation of radiation-induced apoptosis and G{sub 2}/M block in murine T-lymphoma cells

    SciTech Connect

    Palayoor, S.T.; Macklis, R.M.; Bump, E.A.; Coleman, C.N.

    1995-03-01

    Radiation-induced apoptosis in lymphocyte-derived cell lines is characterized by endonucleolytic cleavage of cellular DNA within hours after radiation exposure. We have studied this phenomenon qualitatively (DNA gel electrophoresis) and quantitatively (diphenylamine reagent assay) in murine EL4 T-lymphoma cells exposed to {sup 137}Cs {gamma} irradiation. Fragmentation was discernible within 18-24 h after exposure. It increased with time and dose and reached a plateau after 8 Gy of {gamma} radiation. We studied the effect of several pharmacological agents on the radiation-induced G{sub 2}/M block and DNA fragmentation. The agents which reduced the radiation-induced G{sub 2}/M-phase arrest (caffeine, theobromine, theophylline and 2-aminopurine) enhanced the degree of DNA fragmentation at 24 h. In contrast, the agents which sustained the radiation-induced G{sub 2}/M-phase arrest (TPA, DBcAMP, IBMX and 3-aminobenzamide) inhibited the DNA fragmentation at 24 h. These studies on EL4 lymphoma cells are consistent with the hypothesis that cells with radiation-induced genetic damage are eliminated by apoptosis subsequent to a G{sub 2}/M block. Furthermore, it may be possible to modulate the process of radiation-induced apoptosis in lymphoma cells with pharmacological agents that modify the radiation-induced G{sub 2}/M block, and to use this effect in the treatment of patients with malignant disease. 59 refs., 7 figs.

  8. Ionizing radiation induces senescence and differentiation of human dental pulp stem cells.

    PubMed

    Havelek, R; Soukup, T; Ćmielová, J; Seifrtová, M; Suchánek, J; Vávrová, J; Mokrý, J; Muthná, D; Řezáčová, M

    2013-01-01

    Head and neck cancer is one of the most common cancers in Europe. Many current anti-cancer treatments, including ionizing radiation, induce apoptosis via DNA damage. Unfortunately, such treatments are non-selective to cancer cells and produce similar toxicity in normal cells, including adult stem cells. One of the fundamental properties of an adult stem cell is that it does not have any tissue-specific structures that allow it to perform specialized functions. However, under certain stimuli, unspecialized adult stem cells can give rise to specialized cells to generate replacements for cells that are lost during one's life or due to injury or disease. Nevertheless, specialization of stem cells must be controlled by specific milieu and also initiated at the proper time, making the entire process beneficial for tissue recovery and maintaining it for a long time. In this paper we assess whether irradiated dental pulp stem cells have maintained open their options to mature into specialized cells, or whether they have lost their unspecialized (immature) state following irradiation. Our findings showed radiation-induced premature differentiation of dental pulp stem cells towards odonto-/osteoblast lineages in vitro. Matrix calcification was visualized from Day 6 or Day 9 following irradiation of cells expressing low or high levels of CD146, respectively.

  9. Radiation induced bystander effect by GAP junction channels in human fibroblast cell

    NASA Astrophysics Data System (ADS)

    Furusawa, Y.; Shao, C.; Aoki, M.; Kobayashi, Y.; Funayama, T.; Ando, K.

    The chemical factor involved in bystander effect and its transfer pathway were investigated in a confluent human fibroblast cell (AG1522) population. Micronuclei (MN) and G1-phase arrest were detected in cells irradiated by carbon (~100 keV/μm) ions at HIMAC. A very low dose irradiation showed a high effectiveness in producing MN, suggesting a bystander effect. This effectiveness was enhanced by 8-Br-cAMP treatment that increases gap junctional intercellular communication (GJIC). On the other hand, the effect was reduced by 5% DMSO treatment, which reduce the reactive oxygen species (ROS), and suppressed by 100 μM lindane treatment, an inhibitor of GJIC. In addition, the radiation-induced G1-phase arrest was also enhanced by cAMP, and reduced or suppressed by DMSO or lindane. A microbeam device (JAERI) was also used for these studies. It was found that exposing one single cell in a confluent cell population to exactly one argon (~1260 keV/μm) or neon (~430 keV/ μm) ion, additional MN could be detected in many other unirradiated cells. The yield of MN increased with the number of irradiated cells. However, there was no significant difference in the MN induction when the cells were irradiated by increasing number of particles. MN induction by bystander effect was partly reduced by DMSO, and effectively suppressed by lindane. Our results obtained from both random irradiation and precise numbered irradiation indicate that both GJIC and ROS contributed to the radiation-induced bystander effect, but the cell gap junction channels likely play an essential role in the release and transfer of radiation-induced chemical factors.

  10. Low-dose spiruchostatin-B, a potent histone deacetylase inhibitor enhances radiation-induced apoptosis in human lymphoma U937 cells via modulation of redox signaling.

    PubMed

    Rehman, Mati Ur; Jawaid, Paras; Zhao, Qing Li; Li, Peng; Narita, Koichi; Katoh, Tadashi; Shimizu, Tadamichi; Kondo, Takashi

    2016-06-01

    Spiruchostatin B (SP-B), is a potent histone deacetylase (HDAC) inhibitor, in addition to HDAC inhibition, the pharmacological effects of SP-B are also attributed to its ability to produce intracellular reactive oxygen species (ROS), particularly H2O2. In this study, we investigated the effects of low dose (non-toxic) SP-B on radiation-induced apoptosis in human lymphoma U937 cells in vitro. The treatment of cells with low-dose SP-B induced the acetylation of histones, however, does not induce apoptosis. Whereas, the combined treatment with SP-B and radiation significantly enhanced the radiation-induced apoptosis, suggesting the potential role of this combined treatment for future radiation therapy. Interestingly, the enhancement of apoptosis was accompanied by significant increased in the ROS generation. Pre-treatment with an antioxidant, N-acetyl-l-cysteine (NAC) significantly inhibited the enhancement of apoptosis induced by combined treatment, indicating that ROS play an essential role. It was also found that SP-B combined with radiation caused the activation of death receptor and intrinsic apoptotic pathways, via modulation of ROS-mediated signaling. Moreover, SP-B also significantly enhanced the radiation-induced apoptosis in other lymphoma cell lines such as Molt-4 and HL-60. Taken together, our findings suggest that the low-dose SP-B enhances radiation-induced apoptosis via modulation of redox signaling because of its ability to serve as an intracellular ROS generating agent, mainly (H2O2 or [Formula: see text]). This study provides further insights into the mechanism of action of SP-B with radiation and demonstrates that SP-B can be used as a future novel sensitizer for radiation therapy.

  11. Reduction of radiation-induced cell cycle blocks by caffeine does not necessarily lead to increased cell killing

    SciTech Connect

    Musk, S.R. )

    1991-03-01

    The effect of caffeine upon the radiosensitivities of three human tumor lines was examined and correlated with its action upon the radiation-induced S-phase and G2-phase blocks. Caffeine was found to reduce at least partially the S-phase and G2-phase blocks in all the cell lines examined but potentiated cytotoxicity in only one of the three tumor lines. That reductions have been demonstrated to occur in the absence of increased cell killing provides supporting evidence for the hypothesis that reductions may not be causal in those cases when potentiation of radiation-induced cytotoxicity is observed in the presence of caffeine.

  12. Protection against radiation-induced oxidative stress in cultured human epithelial cells by treatment with antioxidant agents

    SciTech Connect

    Wan, X. Steven; Ware, Jeffrey H.; Zhou, Zhaozong; Donahue, Jeremiah J.; Guan, Jun; Kennedy, Ann R. . E-mail: akennedy@mail.med.upenn.edu

    2006-04-01

    Purpose: To evaluate the protective effects of antioxidant agents against space radiation-induced oxidative stress in cultured human epithelial cells. Methods and Materials: The effects of selected concentrations of N-acetylcysteine, ascorbic acid, sodium ascorbate, co-enzyme Q10, {alpha}-lipoic acid, L-selenomethionine, and vitamin E succinate on radiation-induced oxidative stress were evaluated in MCF10 human breast epithelial cells exposed to radiation with X-rays, {gamma}-rays, protons, or high mass, high atomic number, and high energy particles using a dichlorofluorescein assay. Results: The results demonstrated that these antioxidants are effective in protecting against radiation-induced oxidative stress and complete or nearly complete protection was achieved by treating the cells with a combination of these agents before and during the radiation exposure. Conclusion: The combination of antioxidants evaluated in this study is likely be a promising countermeasure for protection against space radiation-induced adverse biologic effects.

  13. Enhanced radiation-induced cell killing by Herbimycin A pre-treatment

    NASA Astrophysics Data System (ADS)

    Noguchi, Miho; Hirayama, Ryoichi; Druzhinin, Sergey; Okayasu, Ryuichi

    2009-12-01

    Herbimycin A (HA), as in Geldanamycin, binds to conserved pockets of heat shock protein 90 (Hsp90) and inhibits its chaperone functions. Hsp90 plays an integral role in cancer cell growth and survival, because it maintains the stability of several key proteins by its chaperone's activity. It is known that some of the proteins associated with radiation responses are functionally stabilized by Hsp90. In this study, we investigated the effect of HA on radiosensitivity in human cancer cells and the mechanism related to the sensitization. In order to gain a mechanistic insight of this sensitization, we examined repair of DNA double-strand breaks (DSBs) in irradiated human cancer cells pre-treated with HA, as unrepaired DSBs are thought to be the main cause of radiation-induced cell death. Cellular radiosensitivity was determined by clonogenic assay, and the DSB rejoining kinetics was examined by constant field gel electrophoresis. SQ-5, a lung squamous carcinoma cell line, showed synergistic increase in radiosensitivity when cells were pre-treated with HA. In addition, HA significantly inhibited repair of radiation-induced DSBs. These results suggest that the combination of HA and ionizing radiation may be a useful therapeutic strategy for treating certain cancer cells.

  14. Consequences of ionizing radiation-induced damage in human neural stem cells.

    PubMed

    Acharya, Munjal M; Lan, Mary L; Kan, Vickie H; Patel, Neal H; Giedzinski, Erich; Tseng, Bertrand P; Limoli, Charles L

    2010-12-15

    Cranial irradiation remains a frontline treatment for brain cancer, but also leads to normal tissue damage. Although low-dose irradiation (≤10 Gy) causes minimal histopathologic change, it can elicit variable degrees of cognitive dysfunction that are associated with the depletion of neural stem cells. To decipher the mechanisms underlying radiation-induced stem cell dysfunction, human neural stem cells (hNSCs) subjected to clinically relevant irradiation (0-5 Gy) were analyzed for survival parameters, cell-cycle alterations, DNA damage and repair, and oxidative stress. hNSCs showed a marked sensitivity to low-dose irradiation that was in part due to elevated apoptosis and the inhibition of cell-cycle progression that manifested as a G2/M checkpoint delay. Efficient removal of DNA double-strand breaks was indicated by the disappearance of γ-H2AX nuclear foci. A dose-responsive and persistent increase in oxidative and nitrosative stress was found in irradiated hNSCs, possibly the result of a higher metabolic activity in the fraction of surviving cells. These data highlight the marked sensitivity of hNSCs to low-dose irradiation and suggest that long-lasting perturbations in the CNS microenvironment due to radiation-induced oxidative stress can compromise the functionality of neural stem cells.

  15. Radiation induces the generation of cancer stem cells: A novel mechanism for cancer radioresistance

    PubMed Central

    Li, Fengsheng; Zhou, Kunming; Gao, Ling; Zhang, Bin; Li, Wei; Yan, Weijuan; Song, Xiujun; Yu, Huijie; Wang, Sinian; Yu, Nan; Jiang, Qisheng

    2016-01-01

    Radioresistance remains a major obstacle for the radiotherapy treatment of cancer. Previous studies have demonstrated that the radioresistance of cancer is due to the existence of intrinsic cancer stem cells (CSCs), which represent a small, but radioresistant cell subpopulation that exist in heterogeneous tumors. By contrast, non-stem cancer cells are considered to be radiosensitive and thus, easy to kill. However, recent studies have revealed that under conditions of radiation-induced stress, theoretically radiosensitive non-stem cancer cells may undergo dedifferentiation subsequently obtaining the phenotypes and functions of CSCs, including high resistance to radiotherapy, which indicates that radiation may directly result in the generation of novel CSCs from non-stem cancer cells. These findings suggest that in addition to intrinsic CSCs, non-stem cancer cells may also contribute to the relapse and metastasis of cancer following transformation into CSCs. This review aims to investigate the radiation-induced generation of CSCs, its association with epithelial-mesenchymal transition and its significance with regard to the radioresistance of cancer. PMID:27899964

  16. The roles of mitochondria in radiation-induced autophagic cell death in cervical cancer cells.

    PubMed

    Chen, Zongyan; Wang, Benli; Yu, Feifei; Chen, Qiao; Tian, Yuxi; Ma, Shumei; Liu, Xiaodong

    2016-03-01

    Mitochondria as the critical powerhouse of eukaryotic cells play important roles in regulating cell survival or cell death. Under numerous stimuli, impaired mitochondria will generate massive reactive oxygen species (ROS) which participate in the regulation of vital signals and could even determine the fate of cancer cells. While the roles of mitochondria in radiation-induced autophagic cell death still need to be elucidated. Human cervical cancer cell line, Hela, was used, and the SOD2 silencing model (SOD2-Ri) was established by gene engineering. Cell viability was detected by methyl thiazolyl tetrazolium (MTT) assays, MitoTracker Green staining was used to detect mitochondrial mass, Western blot was used to detect protein expression, and the level of ROS, autophagy, and mitochondrial membrane potential (MMP) were analyzed by flow cytometry. Ionizing radiation (IR) could induce the increase of MAPLC3-II/MAPLC3-I ratio, Beclin1 expression, and ROS generation but decrease the MMP in a time-dependent manner. After SOD2 silencing, the IR-induced changes of ROS and the MMP were significantly enhanced. Moreover, both the radio sensitivity and autophagy increased in SOD2-Ri cells. Whereas, compared with SOD2-Ri, the opposite results were obtained by NAC, an antioxidant. After the treatment with the inhibitor of mitochondrial electron-transport chain complex II, thenoyltrifluoroacetone (TTFA), the rate of autophagy, ROS, and the total cell death induced by IR increased. In addition, the decrease of MMP was more obvious. However, these results were reversed by cyclosporine A (CsA). IR could induce ROS generation and mitochondrial damage which lead to autophagic cell death in Hela cells.

  17. Mediators involved in the immunomodulatory effects of apoptotic cells.

    PubMed

    Saas, Philippe; Bonnefoy, Francis; Kury-Paulin, Stephanie; Kleinclauss, François; Perruche, Sylvain

    2007-07-15

    Immunomodulatory properties are attributed to apoptotic cells. These properties have been used to modulate allogeneic immune responses in experimental transplantation settings. In independent studies, apoptotic cell infusion has been shown to favor hematopoietic cell engraftment, to increase heart graft survival, and to delay the lethal onset of graft-versus-host disease (GVHD). The goal of this review was to discuss how apoptotic cell infusion interferes with graft rejection or host rejection (i.e., GVHD) and to focus on the potential mediators or "perpetuators" involved in apoptotic cell-induced immunomodulation. Particular emphasis on apoptotic cell phagocytosis, transforming growth factor (TGF)-beta secretion, and regulatory T cell induction was performed. Stimulating "naturally" immunosuppressive molecules (i.e., TGF-beta) or immunomodulatory cells ("alternatively-activated" macrophages, certain dendritic cell subsets, or regulatory T cells) in a physiological manner by using apoptotic cell infusion can be a promising way to induce tolerance.

  18. Live cell microscopy analysis of radiation-induced DNA double-strand break motion

    PubMed Central

    Jakob, B.; Splinter, J.; Durante, M.; Taucher-Scholz, G.

    2009-01-01

    We studied the spatiotemporal organization of DNA damage processing by live cell microscopy analysis in human cells. In unirradiated U2OS osteosarcoma and HeLa cancer cells, a fast confined and Brownian-like motion of DNA repair protein foci was observed, which was not altered by radiation. By analyzing the motional activity of GFP-53BP1 foci in live cells up to 12-h after irradiation, we detected an additional slower mobility of damaged chromatin sites showing a mean square displacement of ≈0.6 μm2/h after exposure to densely- or sparsely-ionizing radiation, most likely driven by normal diffusion of chromatin. Only occasionally, larger translational motion connected to morphological changes of the whole nucleus could be observed. In addition, there was no general tendency to form repair clusters in the irradiated cells. We conclude that long-range displacements of damaged chromatin domains do not generally occur during DNA double-strand break repair after introduction of multiple damaged sites by charged particles. The occasional and in part transient appearance of cluster formation of radiation-induced foci may represent a higher mobility of chromatin along the ion trajectory. These observations support the hypothesis that spatial proximity of DNA breaks is required for the formation of radiation-induced chromosomal exchanges. PMID:19221031

  19. Mediators involved in the immunomodulatory effects of apoptotic cells

    PubMed Central

    Saas, Philippe; Bonnefoy, Francis; Kury-Paulin, Stephanie; Kleinclauss, François M.; Perruche, Sylvain

    2007-01-01

    Immunomodulatory properties are attributed to apoptotic cells. These properties have been used to modulate allogeneic immune responses in experimental transplantation settings. In independent studies, apoptotic cell infusion has been shown to favor hematopoietic cell engraftment, to increase heart graft survival and to delay the lethal onset of graft-versus-host disease (GVHD). The goal of this review was to discuss how apoptotic cell infusion interferes with graft rejection or host rejection (i.e., GVHD) and to focus on the potential mediators or “perpetuators” involved in apoptotic cell-induced immunomodulation. Particular emphasis on apoptotic cell phagocytosis, TGF-β secretion and regulatory T cell induction was performed. Stimulating “naturally” immunosuppressive molecules (i.e., TGF-β) or immunomodulatory cells (“alternatively-activated” macrophages, certain DC subsets or regulatory T cells) in a physiological manner by using apoptotic cell infusion can be a promising way to induce tolerance. PMID:17632410

  20. Molecular mechanisms of radiation-induced genomic instability in human cells

    SciTech Connect

    Liber, Howard L.

    2003-02-13

    The overall strategy was to create a series of isogenic human cell lines that differ in key elements of cell cycle checkpoint, apoptosis, or DNA repair in response to radiation-induced damage. The goal then was to quantify the fractions of cells within a population that exhibit reduced telomere lengths and relate this to the genetic background of the cell, as well as to the response to ionizing radiation. Association between telomere length and degree of genomic instability in the population is being examined for seven closely related cell lines, that vary in p53 status, bcl-2 status, or ability to repair double strand breaks. Experiments utilize gamma rays at doses of 0, 10, and 200 cGy. During this time period the effort concentrated on generating data with two cell lines. Approximately one-third of the required clones were isolated, and analyses for mutagenesis and chromosome aberrations were undertaken.

  1. Flow cytometric determination of radiation-induced chromosome damage and its correlation with cell survival

    SciTech Connect

    Welleweerd, J.; Wilder, M.E.; Carpenter, S.G.; Raju, M.R.

    1984-07-01

    Chinese hamster M3-1 cells were irradiated with several doses of x rays or ..cap alpha.. particles from /sup 238/Pu. Propidium iodide-stained chromosome suspensions were prepared at different times after irradiation; cells were also assayed for survival. The DNA histograms of these chromosomes showed increased background counts with increased doses of radiation. This increase in background was cell-cycle dependent and was correlated with cell survival. The correlation between radiation-induced chromosome damage and cell survival was the same for X rays and ..cap alpha.. particles. Data are presented which indicate that flow cytometric analysis of chromosomes of irradiated cell populations can be a useful adjunct to classical cytogenic analysis of irradiation-induced chromosomal damage by virtue of its ability to express and measure chromosomal damage not seen by classical cytogenic methods.

  2. Protective effect of mild endoplasmic reticulum stress on radiation-induced bystander effects in hepatocyte cells.

    PubMed

    Xie, Yuexia; Ye, Shuang; Zhang, Jianghong; He, Mingyuan; Dong, Chen; Tu, Wenzhi; Liu, Peifeng; Shao, Chunlin

    2016-12-13

    Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the defense and self-protective mechanisms of bystander normal cells are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 cells under either normoxia or hypoxia, where the ratio of the yield of bystander MN induction to the yield of radiation-induced MN formation under hypoxia was much higher than that of normoxia. Nonetheless, thapsigargin induced endoplasmic reticulum (ER) stress and dramatically suppressed this bystander response manifested as the decrease of MN and apoptosis inductions. Meanwhile, the interference of BiP gene, a major ER chaperone, amplified the detrimental RIBE. More precisely, thapsigargin provoked ER sensor of PERK to initiate an instantaneous and moderate ER stress thus defensed the hazard form RIBE, while BiP depletion lead to persistently destroyed homeostasis of ER and exacerbated cell injury. These findings provide new insights that the mild ER stress through BiP-PERK-p-eIF2α signaling pathway has a profound role in protecting cellular damage from RIBE and hence may decrease the potential secondary cancer risk after cancer radiotherapy.

  3. Protective effect of mild endoplasmic reticulum stress on radiation-induced bystander effects in hepatocyte cells

    PubMed Central

    Xie, Yuexia; Ye, Shuang; Zhang, Jianghong; He, Mingyuan; Dong, Chen; Tu, Wenzhi; Liu, Peifeng; Shao, Chunlin

    2016-01-01

    Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the defense and self-protective mechanisms of bystander normal cells are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 cells under either normoxia or hypoxia, where the ratio of the yield of bystander MN induction to the yield of radiation-induced MN formation under hypoxia was much higher than that of normoxia. Nonetheless, thapsigargin induced endoplasmic reticulum (ER) stress and dramatically suppressed this bystander response manifested as the decrease of MN and apoptosis inductions. Meanwhile, the interference of BiP gene, a major ER chaperone, amplified the detrimental RIBE. More precisely, thapsigargin provoked ER sensor of PERK to initiate an instantaneous and moderate ER stress thus defensed the hazard form RIBE, while BiP depletion lead to persistently destroyed homeostasis of ER and exacerbated cell injury. These findings provide new insights that the mild ER stress through BiP-PERK-p-eIF2α signaling pathway has a profound role in protecting cellular damage from RIBE and hence may decrease the potential secondary cancer risk after cancer radiotherapy. PMID:27958308

  4. Radiation induced endothelial cell retraction in vitro: correlation with acute pulmonary edema.

    PubMed

    Onoda, J M; Kantak, S S; Diglio, C A

    1999-01-01

    We determined the effects of low dose radiation (<200 cGy) on the cell-cell integrity of confluent monolayers of pulmonary microvascular endothelial cells (PMEC). We observed dose- and time-dependent reversible radiation induced injuries to PMEC monolayers characterized by retraction (loss of cell-cell contact) mediated by cytoskeletal F-actin reorganization. Radiation induced reorganization of F-actin microfilament stress fibers was observed > or =30 minutes post irradiation and correlated positively with loss of cell-cell integrity. Cells of irradiated monolayers recovered to form contact inhibited monolayers > or =24 hours post irradiation; concomitantly, the depolymerized microfilaments organized to their pre-irradiated state as microfilament stress fibers arrayed parallel to the boundaries of adjacent contact-inhibited cells. Previous studies by other investigators have measured slight but significant increases in mouse lung wet weight >1 day post thoracic or whole body radiation (> or =500 cGy). Little or no data is available concerning time intervals <1 day post irradiation, possibly because of the presumption that edema is mediated, at least in part, by endothelial cell death or irreversible loss of barrier permeability functions which may only arise 1 day post irradiation. However, our in vitro data suggest that loss of endothelial barrier function may occur rapidly and at low dose levels (< or =200 cGy). Therefore, we determined radiation effects on lung wet weight and observed significant increases in wet weight (standardized per dry weight or per mouse weight) in < or =5 hours post thoracic exposure to 50 200 cGy x-radiation. We suggest that a single fraction of radiation even at low dose levels used in radiotherapy, may induce pulmonary edema by a reversible loss of endothelial cell-cell integrity and permeability barrier function.

  5. Enhanced radiation-induced cytotoxic effect by 2-ME in glioma cells is mediated by induction of cell cycle arrest and DNA damage via activation of ATM pathways.

    PubMed

    Zou, Huichao; Zhao, Shiguang; Zhang, Jianhua; Lv, Gongwei; Zhang, Xu; Yu, Hongwei; Wang, Huibo; Wang, Ligang

    2007-12-14

    Glioblastoma multiform is the most common malignant primary brain tumor in adults, but there remains no effective therapeutic approach. 2-methoxyestradiol (2-ME), which is a naturally occurring metabolite of 17beta-estradiol, was shown to enhance radiotherapeutic effect in certain tumors; however, whether 2-ME can also enhance the sensitivity of glioma cells to radiotherapy remains unknown. The present study, therefore, was to address this issue using two human glioma cell lines (T98G and U251MG). These cells were irradiated with and without 2-ME and then clonogenic assay, apoptosis assay, DNA damage, and cell cycle change were examined. Results showed that 2-ME significantly enhances radiation-induced cell death in both glioma cells, shown by decreasing cell viability and increasing apoptotic cell death. No such radiosensitizing effect was observed if cells pre-treated with Estrodiol, suggesting the specifically radiosensitizing effect of 2-ME rather than a general effect of estrodials. The enhanced radio-cytotoxic effect in glioma cells by 2-ME was found to be associated with its enhancement of G(2)/M arrest and DNA damage, and phosphorylated ATM protein kinases as well as cell cycle checkpoint protein Chk2. Furthermore, inhibition of ATM by ATM inhibitor abolished 2-ME-activated Chk2 and enhanced radio-cytotoxic effects. These results suggest that 2-ME enhancement of the sensitivity of glioma cell lines to radiotherapy is mediated by induction of G2/M cell cycle arrest and increased DNA damage via activation of ATM kinases.

  6. Transmission of persistent ionizing radiation-induced foci through cell division in human primary cells.

    PubMed

    Vaurijoux, Aurelie; Voisin, Pascale; Freneau, Amelie; Barquinero, Joan Francesc; Gruel, Gaetan

    2017-03-01

    Unrepaired DNA double-strand breaks (DSBs) induced by ionizing radiation are associated with lethal effects and genomic instability. After the initial breaks and chromatin destabilization, a set of post-translational modifications of histones occurs, including phosphorylation of serine 139 of histone H2AX (γH2AX), which leads to the formation of ionizing radiation-induced foci (IRIF). DSB repair results in the disappearance of most IRIF within hours after exposure, although some remain 24h after irradiation. Their relation to unrepaired DSBs is generally accepted but still controversial. This study evaluates the frequency and kinetics of persistent IRIF and analyzes their impact on cell proliferation. We observed persistent IRIF up to 7 days postirradiation, and more than 70% of cells exposed to 5Gy had at least one of these persistent IRIF 24h after exposure. Moreover we demonstrated that persistent IRIF did not block cell proliferation definitively. The frequency of IRIF was lower in daughter cells, due to asymmetric distribution of IRIF between some of them. We report a positive association between the presence of IRIF and the likelihood of DNA missegregation. Hence, the structure formed after the passage of a persistent IRI focus across the S and G2 phases may impede the correct segregation of the affected chromosome's sister chromatids. The ensuing abnormal resolution of anaphase might therefore cause the nature of IRIF in daughter-cell nuclei to differ before and after the first cell division. The resulting atypical chromosomal assembly may be lethal or result in a gene dosage imbalance and possibly enhanced genomic instability, in particular in the daughter cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. REDD1 protects osteoblast cells from gamma radiation-induced premature senescence.

    PubMed

    Li, Xiang Hong; Ha, Cam T; Fu, Dadin; Xiao, Mang

    2012-01-01

    Radiotherapy is commonly used for cancer treatment. However, it often results in side effects due to radiation damage in normal tissue, such as bone marrow (BM) failure. Adult hematopoietic stem and progenitor cells (HSPC) reside in BM next to the endosteal bone surface, which is lined primarily by hematopoietic niche osteoblastic cells. Osteoblasts are relatively more radiation-resistant than HSPCs, but the mechanisms are not well understood. In the present study, we demonstrated that the stress response gene REDD1 (regulated in development and DNA damage responses 1) was highly expressed in human osteoblast cell line (hFOB) cells after γ irradiation. Knockdown of REDD1 with siRNA resulted in a decrease in hFOB cell numbers, whereas transfection of PCMV6-AC-GFP-REDD1 plasmid DNA into hFOB cells inhibited mammalian target of rapamycin (mTOR) and p21 expression and protected these cells from radiation-induced premature senescence (PS). The PS in irradiated hFOB cells were characterized by significant inhibition of clonogenicity, activation of senescence biomarker SA-β-gal, and the senescence-associated cytokine secretory phenotype (SASP) after 4 or 8 Gy irradiation. Immunoprecipitation assays demonstrated that the stress response proteins p53 and nuclear factor κ B (NFkB) interacted with REDD1 in hFOB cells. Knockdown of NFkB or p53 gene dramatically suppressed REDD1 protein expression in these cells, indicating that REDD1 was regulated by both factors. Our data demonstrated that REDD1 is a protective factor in radiation-induced osteoblast cell premature senescence.

  8. Gene-modified Mesenchymal Stem Cells Protect Against Radiation-induced Lung Injury

    PubMed Central

    Xue, Jianxin; Li, Xin; Lu, You; Gan, Lu; Zhou, Lin; Wang, Yongsheng; Lan, Jie; Liu, Shurui; Sun, Lan; Jia, Li; Mo, Xianming; Li, Jian

    2013-01-01

    Radiation-induced lung injury (RILI) presents a common and major obstacle in the radiotherapy of thoracic cancers. The aim of this study was to examine whether RILI could be alleviated by mesenchymal stem cells (MSCs) expressing soluble transforming growth factor-β (TGF-β) type II receptor via an adenovirus (Ad-sTβR). Here, we systemically administered male MSCs into female mice challenged with thoracic irradiation. The data showed that either MSCs or Ad-sTβR transduced MSCs (Ad-sTβR-MSCs) specifically migrated into radiation-injured lung. Ad-sTβR-MSCs obviously alleviated lung injury, as reflected by survival and histopathology data, as well as the assays of malondialdehyde (MDA), hydroxyproline, plasma cytokines, and the expression of connective tissue growth factor (CTGF) and α-smooth muscle actin (α-SMA). Furthermore, MSCs and Ad-sTβR-MSCs could adopt the characteristics of alveolar type II (ATII) cells. However, the MSCs levels in the lungs were relatively low to account for the noted therapeutic effects, suggesting the presence of other mechanisms. In vivo, MSCs-conditioned medium (MSCs CM) significantly attenuated RILI. In vitro, MSCs CM protected ATII cells against radiation-induced apoptosis and DNA damage, and modulated the inflammatory response, indicating the beneficial effects of MSCs are largely due to its paracrine activity. Our results provide a novel insight for RILI therapy that currently lack efficient treatments. PMID:23299797

  9. Radiation-Induced Dermatitis is Mediated by IL17-Expressing γδ T Cells.

    PubMed

    Liao, Wupeng; Hei, Tom K; Cheng, Simon K

    2017-04-01

    Radiation dermatitis is a serious cutaneous injury caused by radiation therapy or upon accidental nuclear exposure. However, the pathogenic immune mechanisms underlying this injury are still poorly understood. We seek to discover how the dysregulated immune response after irradiation orchestrates skin inflammation. The skin on the left flank of C57BL/6J wild-type and C57BL/6J Tcrd(-/-) mice, which are deficit in γδ T cells, was exposed to a single X-ray dose of 25 Gy, and the right-flank skin was used as a sham-irradiated control. At 4 weeks postirradiation, the wild-type skin exhibited signs of depilation, erythema and desquamation. Histological analysis showed hyperproliferation of keratinocytes and acanthosis. Dramatic elevation of IL17-expressing T cells was identified from the irradiated skin, which was mainly contributed by γδ T cells and innate lymphoid cells, rather than Th17 cells. Furthermore, protein levels of critical cytokines for IL17-expressing γδ T cell activation, IL1β and IL23 were found markedly upregulated. Lastly, radiation-induced dermatitis was significantly attenuated in γδ T cell knockout mice. In vitro, normal human epidermal keratinocytes (NHEKs) could be initiator cells of inflammation by providing a great number of pro-inflammatory mediators upon radiation, and as well as effector cells of epidermal hyperplasia in response to exogenous IL17 and/or IL22 treatment. Our findings implicate a novel role of IL17-expressing γδ T cells in mediating radiation-induced skin inflammation. This study reveals the innate immune response pathway as a potential therapeutic target for radiation skin injury.

  10. Radiation-Induced Dermatitis is Mediated by IL17-Expressing γδ T Cells.

    PubMed

    Liao, Wupeng; Hei, Tom K; Cheng, Simon K

    2017-02-08

    Radiation dermatitis is a serious cutaneous injury caused by radiation therapy or upon accidental nuclear exposure. However, the pathogenic immune mechanisms underlying this injury are still poorly understood. We seek to discover how the dysregulated immune response after irradiation orchestrates skin inflammation. The skin on the left flank of C57BL/6J wild-type and C57BL/6J Tcrd(-/-) mice, which are deficit in γδ T cells, was exposed to a single X-ray dose of 25 Gy, and the right-flank skin was used as a sham-irradiated control. At 4 weeks postirradiation, the wild-type skin exhibited signs of depilation, erythema and desquamation. Histological analysis showed hyperproliferation of keratinocytes and acanthosis. Dramatic elevation of IL17-expressing T cells was identified from the irradiated skin, which was mainly contributed by γδ T cells and innate lymphoid cells, rather than Th17 cells. Furthermore, protein levels of critical cytokines for IL17-expressing γδ T cell activation, IL1β and IL23 were found markedly upregulated. Lastly, radiation-induced dermatitis was significantly attenuated in γδ T cell knockout mice. In vitro, normal human epidermal keratinocytes (NHEKs) could be initiator cells of inflammation by providing a great number of pro-inflammatory mediators upon radiation, and as well as effector cells of epidermal hyperplasia in response to exogenous IL17 and/or IL22 treatment. Our findings implicate a novel role of IL17-expressing γδ T cells in mediating radiation-induced skin inflammation. This study reveals the innate immune response pathway as a potential therapeutic target for radiation skin injury.

  11. Apoptotic cell clearance: basic biology and therapeutic potential

    PubMed Central

    Poon, Ivan K. H.; Lucas, Christopher D.

    2014-01-01

    Prompt removal of apoptotic cells by phagocytes is important for maintaining tissue homeostasis. The molecular and cellular events that underpin apoptotic cell recognition and uptake, and the subsequent biological responses are increasingly better defined. The detection and disposal of apoptotic cells generally promote an anti-inflammatory response at the tissue level, as well as immunological tolerance. Consequently, defects in apoptotic cell clearance have been linked with a variety of inflammatory diseases and autoimmunity. Conversely, under certain conditions such as killing tumour cells by specific cell death inducers, the recognition of apoptotic tumour cells can promote an immunogenic response and anti-tumour immunity. Here, we review the current understanding of the complex process of apoptotic cell clearance in physiology and pathology, and discuss how this knowledge could be harnessed for new therapeutic strategies. PMID:24481336

  12. Radiation-induced division delay in Chinese hamster ovary fibroblast and carcinoma cells: dose effect and ploidy. [X-ray

    SciTech Connect

    Kimler, B.F.; Leeper, D.B.; Schneiderman, M.H.

    1981-02-01

    The mitotic selection procedure for cell cycle analysis was utilized to investigate the G/sub 2/ transition point for and the duration of radiation-induced division delay in diploid and tetraploid Chinese hamster ovary (CHO) fibroblasts and in Chinese hamster ovarian carcinoma cells. The location of the radiation-induced division delay transition point was dose independent at high doses and located approximately 42 min before division. At lower doses only an estimate of the point of blockade was possible; but the G/sub 2/ transition point appeared to be earlier in the cell cycle. The duration of radiation-induced division delay was dose dependent. This response is consistent with a sensitive population of cells in late G/sub 2/ that define the location of the transition point and the length of division delay. There was no difference observed in the dose response for radiation-induced division delay between the pseudotetraploid cell line of CHO and the pseudodiploid parent strain. However, in the cell line derived from a spontaneous Chinese hamster ovarian carcinoma the division delay was 39 +- 4 min/Gy. Therefore, radiation-induced division delay is independent of chromosome ploidy, but can show intraspecies cell line specificity.

  13. BK K+ channel blockade inhibits radiation-induced migration/brain infiltration of glioblastoma cells

    PubMed Central

    Klumpp, Lukas; Haehl, Erik; Schilbach, Karin; Lukowski, Robert; Kühnle, Matthias; Bernhardt, Günther; Buschauer, Armin; Zips, Daniel; Ruth, Peter; Huber, Stephan M.

    2016-01-01

    Infiltration of the brain by glioblastoma cells reportedly requires Ca2+ signals and BK K+ channels that program and drive glioblastoma cell migration, respectively. Ionizing radiation (IR) has been shown to induce expression of the chemokine SDF-1, to alter the Ca2+ signaling, and to stimulate cell migration of glioblastoma cells. Here, we quantified fractionated IR-induced migration/brain infiltration of human glioblastoma cells in vitro and in an orthotopic mouse model and analyzed the role of SDF-1/CXCR4 signaling and BK channels. To this end, the radiation-induced migratory phenotypes of human T98G and far-red fluorescent U-87MG-Katushka glioblastoma cells were characterized by mRNA and protein expression, fura-2 Ca2+ imaging, BK patch-clamp recording and transfilter migration assay. In addition, U-87MG-Katushka cells were grown to solid glioblastomas in the right hemispheres of immunocompromised mice, fractionated irradiated (6 MV photons) with 5 × 0 or 5 × 2 Gy, and SDF-1, CXCR4, and BK protein expression by the tumor as well as glioblastoma brain infiltration was analyzed in dependence on BK channel targeting by systemic paxilline application concomitant to IR. As a result, IR stimulated SDF-1 signaling and induced migration of glioblastoma cells in vitro and in vivo. Importantly, paxilline blocked IR-induced migration in vivo. Collectively, our data demonstrate that fractionated IR of glioblastoma stimulates and BK K+ channel targeting mitigates migration and brain infiltration of glioblastoma cells in vivo. This suggests that BK channel targeting might represent a novel approach to overcome radiation-induced spreading of malignant brain tumors during radiotherapy. PMID:26893360

  14. α3β1 integrin promotes radiation-induced migration of meningioma cells.

    PubMed

    Gogineni, Venkateswara Rao; Nalla, Arun Kumar; Gupta, Reshu; Gujrati, Meena; Klopfenstein, Jeffrey D; Mohanam, Sanjeeva; Rao, Jasti S

    2011-06-01

    Cell motility is influenced by the microenvironment, signal transduction and cytoskeleton rearrangement. Cancer cells become resistant to these control mechanisms and gain the ability to move throughout the body and invade healthy tissues, which leads to metastatic disease. Integrins respond to context-dependent cues and promote cell migration and survival in cancer cells. In the present study, we analyzed the role of integrins in radiation-induced migration of meningioma cells. Migration and cell proliferation assays revealed that radiation treatment (7 Gy) significantly increased migration and decreased proliferation in two cell lines, IOMM-Lee and CH-157-MN. α3 and β1 integrins were overexpressed at both the protein and transcript levels after radiation treatment and a function-blocking α3β1 antibody inhibited the radiation-induced migration. Immunofluorescence studies illustrated the localization of α3 integrin and F-actin at the migration front of irradiated cells. Further, an increase in phosphorylation of FAK and ERK was observed, while both FAK phosphorylation inhibitor and FAK shRNA inhibited ERK phosphorylation and downregulated uPA and vinculin. In addition to the co-localization of FAK and ERK at the migration front, these FAK-inhibition results link the downstream effects of ERK to FAK. Correspondingly, U0126 quenched ERK phosphorylation and reduced the expression of molecules involved in migration. Furthermore, brain sections of the animals implanted with tumors followed by radiation treatment showed elevated levels of α3 integrin and active ERK. Taken together, our results show that radiation treatment enhances the migration of meningioma cells with the involvement of α3β1 integrin-mediated signaling via FAK and ERK.

  15. Apoptotic cells subjected to cold/warming exposure disorganize apoptotic microtubule network and undergo secondary necrosis.

    PubMed

    Oropesa-Ávila, Manuel; Fernández-Vega, Alejandro; de la Mata, Mario; Garrido-Maraver, Juan; Cotán, David; Paz, Marina Villanueva; Pavón, Ana Delgado; Cordero, Mario D; Alcocer-Gómez, Elizabet; de Lavera, Isabel; Lema, Rafael; Zaderenko, Ana Paula; Sánchez-Alcázar, José A

    2014-09-01

    Apoptotic microtubule network (AMN) is organized during apoptosis, forming a cortical structure beneath the plasma membrane which plays a critical role in preserving cell morphology and plasma membrane integrity. The aim of this study was to examine the effect of cold/warming exposure on apoptotic microtubules and plasma membrane integrity during the execution phase of apoptosis. We demonstrated in camptothecin-induced apoptotic H460 cells that cold/warming exposure disorganized apoptotic microtubules and allowed the access of active caspases to the cellular cortex and the cleavage of essential proteins in the preservation of plasma membrane permeability. Cleavage of cellular cortex and plasma membrane proteins, such as α-spectrin, paxilin, focal adhesion kinase and calcium ATPase pump (PMCA-4) involved in cell calcium extrusion resulted in increased plasma permeability and calcium overload leading apoptotic cells to secondary necrosis. The essential role of caspase-mediated cleavage in this process was demonstrated because the addition of the pan-caspase inhibitor z-VAD during cold/warming exposure that induces AMN depolymerization avoided the cleavage of cortical and plasma membrane proteins and prevented apoptotic cells to undergo secondary necrosis. Likewise, apoptotic microtubules stabilization by taxol during cold/warming exposure also prevented cellular cortex and plasma membrane protein cleavage and secondary necrosis. Furthermore, microtubules stabilization or caspase inhibition during cold/warming exposure was also critical for proper phosphatidylserine externalization and apoptotic cell clearance by macrophages. These results indicate that cold/warming exposure of apoptotic cells induces secondary necrosis which can be prevented by both, microtubule stabilization or caspase inhibition.

  16. Radiofrequency radiation-induced calcium ion efflux enhancement from human and other neuroblastoma cells in culture

    SciTech Connect

    Dutta, S.K.; Ghosh, B.; Blackman, C.F.

    1989-01-01

    To test the generality of radiofrequency radiation-induced changes in /sup 45/Ca2+ efflux from avian and feline brain tissues, human neuroblastoma cells were exposed to electromagnetic radiation at 147 MHz, amplitude-modulated (AM) at 16 Hz, at specific absorption rates (SAR) of 0.1, 0.05, 0.01, 0.005, 0.001, and 0.0005 W/kg. Significant /sup 45/Ca2+ efflux was obtained at SAR values of 0.05 and 0.005 W/kg. Enhanced efflux at 0.05 W/kg peaked at the 13-16 Hz and at the 57.5-60 Hz modulation ranges. A Chinese hamster-mouse hybrid neuroblastoma was also shown to exhibit enhanced radiation-induced /sup 45/Ca2+ efflux at an SAR of 0.05 W/kg, using 147 MHz, AM at 16 Hz. These results confirm that amplitude-modulated radiofrequency radiation can induce responses in cells of nervous tissue origin from widely different animal species, including humans. The results are also consistent with the reports of similar findings in avian and feline brain tissues and indicate the general nature of the phenomenon.

  17. Cell to Cell Variability of Radiation-Induced Foci: Relation between Observed Damage and Energy Deposition.

    PubMed

    Gruel, Gaëtan; Villagrasa, Carmen; Voisin, Pascale; Clairand, Isabelle; Benderitter, Marc; Bottollier-Depois, Jean-François; Barquinero, Joan Francesc

    2016-01-01

    Most studies that aim to understand the interactions between different types of photon radiation and cellular DNA assume homogeneous cell irradiation, with all cells receiving the same amount of energy. The level of DNA damage is therefore generally determined by averaging it over the entire population of exposed cells. However, evaluating the molecular consequences of a stochastic phenomenon such as energy deposition of ionizing radiation by measuring only an average effect may not be sufficient for understanding some aspects of the cellular response to this radiation. The variance among the cells associated with this average effect may also be important for the behaviour of irradiated tissue. In this study, we accurately estimated the distribution of the number of radiation-induced γH2AX foci (RIF) per cell nucleus in a large population of endothelial cells exposed to 3 macroscopic doses of gamma rays from 60Co. The number of RIF varied significantly and reproducibly from cell to cell, with its relative standard deviation ranging from 36% to 18% depending on the macroscopic dose delivered. Interestingly, this relative cell-to-cell variability increased as the dose decreased, contrary to the mean RIF count per cell. This result shows that the dose effect, in terms of the number of DNA lesions indicated by RIF is not as simple as a purely proportional relation in which relative SD is constant with dose. To analyse the origins of this observed variability, we calculated the spread of the specific energy distribution for the different target volumes and subvolumes in which RIF can be generated. Variances, standard deviations and relative standard deviations all changed similarly from dose to dose for biological and calculated microdosimetric values. This similarity is an important argument that supports the hypothesis of the conservation of the association between the number of RIF per nucleus and the specific energy per DNA molecule. This comparison allowed us to

  18. Sialylation of Integrin beta1 is Involved in Radiation-Induced Adhesion and Migration in Human Colon Cancer Cells

    SciTech Connect

    Lee, Minyoung; Lee, Hae-June; Seo, Woo Duck; Park, Ki Hun; Lee, Yun-Sil

    2010-04-15

    Purpose: Previously, we reported that radiation-induced ST6 Gal I gene expression was responsible for an increase of integrin beta1 sialylation. In this study, we have further investigated the function of radiation-mediated integrin beta1 sialylation in colon cancer cells. Methods and Materials: We performed Western blotting and lectin affinity assay to analyze the expression and level of sialylated integrin beta1. After exposure to ionizing radiation (IR), adhesion and migration of cells were measured by in vitro adhesion and migration assay. Results: IR increased sialylation of integrin beta1 responsible for its increased protein stability and adhesion and migration of colon cancer cells. However, for cells with an N-glycosylation site mutant of integrin beta1 located on the I-like domain (Mu3), these effects were dramatically inhibited. In addition, integrin beta1-mediated radioresistance was not observed in cells containing this mutant. When sialylation of integrin beta1 was targeted with a sulfonamide chalcone compound, inhibition of radiation-induced sialylation of integrin beta1 and inhibition of radiation-induced adhesion and migration occurred. Conclusion: The increase of integrin beta1 sialylation by ST6 Gal I is critically involved in radiation-mediated adhesion and migration of colon cancer cells. From these findings, integrin beta1 sialylation may be a novel target for overcoming radiation-induced survival, especially radiation-induced adhesion and migration.

  19. Dying Cells Protect Survivors from Radiation-Induced Cell Death in Drosophila

    PubMed Central

    Bilak, Amber; Uyetake, Lyle; Su, Tin Tin

    2014-01-01

    We report a phenomenon wherein induction of cell death by a variety of means in wing imaginal discs of Drosophila larvae resulted in the activation of an anti-apoptotic microRNA, bantam. Cells in the vicinity of dying cells also become harder to kill by ionizing radiation (IR)-induced apoptosis. Both ban activation and increased protection from IR required receptor tyrosine kinase Tie, which we identified in a genetic screen for modifiers of ban. tie mutants were hypersensitive to radiation, and radiation sensitivity of tie mutants was rescued by increased ban gene dosage. We propose that dying cells activate ban in surviving cells through Tie to make the latter cells harder to kill, thereby preserving tissues and ensuring organism survival. The protective effect we report differs from classical radiation bystander effect in which neighbors of irradiated cells become more prone to death. The protective effect also differs from the previously described effect of dying cells that results in proliferation of nearby cells in Drosophila larval discs. If conserved in mammals, a phenomenon in which dying cells make the rest harder to kill by IR could have implications for treatments that involve the sequential use of cytotoxic agents and radiation therapy. PMID:24675716

  20. Dying cells protect survivors from radiation-induced cell death in Drosophila.

    PubMed

    Bilak, Amber; Uyetake, Lyle; Su, Tin Tin

    2014-03-01

    We report a phenomenon wherein induction of cell death by a variety of means in wing imaginal discs of Drosophila larvae resulted in the activation of an anti-apoptotic microRNA, bantam. Cells in the vicinity of dying cells also become harder to kill by ionizing radiation (IR)-induced apoptosis. Both ban activation and increased protection from IR required receptor tyrosine kinase Tie, which we identified in a genetic screen for modifiers of ban. tie mutants were hypersensitive to radiation, and radiation sensitivity of tie mutants was rescued by increased ban gene dosage. We propose that dying cells activate ban in surviving cells through Tie to make the latter cells harder to kill, thereby preserving tissues and ensuring organism survival. The protective effect we report differs from classical radiation bystander effect in which neighbors of irradiated cells become more prone to death. The protective effect also differs from the previously described effect of dying cells that results in proliferation of nearby cells in Drosophila larval discs. If conserved in mammals, a phenomenon in which dying cells make the rest harder to kill by IR could have implications for treatments that involve the sequential use of cytotoxic agents and radiation therapy.

  1. Ionizing radiation-induced instant pairing of heterochromatin of homologous chromosomes in human cells.

    PubMed

    Abdel-Halim, H I; Imam, S A; Badr, F M; Natarajan, A T; Mullenders, L H F; Boei, J J W A

    2004-01-01

    Using fluorescence in situ hybridization with human band-specific DNA probes we examined the effect of ionizing radiation on the intra-nuclear localization of the heterochromatic region 9q12-->q13 and the euchromatic region 8p11.2 of similar sized chromosomes 9 and 8 respectively in confluent (G1) primary human fibroblasts. Microscopic analysis of the interphase nuclei revealed colocalization of the homologous heterochromatic regions from chromosome 9 in a proportion of cells directly after exposure to 4 Gy X-rays. The percentage of cells with paired chromosomes 9 gradually decreased to control levels during a period of one hour. No significant changes in localization were observed for chromosome 8. Using 2-D image analysis, radial and inter-homologue distances were measured for both chromosome bands. In unexposed cells, a random distribution of the chromosomes over the interphase nucleus was found. Directly after irradiation, the average inter-homologue distance decreased for chromosome 9 without alterations in radial distribution. The percentage of cells with inter-homologue distance <3 micro m increased from 11% in control cells to 25% in irradiated cells. In contrast, irradiation did not result in significant changes in the inter-homologue distance for chromosome 8. Colocalization of the heterochromatic regions of homologous chromosomes 9 was not observed in cells irradiated on ice. This observation, together with the time dependency of the colocalization, suggests an underlying active cellular process. The biological relevance of the observed homologous pairing remains unclear. It might be related to a homology dependent repair process of ionizing radiation induced DNA damage that is specific for heterochromatin. However, also other more general cellular responses to radiation-induced stress or change in chromatin organization might be responsible for the observed pairing of heterochromatic regions.

  2. The suppression of radiation-induced NF-{kappa}B activity by dexamethasone correlates with increased cell death in vivo

    SciTech Connect

    Nam, Seon Young; Chung, Hee-Yong . E-mail: hychung@hanyang.ac.kr

    2005-10-21

    In this study, we show that dexamethasone treatment increases ionizing radiation-induced cell death by inducing the inhibitory {kappa}B{alpha} (I{kappa}B{alpha}) pathway in mice. The effect of dexamethasone on radiation-induced cell death was assessed by changes in total spleen cellularity and bone marrow colony-forming unit-granulocyte-macrophage (CFU-GM) contents after total body irradiation. While in vivo treatment of mice with dexamethasone alone (1 mg/kg/day, for 2 days) failed to elicit cell death in spleen cells, the combined treatment with dexamethasone (1 mg/kg/day, for 2 days) and {gamma}-rays (1 or 5 Gy) caused a 50-80% reduction in total cellularity in spleen and CFU-GM contents in bone marrow. These results demonstrate that dexamethasone has a synergistic effect on radiation-induced cellular damages in vivo. Immunoblot analysis showed that dexamethasone treatment significantly increases I{kappa}B{alpha} expression in the spleens of irradiated mice. In addition, the dexamethasone treatment significantly reduced radiation-induced nuclear translocation of the nucleus factor-{kappa}B in the spleens of irradiated mice. These results indicate that dexamethasone treatment in vivo may increase radiation-induced cell damages by increasing I{kappa}B{alpha} expression in hematopoietic organs such as spleen and bone marrow.

  3. Innate recognition of apoptotic cells: novel apoptotic cell-associated molecular patterns revealed by crossreactivity of anti-LPS antibodies.

    PubMed

    Tennant, I; Pound, J D; Marr, L A; Willems, J J L P; Petrova, S; Ford, C A; Paterson, M; Devitt, A; Gregory, C D

    2013-05-01

    Cells dying by apoptosis are normally cleared by phagocytes through mechanisms that can suppress inflammation and immunity. Molecules of the innate immune system, the pattern recognition receptors (PRRs), are able to interact not only with conserved structures on microbes (pathogen-associated molecular patterns, PAMPs) but also with ligands displayed by apoptotic cells. We reasoned that PRRs might therefore interact with structures on apoptotic cells - apoptotic cell-associated molecular patterns (ACAMPs) - that are analogous to PAMPs. Here we show that certain monoclonal antibodies raised against the prototypic PAMP, lipopolysaccharide (LPS), can crossreact with apoptotic cells. We demonstrate that one such antibody interacts with a constitutively expressed intracellular protein, laminin-binding protein, which translocates to the cell surface during apoptosis and can interact with cells expressing the prototypic PRR, mCD14 as well as with CD14-negative cells. Anti-LPS cross reactive epitopes on apoptotic cells colocalised with annexin V- and C1q-binding sites on vesicular regions of apoptotic cell surfaces and were released associated with apoptotic cell-derived microvesicles (MVs). These results confirm that apoptotic cells and microbes can interact with the immune system through common elements and suggest that anti-PAMP antibodies could be used strategically to characterise novel ACAMPs associated not only with apoptotic cells but also with derived MVs.

  4. Comparative ultrastructure analysis of radiation-induced radioresistant laryngeal cancer hep-2 cell line.

    PubMed

    Yang, Bo; Tang, Fuqiu; Zhang, Bicheng; Zhao, Yong; Ding, Shifang; Rao, Zhiguo

    2014-08-01

    Radioresistance is one of the main reasons for the failure of radiotherapy in laryngeal cancer. However, the mechanisms of radioresistance of tumor cells have remained elusive. This study was conducted to identify the ultrastructural changes of radiation-induced radioresistant laryngeal cancer hep-2 cell line. First, a radioresistant hep-2R cell line was generated after prolonged exposure to γ-rays for 60 Gy (6 Gy/day, 2 days/week) and was confirmed by clonogenic assay. Next, the ultrastructural differences between hep-2R cells and hep-2 cells were compared by transmission electron microscopy. Finally, the results showed that hep-2R cells showed significant resistance to radiation compared with parental hep-2 cells. Increased cell nucleus atypia, more rough endoplasmic reticulum and less mitochondria were observed in hep-2R cells. The amount of microvilli of hep-2R was similar to hep-2 cell. In summary, these ultrastructural differences revealed the morphological mechanism that hep-2R cells had stronger radioresistance than hep-2 cells.

  5. Therapy with Multipotent Mesenchymal Stromal Cells Protects Lungs from Radiation-Induced Injury and Reduces the Risk of Lung Metastasis.

    PubMed

    Klein, Diana; Schmetter, Alexandra; Imsak, Roze; Wirsdörfer, Florian; Unger, Kristian; Jastrow, Holger; Stuschke, Martin; Jendrossek, Verena

    2016-01-10

    Previous thorax irradiation promotes metastatic spread of tumor cells to the lung. We hypothesized that vascular damage facilitates lung metastasis after thorax irradiation and that therapeutically applied multipotent mesenchymal stromal cells (MSCs) with reported repair activity may prevent these adverse effects of ionizing radiation by protecting lung endothelia from radiation-induced damage. Previous whole-thorax irradiation (WTI) with 15 Gy significantly enhanced seeding and metastatic growth of tumor cells in the lung. WTI was further associated with endothelial cell damage, senescence of lung epithelial cells, and upregulation of invasion- and inflammation-promoting soluble factors, for example, endothelial matrix metalloproteinase 2 (Mmp2), its activator Mmp14, the cofactor tissue inhibitor of metalloproteinases 2 (Timp2), chemokine (C-C motif) ligand 2 (Ccl2), and urokinase-type plasminogen activator (Plau/uPA), and recruitment of CD11b+CD11c- myelomonocytic cells. Inhibition of Mmp2 counteracted radiation-induced vascular dysfunction without preventing increased metastasis. In contrast, therapy with bone marrow or aorta-derived MSCs within 2 weeks postirradiation antagonized radiation-induced damage to resident cells as well as the resulting secretome changes and abrogated the metastasis-promoting effects of WTI. Therapy with MSCs protects lungs from radiation-induced injury and reduces the risk of lung metastasis. MSC-mediated inhibition of Mmp2 mediates their protective effects at the vasculature. Furthermore, local and systemic effects such as inhibition of radiation-induced senescence of bronchial epithelial cells and associated secretion of immunomodulatory factors may participate in the inhibitory effect of MSCs on lung metastasis. MSC therapy is a promising strategy to prevent radiation-induced lung injury and the resulting increased risk of metastasis.

  6. Radiation-inducible immunotherapy for cancer: senescent tumor cells as a cancer vaccine.

    PubMed

    Meng, Yuru; Efimova, Elena V; Hamzeh, Khaled W; Darga, Thomas E; Mauceri, Helena J; Fu, Yang-Xin; Kron, Stephen J; Weichselbaum, Ralph R

    2012-05-01

    Radiotherapy offers an effective treatment for advanced cancer but local and distant failures remain a significant challenge. Here, we treated melanoma and pancreatic carcinoma in syngeneic mice with ionizing radiation (IR) combined with the poly(ADP-ribose) polymerase inhibitor (PARPi) veliparib to inhibit DNA repair and promote accelerated senescence. Based on prior work implicating cytotoxic T lymphocytes (CTLs) as key mediators of radiation effects, we discovered that senescent tumor cells induced by radiation and veliparib express immunostimulatory cytokines to activate CTLs that mediate an effective antitumor response. When these senescent tumor cells were injected into tumor-bearing mice, an antitumor CTL response was induced which potentiated the effects of radiation, resulting in elimination of established tumors. Applied to human cancers, radiation-inducible immunotherapy may enhance radiotherapy responses to prevent local recurrence and distant metastasis.

  7. HGF Gene Modification in Mesenchymal Stem Cells Reduces Radiation-Induced Intestinal Injury by Modulating Immunity

    PubMed Central

    Li, Yang; Yang, Yue-Feng; Xiao, Feng-Jun; Zhang, Yi-Kun; Wang, Shao-Xia; Sun, Hui-Yan; Zhang, Qun-Wei; Wu, Chu-Tse; Wang, Li-Sheng

    2015-01-01

    Background Effective therapeutic strategies to address intestinal complications after radiation exposure are currently lacking. Mesenchymal stem cells (MSCs), which display the ability to repair the injured intestine, have been considered as delivery vehicles for repair genes. In this study, we evaluated the therapeutic effect of hepatocyte growth factor (HGF)-gene-modified MSCs on radiation-induced intestinal injury (RIII). Methods Female 6- to 8-week-old mice were radiated locally at the abdomen with a single 13-Gy dose of radiation and then treated with saline control, Ad-HGF or Ad-Null-modified MSCs therapy. The transient engraftment of human MSCs was detected via real-time PCR and immunostaining. The therapeutic effects of non- and HGF-modified MSCs were evaluated via FACS to determine the lymphocyte immunophenotypes; via ELISA to measure cytokine expression; via immunostaining to determine tight junction protein expression; via PCNA staining to examine intestinal epithelial cell proliferation; and via TUNEL staining to detect intestinal epithelial cell apoptosis. Results The histopathological recovery of the radiation-injured intestine was significantly enhanced following non- or HGF-modified MSCs treatment. Importantly, the radiation-induced immunophenotypic disorders of the mesenteric lymph nodes and Peyer’s patches were attenuated in both MSCs-treated groups. Treatment with HGF-modified MSCs reduced the expression and secretion of inflammatory cytokines, including tumor necrosis factor alpha (TNF-α) and interferon-gamma (IFN-γ), increased the expression of the anti-inflammatory cytokine IL-10 and the tight junction protein ZO-1, and promoted the proliferation and reduced the apoptosis of intestinal epithelial cells. Conclusions Treatment of RIII with HGF-gene-modified MSCs reduces local inflammation and promotes the recovery of small intestinal histopathology in a mouse model. These findings might provide an effective therapeutic strategy for RIII

  8. HGF Gene Modification in Mesenchymal Stem Cells Reduces Radiation-Induced Intestinal Injury by Modulating Immunity.

    PubMed

    Wang, Hua; Sun, Rui-Ting; Li, Yang; Yang, Yue-Feng; Xiao, Feng-Jun; Zhang, Yi-Kun; Wang, Shao-Xia; Sun, Hui-Yan; Zhang, Qun-Wei; Wu, Chu-Tse; Wang, Li-Sheng

    2015-01-01

    Effective therapeutic strategies to address intestinal complications after radiation exposure are currently lacking. Mesenchymal stem cells (MSCs), which display the ability to repair the injured intestine, have been considered as delivery vehicles for repair genes. In this study, we evaluated the therapeutic effect of hepatocyte growth factor (HGF)-gene-modified MSCs on radiation-induced intestinal injury (RIII). Female 6- to 8-week-old mice were radiated locally at the abdomen with a single 13-Gy dose of radiation and then treated with saline control, Ad-HGF or Ad-Null-modified MSCs therapy. The transient engraftment of human MSCs was detected via real-time PCR and immunostaining. The therapeutic effects of non- and HGF-modified MSCs were evaluated via FACS to determine the lymphocyte immunophenotypes; via ELISA to measure cytokine expression; via immunostaining to determine tight junction protein expression; via PCNA staining to examine intestinal epithelial cell proliferation; and via TUNEL staining to detect intestinal epithelial cell apoptosis. The histopathological recovery of the radiation-injured intestine was significantly enhanced following non- or HGF-modified MSCs treatment. Importantly, the radiation-induced immunophenotypic disorders of the mesenteric lymph nodes and Peyer's patches were attenuated in both MSCs-treated groups. Treatment with HGF-modified MSCs reduced the expression and secretion of inflammatory cytokines, including tumor necrosis factor alpha (TNF-α) and interferon-gamma (IFN-γ), increased the expression of the anti-inflammatory cytokine IL-10 and the tight junction protein ZO-1, and promoted the proliferation and reduced the apoptosis of intestinal epithelial cells. Treatment of RIII with HGF-gene-modified MSCs reduces local inflammation and promotes the recovery of small intestinal histopathology in a mouse model. These findings might provide an effective therapeutic strategy for RIII.

  9. Methylglyoxal-bis(guanylhydrazone), a polyamine analogue, sensitized γ-radiation-induced cell death in HL-60 leukemia cells Sensitizing effect of MGBG on γ-radiation-induced cell death.

    PubMed

    Kim, Jin Sik; Lee, Jin; Chung, Hai Won; Choi, Han; Paik, Sang Gi; Kim, In Gyu

    2006-09-01

    Methylglyoxal-bis(guanylhydrazone) (MGBG), a polyamine analogue, has been known to inhibit the biosynthesis of polyamines, which are important in cell proliferation. We showed that MGBG treatment significantly affected γ-radiation-induced cell cycle transition (G(1)/G(0)→S→G(2)/M) and thus γ-radiation-induced cell death. As determined by micronuclei and comet assay, we showed that it sensitized the cytotoxic effect induced by γ-radiation. One of the reasons is that polyamine depletion by MGBG treatment did not effectively protect against the chemical (OH) or physical damage to DNA caused by γ-radiation. Through in vitro experiment, we confirmed that DNA strand breaks induced by γ-radiation was prevented more effectively in the presence of polyamines (spermine and spermidine) than in the absence of polyamines. MGBG also blocks the cell cycle transition caused by γ-radiation (G(2) arrest), which helps protect cells by allowing time for DNA repair before entry into mitosis or apoptosis, via the down regulation of cyclin D1, which mediates the transition from G(1) to S phase of cell cycle, and ataxia telangiectasia mutated, which is involved in the DNA sensing, repair and cell cycle check point. Therefore, the abrogation of G(2) arrest sensitizes cells to the effect of γ-radiation. As a result, γ-radiation-induced cell death increased by about 2.5-3.0-fold in cells treated with MGBG. However, exogenous spermidine supplement partially relieved this γ-radiation-induced cytotoxicity and cell death. These findings suggest a potentially therapeutic strategy for increasing the cytotoxic efficacy of γ-radiation.

  10. Organotypic culture in three dimensions prevents radiation-induced transformation in human lung epithelial cells

    NASA Astrophysics Data System (ADS)

    El-Ashmawy, Mariam; Coquelin, Melissa; Luitel, Krishna; Batten, Kimberly; Shay, Jerry W.

    2016-08-01

    The effects of radiation in two-dimensional (2D) cell culture conditions may not recapitulate tissue responses as modeled in three-dimensional (3D) organotypic culture. In this study, we determined if the frequency of radiation-induced transformation and cancer progression differed in 3D compared to 2D culture. Telomerase immortalized human bronchial epithelial cells (HBECs) with shTP53 and mutant KRas expression were exposed to various types of radiation (gamma, +H, 56Fe) in either 2D or 3D culture. After irradiation, 3D structures were dissociated and passaged as a monolayer followed by measurement of transformation, cell growth and expression analysis. Cells irradiated in 3D produced significantly fewer and smaller colonies in soft agar than their 2D-irradiated counterparts (gamma P = 0.0004 +H P = 0.049 56Fe P < 0.0001). The cell culture conditions did not affect cell killing, the ability of cells to survive in a colony formation assay, and proliferation rates after radiation—implying there was no selection against cells in or dissociated from 3D conditions. However, DNA damage repair and apoptosis markers were increased in 2D cells compared to 3D cells after radiation. Ideally, expanding the utility of 3D culture will allow for a better understanding of the biological consequences of radiation exposure.

  11. Organotypic culture in three dimensions prevents radiation-induced transformation in human lung epithelial cells

    PubMed Central

    El-Ashmawy, Mariam; Coquelin, Melissa; Luitel, Krishna; Batten, Kimberly; Shay, Jerry W.

    2016-01-01

    The effects of radiation in two-dimensional (2D) cell culture conditions may not recapitulate tissue responses as modeled in three-dimensional (3D) organotypic culture. In this study, we determined if the frequency of radiation-induced transformation and cancer progression differed in 3D compared to 2D culture. Telomerase immortalized human bronchial epithelial cells (HBECs) with shTP53 and mutant KRas expression were exposed to various types of radiation (gamma, +H, 56Fe) in either 2D or 3D culture. After irradiation, 3D structures were dissociated and passaged as a monolayer followed by measurement of transformation, cell growth and expression analysis. Cells irradiated in 3D produced significantly fewer and smaller colonies in soft agar than their 2D-irradiated counterparts (gamma P = 0.0004; +H P = 0.049; 56Fe P < 0.0001). The cell culture conditions did not affect cell killing, the ability of cells to survive in a colony formation assay, and proliferation rates after radiation—implying there was no selection against cells in or dissociated from 3D conditions. However, DNA damage repair and apoptosis markers were increased in 2D cells compared to 3D cells after radiation. Ideally, expanding the utility of 3D culture will allow for a better understanding of the biological consequences of radiation exposure. PMID:27539227

  12. 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.

  13. Acute high-dose X-radiation-induced genomic changes in A549 cells.

    PubMed

    Muradyan, A; Gilbertz, K; Stabentheiner, S; Klause, S; Madle, H; Meineke, V; Ullmann, R; Scherthan, H

    2011-06-01

    Accidents with ionizing radiation often involve single, acute high-dose exposures that can lead to acute radiation syndrome and late effects such as carcinogenesis. To study such effects at the cellular level, we investigated acute ionizing radiation-induced chromosomal aberrations in A549 adenocarcinoma cells at the genome-wide level by exposing the cells to an acute dose of 6 Gy 240 kV X rays. One sham-irradiated clone and four surviving irradiated clones were recovered by minimal dilution and further expanded and analyzed by chromosome painting and tiling-path array CGH, with the nonirradiated clone 0 serving as the control. Acute X-ray exposure induced specific translocations and changes in modal chromosome number in the four irradiated clones. Array CGH disclosed unique and recurrent genomic changes, predominantly losses, and revealed that the fragile sites FRA3B and FRA16D were preferential regions of genomic alterations in all irradiated clones, which is likely related to radioresistant S-phase progression and genomic stress. Furthermore, clone 4 displayed an increased radiosensitivity at doses >5 Gy. Pairwise comparisons of the gene expression patterns of all irradiated clones to the sham-irradiated clone 0 revealed an enrichment of the Gene Ontology term "M Phase" (P = 6.2 × 10(-7)) in the set of differentially expressed genes of clone 4 but not in those of clones 1-3. Ionizing radiation-induced genomic changes and fragile site expression highlight the capacity of a single acute radiation exposure to affect the genome of exposed cells by inflicting genomic stress.

  14. Nimotuzumab Enhances the Radiosensitivity of Cancer Cells In Vitro by Inhibiting Radiation-Induced DNA Damage Repair

    PubMed Central

    Qu, Yuan-yuan; Hu, Song-liu; Xu, Xiang-ying; Wang, Rui-zhi; Yu, Hong-yang; Xu, Jian-yu; Chen, Lin; Dong, Guang-lu

    2013-01-01

    Background Nimotuzumab is a humanized IgG1 monoclonal antibody specifically targeting EGFR. In this study, we aimed to investigate the molecular mechanisms of nimotuzumab in its effects of enhancing cancer cell radiosensitivity. Principal Finding Lung cancer A549 cells and breast cancer MCF-7 cells were pretreated with or without nimotuzumab for 24 h before radiation to perform the clonogenic survival assay and to analyze the cell apoptosis by flow ctyometry. γ-H2AX foci were detected by confocal microscopy to assess the effect of nimotuzumab on radiation induced DNA repair. EGFR activation was examined and the levels of DNA damage repair related proteins in A549 cells at different time point and at varying doses exposure after nimotuzumab and radiation treatment were examined by Western blot. Pretreatment with nimotuzumab reduced clonogenic survival after radiation, inhibited radiation-induced EGFR activation and increased the radiation-induced apoptosis in both A549 cells and MCF-7 cells. The foci of γ-H2AX 24 h after radiation significantly increased in nimotuzumab pretreated cells with different doses. The phosphorylation of AKT and DNA-PKcs were remarkably inhibited in the combination group at each dose point as well as time point. Conclusions Our results revealed that the possible mechanism of nimotuzumab enhancing the cancer radiosensitivity is that nimotuzumab inhibited the radiation-induced activation of DNA-PKcs through blocking the PI3K/AKT pathway, which ultimately affected the DNA DSBs repair. PMID:23976954

  15. Nimotuzumab enhances the radiosensitivity of cancer cells in vitro by inhibiting radiation-induced DNA damage repair.

    PubMed

    Qu, Yuan-yuan; Hu, Song-liu; Xu, Xiang-ying; Wang, Rui-zhi; Yu, Hong-yang; Xu, Jian-yu; Chen, Lin; Dong, Guang-lu

    2013-01-01

    Nimotuzumab is a humanized IgG1 monoclonal antibody specifically targeting EGFR. In this study, we aimed to investigate the molecular mechanisms of nimotuzumab in its effects of enhancing cancer cell radiosensitivity. Lung cancer A549 cells and breast cancer MCF-7 cells were pretreated with or without nimotuzumab for 24 h before radiation to perform the clonogenic survival assay and to analyze the cell apoptosis by flow ctyometry. γ-H2AX foci were detected by confocal microscopy to assess the effect of nimotuzumab on radiation induced DNA repair. EGFR activation was examined and the levels of DNA damage repair related proteins in A549 cells at different time point and at varying doses exposure after nimotuzumab and radiation treatment were examined by Western blot. Pretreatment with nimotuzumab reduced clonogenic survival after radiation, inhibited radiation-induced EGFR activation and increased the radiation-induced apoptosis in both A549 cells and MCF-7 cells. The foci of γ-H2AX 24 h after radiation significantly increased in nimotuzumab pretreated cells with different doses. The phosphorylation of AKT and DNA-PKcs were remarkably inhibited in the combination group at each dose point as well as time point. Our results revealed that the possible mechanism of nimotuzumab enhancing the cancer radiosensitivity is that nimotuzumab inhibited the radiation-induced activation of DNA-PKcs through blocking the PI3K/AKT pathway, which ultimately affected the DNA DSBs repair.

  16. Gamma radiation induced cell cycle perturbations and DNA damage in Catla Catla as measured by flow cytometry.

    PubMed

    Anbumani, S; Mohankumar, Mary N

    2015-03-01

    Gamma radiation induced cell cycle perturbations and DNA damage in Catla catla were analyzed in erythrocytes at different time points using flow cytometry (FCM). Protracted exposure to radiation induced damage between days 12 and 45. Disturbances in cell cycle machinery, i.e., proportional increase and decrease in Gap0 or quiescent/Gap1 (G0/G1), Synthesis (S) and Gap2/Mitotic (G2/M) phases were observed at both acute and protracted treatments. Both acute and protracted exposures induced apoptosis with a notable significance between days 3 and 6 at protracted and on day 45 at acute doses. Fish exposed protractedly avail some DNA repair mechanisms than acutely exposed. This is the first study to analyze radiation induced DNA damage under laboratory conditions and suggests that flow cytometry can also be an alternate tool to screen genotoxicity induced by ionizing radiation in fish. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. A Human Espophageal Epithelial Cell Model for Study of Radiation Induced Cancer and DNA Damage Repair

    NASA Technical Reports Server (NTRS)

    Huff, Janice L.; Patel, Zarana S.; Hada, Megumi; Cucinotta, Francis A.

    2008-01-01

    For cancer risk assessment in astronauts and for countermeasure development, it is essential to understand the molecular mechanisms of radiation carcinogenesis and how these mechanisms are influenced by exposure to the types of radiation found in space. We are developing an in vitro model system for the study of radiation-induced initiation and progression of esophageal carcinoma, a type of cancer found to have a significant enhancement in incidence in the survivors of the atomic bomb detonations in Japan. Here we present the results of our preliminary characterization of both normal and hTERT immortalized esophageal epithelial cells grown in 2-dimensional culture. We analyzed DNA repair capacity by measuring the kinetics of formation and loss of - H2AX foci following radiation exposure. Additionally, we analyzed induction of chromosomal aberrations using 3-color fluorescence in situ hybridization (FISH). Data were generated using both low LET (gamma rays) and high LET ions (1000 MeV/nucleon iron).

  18. A Human Espophageal Epithelial Cell Model for Study of Radiation Induced Cancer and DNA Damage Repair

    NASA Technical Reports Server (NTRS)

    Huff, Janice L.; Patel, Zarana S.; Hada, Megumi; Cucinotta, Francis A.

    2008-01-01

    For cancer risk assessment in astronauts and for countermeasure development, it is essential to understand the molecular mechanisms of radiation carcinogenesis and how these mechanisms are influenced by exposure to the types of radiation found in space. We are developing an in vitro model system for the study of radiation-induced initiation and progression of esophageal carcinoma, a type of cancer found to have a significant enhancement in incidence in the survivors of the atomic bomb detonations in Japan. Here we present the results of our preliminary characterization of both normal and hTERT immortalized esophageal epithelial cells grown in 2-dimensional culture. We analyzed DNA repair capacity by measuring the kinetics of formation and loss of - H2AX foci following radiation exposure. Additionally, we analyzed induction of chromosomal aberrations using 3-color fluorescence in situ hybridization (FISH). Data were generated using both low LET (gamma rays) and high LET ions (1000 MeV/nucleon iron).

  19. Polymeric nanocomposite proton exchange membranes prepared by radiation-induced polymerization for direct methanol fuel cell

    NASA Astrophysics Data System (ADS)

    Kim, Young-Seok; Seo, Kwang-Seok; Choi, Seong-Ho

    2016-01-01

    The vinyl group-modified montmorillonite clay (F-MMT), vinyl group-modified graphene oxide (F-GO), and vinyl group-modified multi-walled carbon nanotube (F-MWNT) were first prepared by ion exchange reaction of 1-[(4-ethylphenyl)methyl]-3-butyl-imidazolium chloride in order to use the materials for protection against methanol cross-over in direct methanol fuel cell (DMFC) membrane. Then polymeric nanocomposite membranes with F-MMT, F-GO, and F-MWNT were prepared by the solvent casting method after radiation-induced polymerization of vinyl monomers in water-methanol mixture solvents. The proton conductivity, water uptake, ion-exchange capacity, methanol permeability, and DMFC performance of the polymeric nanocomposite membranes with F-MMT, F-GO, and F-MWNT were evaluated.

  20. Repair of radiation-induced damage to the cell division mechanism of Escherichia coli.

    PubMed

    Adler, H I; Fisher, W D; Hardigree, A A; Stapleton, G E

    1966-02-01

    Adler, Howard I. (Oak Ridge National Laboratory, Oak Ridge, Tenn.), William D. Fisher, Alice A. Hardigree, and George E. Stapleton. Repair of radiation-induced damage to the cell division mechanism of Escherichia coli. J. Bacteriol. 91:737-742. 1966.-Microscopic observations of irradiated populations of filamentous Escherichia coli cells indicated that filaments can be induced to divide by a substance donated by neighboring cells. We have made this observation the basis for a quantitative technique in which filaments are incubated in the presence of nongrowing donor cells. The presence of "donor" organisms promotes division and subsequent colony formation in filaments. "Donor" bacteria do not affect nonfilamentous cells. An extract of "donor" cells retains the division-promoting activity. The extract has been partially fractionated, and consists of a heat-stable and a heat-labile component. The heat-stable component is inactive in promoting cell division, but enhances the activity of the heat-labile component. The division-promoting system is discussed as a radiation repair mechanism and as a normal component of the cell division system in E. coli.

  1. Role of ROS-mediated autophagy in radiation-induced bystander effect of hepatoma cells.

    PubMed

    Wang, Xiangdong; Zhang, Jianghong; Fu, Jiamei; Wang, Juan; Ye, Shuang; Liu, Weili; Shao, Chunlin

    2015-05-01

    Autophagy plays a crucial role in cellular response to ionizing radiation, but it is unclear whether autophagy can modulate radiation-induced bystander effect (RIBE). Here, we investigated the relationship between bystander damage and autophagy in human hepatoma cells of HepG2. HepG2 cells were treated with conditioned medium (CM) collected from 3 Gy γ-rays irradiated hepatoma HepG2 cells for 4, 12, or 24 h, followed by the measurement of micronuclei (MN), intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and protein expressions of microtubule-associated protein 1 light chain 3 (LC3) and Beclin-1 in the bystander HepG2 cells. In some experiments, the bystander HepG2 cells were respectively transfected with LC3 small interfering RNA (siRNA), Beclin-1 siRNA or treated with 1% dimethyl sulfoxide (DMSO). Additional MN and mitochondrial dysfunction coupled with ROS were induced in the bystander cells. The expressions of protein markers of autophagy, LC3-II/LC3-I and Beclin-1, increased in the bystander cells. The inductions of bystander MN and overexpressions of LC3 and Beclin-1 were significantly diminished by DMSO. However, when the bystander cells were transfected with LC3 siRNA or Beclin-1 siRNA, the yield of bystander MN was significantly enhanced. The elevated ROS have bi-functions in balancing the bystander effects. One is to cause MN and the other is to induce protective autophagy.

  2. Apoptotic regulation and mutagenesis in human cells exposes to charged particles of importance for spaceflight

    NASA Astrophysics Data System (ADS)

    Kronenberg, A.; Gauny, S.; Hain, J.; Wu, P.; Wiese, C.

    Exposure to ionizing radiation can elicit two modes of cell death - necrosis or apoptosis. In human lymphoid cells, the predominant mechanism of radiation- induced cell death is apoptosis. The most likely exposure of individual human cells to heavy ions (e.g. Fe or Si) during spaceflight will result from single particle traversals. Here we report the fluence-response for apoptosis in human TK6 B- lymp hoblasts and provide evidence that single Fe ion traversals can stimulate an apoptotic response. The apoptotic response to charged particle exposures includes scrambling of the phospholipid bilayer in the cell membrane, activation of caspase signaling cascades and degradation of DNA into oligonucleosomes. We have also explored the importance of apoptotic regulation on the frequency and spectrum of mutations arising after exposure to charged particles. We used isogenic derivatives of TK6 cells stably transfected with pSFFV-neo-bcl-xL (encoding the anti-apoptotic gene BCL-XL and the neomycin resistance gene) or with pSFFV neo (encoding only- the neomycin resistance gene). TK6-bclxL cells were more susceptible to mutations at the TK1 locus than TK6-neo cells following exposure to protons, silicon ions or Fe ions. Molecular analysis demonstrated that most Fe-ion-induced mutations arose by loss of heterozygosity (LOH). In TK6-bclxL cells, more of the LOH occurred via mitotic recombination than in TK6-neo cells where the predominant mode of LOH was via deletion. We are currently mapping the LOH tracts to further define the biological bases for the differential sensitivity to Fe-ion-induced mutagenesis as a function of the genotype of the cell at risk. Supported by NASA grant T-964W to A. Kronenberg

  3. Vascular endothelial growth factor enhances macrophage clearance of apoptotic cells

    PubMed Central

    Dalal, Samay; Horstmann, Sarah A.; Richens, Tiffany R.; Tanaka, Takeshi; Doe, Jenna M.; Boe, Darren M.; Voelkel, Norbert F.; Taraseviciene-Stewart, Laimute; Janssen, William J.; Lee, Chun G.; Elias, Jack A.; Bratton, Donna; Tuder, Rubin M.; Henson, Peter M.; Vandivier, R. William

    2012-01-01

    Efficient clearance of apoptotic cells from the lung by alveolar macrophages is important for the maintenance of tissue structure and function. Lung tissue from humans with emphysema contains increased numbers of apoptotic cells and decreased levels of vascular endothelial growth factor (VEGF). Mice treated with VEGF receptor inhibitors have increased numbers of apoptotic cells and develop emphysema. We hypothesized that VEGF regulates apoptotic cell clearance by alveolar macrophages (AM) via its interaction with VEGF receptor 1 (VEGF R1). Our data show that the uptake of apoptotic cells by murine AMs and human monocyte-derived macrophages is inhibited by depletion of VEGF and that VEGF activates Rac1. Antibody blockade or pharmacological inhibition of VEGF R1 activity also decreased apoptotic cell uptake ex vivo. Conversely, overexpression of VEGF significantly enhanced apoptotic cell uptake by AMs in vivo. These results indicate that VEGF serves a positive regulatory role via its interaction with VEGF R1 to activate Rac1 and enhance AM apoptotic cell clearance. PMID:22307908

  4. Repeated Autologous Bone Marrow-Derived Mesenchymal Stem Cell Injections Improve Radiation-Induced Proctitis in Pigs

    PubMed Central

    Busson, Elodie; Holler, Valerie; Strup-Perrot, Carine; Lacave-Lapalun, Jean-Victor; Lhomme, Bruno; Prat, Marie; Devauchelle, Patrick; Sabourin, Jean-Christophe; Simon, Jean-Marc; Bonneau, Michel; Lataillade, Jean-Jacques; Benderitter, Marc

    2013-01-01

    The management of proctitis in patients who have undergone very-high-dose conformal radiotherapy is extremely challenging. The fibrosis-necrosis, fistulae, and hemorrhage induced by pelvic overirradiation have an impact on morbidity. Augmenting tissue repair by the use of mesenchymal stem cells (MSCs) may be an important advance in treating radiation-induced toxicity. Using a preclinical pig model, we investigated the effect of autologous bone marrow-derived MSCs on high-dose radiation-induced proctitis. Irradiated pigs received repeated intravenous administrations of autologous bone marrow-derived MSCs. Immunostaining and real-time polymerase chain reaction analysis were used to assess the MSCs' effect on inflammation, extracellular matrix remodeling, and angiogenesis, in radiation-induced anorectal and colon damages. In humans, as in pigs, rectal overexposure induces mucosal damage (crypt depletion, macrophage infiltration, and fibrosis). In a pig model, repeated administrations of MSCs controlled systemic inflammation, reduced in situ both expression of inflammatory cytokines and macrophage recruitment, and augmented interleukin-10 expression in rectal mucosa. MSC injections limited radiation-induced fibrosis by reducing collagen deposition and expression of col1a2/col3a1 and transforming growth factor-β/connective tissue growth factor, and by modifying the matrix metalloproteinase/TIMP balance. In a pig model of proctitis, repeated injections of MSCs effectively reduced inflammation and fibrosis. This treatment represents a promising therapy for radiation-induced severe rectal damage. PMID:24068742

  5. Repeated autologous bone marrow-derived mesenchymal stem cell injections improve radiation-induced proctitis in pigs.

    PubMed

    Linard, Christine; Busson, Elodie; Holler, Valerie; Strup-Perrot, Carine; Lacave-Lapalun, Jean-Victor; Lhomme, Bruno; Prat, Marie; Devauchelle, Patrick; Sabourin, Jean-Christophe; Simon, Jean-Marc; Bonneau, Michel; Lataillade, Jean-Jacques; Benderitter, Marc

    2013-11-01

    The management of proctitis in patients who have undergone very-high-dose conformal radiotherapy is extremely challenging. The fibrosis-necrosis, fistulae, and hemorrhage induced by pelvic overirradiation have an impact on morbidity. Augmenting tissue repair by the use of mesenchymal stem cells (MSCs) may be an important advance in treating radiation-induced toxicity. Using a preclinical pig model, we investigated the effect of autologous bone marrow-derived MSCs on high-dose radiation-induced proctitis. Irradiated pigs received repeated intravenous administrations of autologous bone marrow-derived MSCs. Immunostaining and real-time polymerase chain reaction analysis were used to assess the MSCs' effect on inflammation, extracellular matrix remodeling, and angiogenesis, in radiation-induced anorectal and colon damages. In humans, as in pigs, rectal overexposure induces mucosal damage (crypt depletion, macrophage infiltration, and fibrosis). In a pig model, repeated administrations of MSCs controlled systemic inflammation, reduced in situ both expression of inflammatory cytokines and macrophage recruitment, and augmented interleukin-10 expression in rectal mucosa. MSC injections limited radiation-induced fibrosis by reducing collagen deposition and expression of col1a2/col3a1 and transforming growth factor-β/connective tissue growth factor, and by modifying the matrix metalloproteinase/TIMP balance. In a pig model of proctitis, repeated injections of MSCs effectively reduced inflammation and fibrosis. This treatment represents a promising therapy for radiation-induced severe rectal damage.

  6. Radiation-induced bystander effects enhanced by elevated sodium chloride through sensitizing cells to bystander factors.

    PubMed

    Zhu, Lingyan; Han, Wei; Chen, Shaopeng; Zhao, Ye; Jiang, Erkang; Bao, Lingzhi; Pei, Bei; Yang, Gen; Zhao, Guoping; Wang, Jun; Xu, An; Wu, Lijun

    2008-09-26

    Radiation-induced bystander effects (RIBE) have been demonstrated to occur widely in various cell lines. However, very little data is available on the genotoxic effects of RIBE combined with other factor(s). We reported previously that with a low dose of alpha-particle irradiation, the fraction of gamma-H2AX foci-positive cells in non-irradiated bystander cells was significantly increased under elevated NaCl culture conditions. In this study, we further investigated the functional role of NaCl in the enhancement of RIBE using a specially designed co-culture system and micronucleus (MN) test. It was shown that the MN frequency was not increased significantly by elevated NaCl (9.0 g/L) alone or by medium exposure. However, with 1.0 cGy alpha-particle irradiation, the induced MN frequency increased significantly in both irradiated and non-irradiated bystander regions. Additional studies showed that elevated NaCl made the non-irradiated bystander cells more vulnerable to bystander factors. Furthermore, it was found that the induced MN frequency in cells both in irradiated and non-irradiated bystander regions was weakened when the hypertonic medium was changed to normotonic medium for 2h before irradiation. Such observations were quite similar to the co-effect of NaCl and hydrogen peroxide (H(2)O(2)), indicating that elevated NaCl might sensitize non-irradiated cells to bystander factors-induced oxidative stress.

  7. Relationship between the repair of radiation-induced DNA damage and recovery from potentially lethal damage in 9L rat brain tumor cells. [Gamma radiation

    SciTech Connect

    vanAnkeren, S.C.; Wheeler, K.T.

    1984-03-01

    The kinetics of repair of radiation-induced DNA damage and recovery from radiation-induced potentially lethal damage (PLD) for fed plateau-phase 9L/Ro rat brain tumor cells were compared after single doses of gamma-radiation and after combined treatment with 3 micrograms of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU)/ml given 16 hr prior to irradiation. DNA damage and repair were assayed using alkaline filter elution, while cell survival was assayed by colony formation. Repair of radiation-induced DNA damage and recovery from radiation-induced PLD followed statistically identical biphasic kinetics; the fast-phase half-times were 4.1 +/- 0.3 (S.D.) min and 4.0 +/- 0.8 min, while the slow-phase half-times were 59.7 +/- 11.2 min and 78.7 +/- 34.1 min, respectively. Treatment with BCNU prior to irradiation resulted in both additional DNA damage and increased cell kill. When DNA damage and cell survival after the combined treatment were corrected for the contribution from BCNU given alone, no inhibition of either repair of radiation-induced DNA damage or of recovery from radiation-induced PLD was observed. However, postirradiation hypertonic treatment inhibited both DNA repair and recovery from radiation-induced PLD. These correlations between the kinetics of the molecular and cellular repair processes support a role for repair of radiation-induced DNA damage in recovery from radiation-induced PLD. The lack of inhibition by BCNU of both repair of radiation-induced DNA damage and of recovery from radiation-induced PLD also demonstrates that these are not the mechanisms by which BCNU enhances radiation-induced cytotoxicity in 9L cells.

  8. Regulatory T Cells Promote β-Catenin–Mediated Epithelium-to-Mesenchyme Transition During Radiation-Induced Pulmonary Fibrosis

    SciTech Connect

    Xiong, Shanshan; Pan, Xiujie; Xu, Long; Yang, Zhihua; Guo, Renfeng; Gu, Yongqing; Li, Ruoxi; Wang, Qianjun; Xiao, Fengjun; Du, Li; Zhou, Pingkun; Zhu, Maoxiang

    2015-10-01

    Purpose: Radiation-induced pulmonary fibrosis results from thoracic radiation therapy and severely limits radiation therapy approaches. CD4{sup +}CD25{sup +}FoxP3{sup +} regulatory T cells (Tregs) as well as epithelium-to-mesenchyme transition (EMT) cells are involved in pulmonary fibrosis induced by multiple factors. However, the mechanisms of Tregs and EMT cells in irradiation-induced pulmonary fibrosis remain unclear. In the present study, we investigated the influence of Tregs on EMT in radiation-induced pulmonary fibrosis. Methods and Materials: Mice thoraxes were irradiated (20 Gy), and Tregs were depleted by intraperitoneal injection of a monoclonal anti-CD25 antibody 2 hours after irradiation and every 7 days thereafter. Mice were treated on days 3, 7, and 14 and 1, 3, and 6 months post irradiation. The effectiveness of Treg depletion was assayed via flow cytometry. EMT and β-catenin in lung tissues were detected by immunohistochemistry. Tregs isolated from murine spleens were cultured with mouse lung epithelial (MLE) 12 cells, and short interfering RNA (siRNA) knockdown of β-catenin in MLE 12 cells was used to explore the effects of Tregs on EMT and β-catenin via flow cytometry and Western blotting. Results: Anti-CD25 antibody treatment depleted Tregs efficiently, attenuated the process of radiation-induced pulmonary fibrosis, hindered EMT, and reduced β-catenin accumulation in lung epithelial cells in vivo. The coculture of Tregs with irradiated MLE 12 cells showed that Tregs could promote EMT in MLE 12 cells and that the effect of Tregs on EMT was partially abrogated by β-catenin knockdown in vitro. Conclusions: Tregs can promote EMT in accelerating radiation-induced pulmonary fibrosis. This process is partially mediated through β-catenin. Our study suggests a new mechanism for EMT, promoted by Tregs, that accelerates radiation-induced pulmonary fibrosis.

  9. Regulatory T Cells Promote β-Catenin--Mediated Epithelium-to-Mesenchyme Transition During Radiation-Induced Pulmonary Fibrosis.

    PubMed

    Xiong, Shanshan; Pan, Xiujie; Xu, Long; Yang, Zhihua; Guo, Renfeng; Gu, Yongqing; Li, Ruoxi; Wang, Qianjun; Xiao, Fengjun; Du, Li; Zhou, Pingkun; Zhu, Maoxiang

    2015-10-01

    Radiation-induced pulmonary fibrosis results from thoracic radiation therapy and severely limits radiation therapy approaches. CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) as well as epithelium-to-mesenchyme transition (EMT) cells are involved in pulmonary fibrosis induced by multiple factors. However, the mechanisms of Tregs and EMT cells in irradiation-induced pulmonary fibrosis remain unclear. In the present study, we investigated the influence of Tregs on EMT in radiation-induced pulmonary fibrosis. Mice thoraxes were irradiated (20 Gy), and Tregs were depleted by intraperitoneal injection of a monoclonal anti-CD25 antibody 2 hours after irradiation and every 7 days thereafter. Mice were treated on days 3, 7, and 14 and 1, 3, and 6 months post irradiation. The effectiveness of Treg depletion was assayed via flow cytometry. EMT and β-catenin in lung tissues were detected by immunohistochemistry. Tregs isolated from murine spleens were cultured with mouse lung epithelial (MLE) 12 cells, and short interfering RNA (siRNA) knockdown of β-catenin in MLE 12 cells was used to explore the effects of Tregs on EMT and β-catenin via flow cytometry and Western blotting. Anti-CD25 antibody treatment depleted Tregs efficiently, attenuated the process of radiation-induced pulmonary fibrosis, hindered EMT, and reduced β-catenin accumulation in lung epithelial cells in vivo. The coculture of Tregs with irradiated MLE 12 cells showed that Tregs could promote EMT in MLE 12 cells and that the effect of Tregs on EMT was partially abrogated by β-catenin knockdown in vitro. Tregs can promote EMT in accelerating radiation-induced pulmonary fibrosis. This process is partially mediated through β-catenin. Our study suggests a new mechanism for EMT, promoted by Tregs, that accelerates radiation-induced pulmonary fibrosis. Copyright © 2015. Published by Elsevier Inc.

  10. Genistein prevents ultraviolet B radiation-induced nitrosative skin injury and promotes cell proliferation.

    PubMed

    Terra, V A; Souza-Neto, F P; Frade, M A C; Ramalho, L N Z; Andrade, T A M; Pasta, A A C; Conchon, A C; Guedes, F A; Luiz, R C; Cecchini, R; Cecchini, A L

    2015-03-01

    Nitric oxide (NO) levels increase considerably after 24h of exposure of skin to ultraviolet B (UVB) radiation, which leads to nitrosative skin injury. In addition, increased NO levels after exposure to UVB radiation are associated with inhibition of cell proliferation. Compared to the UV-control group, UV-genistein at 10 mg/kg (UV-GEN10) group showed tissue protection, decreased lipid peroxide and nitrotyrosine formation, and low CAT activity. Furthermore, NO levels and iNOS labeling remained high. In this group, the reduction in lipid peroxides and nitrotyrosine was accompanied by upregulation of cell proliferation factors (Ki67 and PCNA), which indicated that prevention of nitrosative skin injury promoted cell proliferation and DNA repair. Genistein also prevented nitrosative events, inhibited ONOO(-) formation, which leads to tissue protection and cell proliferation. The UV-GEN15 group did not result in a greater protective effect compared to that with UV-GEN10 group. In the UV-GEN15 group, histological examination of the epidermis showed morphological alterations without efficient protection against lipid peroxide formation, as well as inhibition of Ki67 and PCNA, and VEGF labeling, which suggested inhibition of cell proliferation. These results help to elucidate the mechanisms underlying the photoprotective effect of genistein and reveal the importance of UVB radiation-induced nitrosative damage. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Ionizing radiation-induced mutant frequencies increase transiently in male germ cells of older mice.

    PubMed

    Xu, Guogang; McMahan, C Alex; Hildreth, Kim; Garcia, Rebecca A; Herbert, Damon C; Walter, Christi A

    2012-05-15

    Spontaneous mutant frequency in the male germline increases with age, thereby increasing the risk of siring offspring with genetic disorders. In the present study we investigated the effect of age on ionizing radiation-induced male germline mutagenesis. lacI transgenic mice were treated with ionizing radiation at 4-, 15- and 26-month-old, and mutant frequencies were determined for pachytene spermatocytes and round spermatids at 15 days or 49 days after ionizing radiation treatment. Cells collected 15 days after treatment were derivatives of irradiated differentiating spermatogenic cells while cells collected 49 days later were derivatives of spermatogonial stem cells. The results showed that (1) spontaneous mutant frequency increased in spermatogenic cells recovered from nonirradiated old mice (26-months-old), particularly in the round spermatids; (2) mutant frequencies were significantly increased in round spermatids obtained from middle-aged mice (15-months-old) and old age mice (26-months-old) at 15 and 49 days after irradiation compared to the sham-treated old mice; and (3) pachytene spermatocytes obtained from 15- or 26-month-old mice displayed a significantly increased mutant frequency at 15 days post irradiation. This study indicates that age modulates the mutagenic response to ionizing radiation in the male germline.

  12. uPAR and cathepsin B inhibition enhanced radiation-induced apoptosis in gliomainitiating cells

    PubMed Central

    Malla, Rama Rao; Gopinath, Sreelatha; Alapati, Kiranmai; Gorantla, Bharathi; Gondi, Christopher S.; Rao, Jasti S.

    2012-01-01

    Glioblastomas present as diffuse tumors with invasion into normal brain tissue and frequently recur or progress after radiation as focal masses because of glioma-initiating cells. The role of the urokinase-type plasminogen activator receptor (uPAR) and cathepsin B in stem-like phenotype has been extensively studied in several solid tumors. In the present study, we demonstrated that selection of glioma-initiating cells using CD133 expression leads to a specific enrichment of CD133+ cells in both U87 and 4910 cells. In addition, CD133+ cells exhibited a considerable amount of other stem cell markers, such as Nestin and Sox-2. Radiation treatment significantly enhanced uPAR and cathepsin B levels in glioma-initiating cells. To downregulate radiation-induced uPAR and cathepsin B expression, we used a bicistronic shRNA construct that simultaneously targets both uPAR and cathepsin B (pCU). Downregulation of uPAR and cathepsin B using pCU decreased radiation-enhanced uPAR and cathepsin B levels and caused DNA damage-induced apoptosis in glioma cell lines and glioma-initiating cells. The most striking finding of this study is that knockdown of uPAR and cathepsin B inhibited ongoing transcription by suppressing BrUTP incorporation at γH2AX foci. In addition, uPAR and cathepsin B gene silencing inversely regulated survivin and H2AX expression in both glioma cells and glioma-initiating cells. Pretreatment with pCU reduced radiation-enhanced expression of uPAR, cathepsin B, and survivin and enhanced DNA damage in pre-established glioma in nude mice. Taken together, our in vitro and in vivo findings suggest that uPAR and cathepsin B inhibition might serve as an adjunct to radiation therapy to target glioma-initiating cells and, therefore, for the treatment of glioma. PMID:22573309

  13. Ion Channels, Cell Volume, Cell Proliferation and Apoptotic Cell Death

    NASA Astrophysics Data System (ADS)

    Lang, Florian; Gulbins, Erich; Szabo, Ildiko; Vereninov, Alexey; Huber, Stephan M.

    At some stage cell proliferation requires an increase in cell volume and a typical hallmark of apoptotic cell death is cell shrinkage. The respective alterations of cell volume are accomplished by altered regulation of ion transport including ion channels. Thus, cell proliferation and apoptosis are both paralleled by altered activity of ion channels, which play an active part in these fundamental cellular mechanisms. Activation of anion channels allows exit of Cl?, osmolyte and HCO3 ? leading to cell shrinkage and acidification of the cytosol. K+ exit through K+ channels leads to cell shrinkage and a decrease in intracellular K+ concentration. K+ channel activity is further important for maintenance of the cell membrane potential - a critical determinant of Ca2+ entry through Ca2+ channels. Cytosolic Ca2+ may both activate mechanisms required for cell proliferation and stimulate enzymes executing apoptosis. The effect of enhanced cytosolic Ca2+ activity depends on the magnitude and temporal organisation of Ca2+ entry. Moreover, a given ion channel may support both cell proliferation and apoptosis, and specific ion channel blockers may abrogate both fundamental cellular mechanisms, depending on cell type, regulatory environment and condition of the cell. Clearly, further experimental effort is needed to clarify the role of ion channels in the regulation of cell proliferation and apoptosis.

  14. Apoptotic Cell Death of Human Interstitial Cells of Cajal

    PubMed Central

    De Giorgio, Roberto; Faussone Pellegrini, Maria Simonetta; Garrity-Park, Megan M.; Miller, Steven M.; Schmalz, Philip F.; Young-Fadok, Tonia M.; Larson, David W.; Dozois, Eric J.; Camilleri, Michael; Stanghellini, Vincenzo; Szurszewski, Joseph H.; Farrugia, Gianrico

    2008-01-01

    Interstitial cells of Cajal (ICC) are specialized mesenchyme-derived cells that regulate contractility and excitability of many smooth muscles with loss of ICC seen in a variety of gut motility disorders. Maintenance of ICC numbers is tightly regulated, with several factors known to regulate proliferation. In contrast, the fate of ICC is not established. The aim of this study was to investigate whether apoptosis plays a role in the regulation of ICC numbers in the normal colon. ICC were identified by immunolabeling for the c-Kit receptor tyrosine kinase and by electron microscopy. Apoptosis was detected in colon tissue by immunolabeling for activated caspase-3, terminal dUTP nucleotide end labeling, and ultrastructural changes in the cells. Apoptotic ICC were identified and counted in double labeled tissue sections. Apoptotic ICC were identified in all layers of the colonic muscle. In the muscularis propria 1.5 ± 0.2% of ICC were positive for activated caspase-3 and in the circular muscle layer 2.1 ± 0.9% of ICC were positive for TUNEL. Apoptotic ICC were identified by electron microscopy. Apoptotic cell death is ongoing in ICC. The level of apoptosis in ICC in healthy colon indicates that these cells must be continually regenerated to maintain intact networks. PMID:18798796

  15. Ionizing radiation-induced mutagenesis: radiation studies in Neurospora predictive for results in mammalian cells

    NASA Technical Reports Server (NTRS)

    Evans, H. H.; DeMarini, D. M.

    1999-01-01

    Ionizing radiation was the first mutagen discovered and was used to develop the first mutagenicity assay. In the ensuing 70+ years, ionizing radiation became a fundamental tool in understanding mutagenesis and is still a subject of intensive research. Frederick de Serres et al. developed and used the Neurospora crassa ad-3 system initially to explore the mutagenic effects of ionizing radiation. Using this system, de Serres et al. demonstrated the dependence of the frequency and spectra of mutations induced by ionizing radiation on the dose, dose rate, radiation quality, repair capabilities of the cells, and the target gene employed. This work in Neurospora predicted the subsequent observations of the mutagenic effects of ionizing radiation in mammalian cells. Modeled originally on the mouse specific-locus system developed by William L. Russell, the N. crassa ad-3 system developed by de Serres has itself served as a model for interpreting the results in subsequent systems in mammalian cells. This review describes the primary findings on the nature of ionizing radiation-induced mutagenesis in the N. crassa ad-3 system and the parallel observations made years later in mammalian cells.

  16. Ionizing radiation-induced mutagenesis: radiation studies in Neurospora predictive for results in mammalian cells

    NASA Technical Reports Server (NTRS)

    Evans, H. H.; DeMarini, D. M.

    1999-01-01

    Ionizing radiation was the first mutagen discovered and was used to develop the first mutagenicity assay. In the ensuing 70+ years, ionizing radiation became a fundamental tool in understanding mutagenesis and is still a subject of intensive research. Frederick de Serres et al. developed and used the Neurospora crassa ad-3 system initially to explore the mutagenic effects of ionizing radiation. Using this system, de Serres et al. demonstrated the dependence of the frequency and spectra of mutations induced by ionizing radiation on the dose, dose rate, radiation quality, repair capabilities of the cells, and the target gene employed. This work in Neurospora predicted the subsequent observations of the mutagenic effects of ionizing radiation in mammalian cells. Modeled originally on the mouse specific-locus system developed by William L. Russell, the N. crassa ad-3 system developed by de Serres has itself served as a model for interpreting the results in subsequent systems in mammalian cells. This review describes the primary findings on the nature of ionizing radiation-induced mutagenesis in the N. crassa ad-3 system and the parallel observations made years later in mammalian cells.

  17. Radiation-Induced Alterations of Osteogenic and Chondrogenic Differentiation of Human Mesenchymal Stem Cells

    PubMed Central

    Cruet-Hennequart, Séverine; Drougard, Carole; Shaw, Georgina; Legendre, Florence; Demoor, Magali; Barry, Frank; Lefaix, Jean-Louis; Galéra, Philippe

    2015-01-01

    While human mesenchymal stem cells (hMSCs), either in the bone marrow or in tumour microenvironment could be targeted by radiotherapy, their response is poorly understood. The oxic effects on radiosensitivity, cell cycle progression are largely unknown, and the radiation effects on hMSCs differentiation capacities remained unexplored. Here we analysed hMSCs viability and cell cycle progression in 21% O2 and 3% O2 conditions after medical X-rays irradiation. Differentiation towards osteogenesis and chondrogenesis after irradiation was evaluated through an analysis of differentiation specific genes. Finally, a 3D culture model in hypoxia was used to evaluate chondrogenesis in conditions mimicking the natural hMSCs microenvironment. The hMSCs radiosensitivity was not affected by O2 tension. A decreased number of cells in S phase and an increase in G2/M were observed in both O2 tensions after 16 hours but hMSCs released from the G2/M arrest and proliferated at day 7. Osteogenesis was increased after irradiation with an enhancement of mRNA expression of specific osteogenic genes (alkaline phosphatase, osteopontin). Osteoblastic differentiation was altered since matrix deposition was impaired with a decreased expression of collagen I, probably through an increase of its degradation by MMP-3. After induction in monolayers, chondrogenesis was altered after irradiation with an increase in COL1A1 and a decrease in both SOX9 and ACAN mRNA expression. After induction in a 3D culture in hypoxia, chondrogenesis was altered after irradiation with a decrease in COL2A1, ACAN and SOX9 mRNA amounts associated with a RUNX2 increase. Together with collagens I and II proteins decrease, associated to a MMP-13 expression increase, these data show a radiation-induced impairment of chondrogenesis. Finally, a radiation-induced impairment of both osteogenesis and chondrogenesis was characterised by a matrix composition alteration, through inhibition of synthesis and/or increased degradation

  18. Radiation-induced strain differences in mouse alveolar inflammatory cell apoptosis.

    PubMed

    O'Brien, Thomas J; Létuvé, Séverine; Haston, Christina K

    2005-01-01

    Whole-thorax irradiation results in the development of the diffuse inflammatory response alveolitis in C3H/HeJ (C3H) mice and a milder alveolitis with fibrosis in C57BL/6J (B6) mice. In this study, we investigate if this mouse strain difference in response to radiation is due to differences in lung inflammatory cell apoptosis. Mice of the C3H and B6 strains were given a radiation dose of 18 Gy to the thorax and the animals were sacrificed at 11 or 18 weeks following exposure or when they were moribund. Active caspase-3 staining was used to identify apoptotic cells in the alveolar space of histological lung sections from the mice. The apoptotic index of B6 mice was greater than that of C3H mice at 11 weeks postirradiation (17.8% of airspace cells vs. 7.8%, p = 0.028) and in mice sacrificed because of illness (27.3% vs. 14.4%, p = 0.036). No C3H mice survived to the later time point. The inflammatory cells undergoing apoptosis in the mouse lungs were morphologically consistent with alveolar macrophages. We conclude that a difference in inflammatory cell apoptosis may contribute to the disparate pulmonary radiation response of these mouse strains.

  19. Protective Role of Hsp27 Protein Against Gamma Radiation-Induced Apoptosis and Radiosensitization Effects of Hsp27 Gene Silencing in Different Human Tumor Cells

    SciTech Connect

    Aloy, Marie-Therese Hadchity, Elie; Bionda, Clara; Diaz-Latoud, Chantal; Claude, Line; Rousson, Robert; Arrigo, Andre-Patrick; Rodriguez-Lafrasse, Claire

    2008-02-01

    Purpose: The ability of heat shock protein 27 (Hsp27) to protect cells from stressful stimuli and its increased levels in tumors resistant to anticancer therapeutics suggest that it may represent a target for sensitization to radiotherapy. In this study, we investigate the protective role of Hsp27 against radiation-induced apoptosis and the effect of its attenuation in highly expressing radioresistant cancer cell lines. Methods and Materials: We examined clonogenic death and the kinetics of apoptotic events in different tumor cell lines overexpressing or underexpressing Hsp27 protein irradiated with photons. The radiosensitive Jurkat cell line, which does not express Hsp27 constitutively or in response to {gamma}-rays, was stably transfected with Hsp27 complementary DNA. Attenuation of Hsp27 expression was accomplished by antisense or RNAi (interfering RNA) strategies in SQ20B head-and-neck squamous carcinoma, PC3 prostate cancer, and U87 glioblastoma radioresistant cells. Results: We measured concentration-dependent protection against the cytotoxic effects of radiation in Jurkat-Hsp27 cells, which led to a 50% decrease in apoptotic cells at 48 hours in the highest expressing cells. Underlying mechanisms leading to radiation resistance involved a significant increase in glutathione levels associated with detoxification of reactive oxygen species, a delay in mitochondrial collapse, and caspase activation. Conversely, attenuation of Hsp27 in SQ20B cells, characterized by their resistance to apoptosis, sensitizes cells to irradiation. This was emphasized by increased apoptosis, decreased glutathione basal level, and clonogenic cell death. Sensitization to irradiation was confirmed in PC3 and U87 radioresistant cells. Conclusion: Hsp27 gene therapy offers a potential adjuvant to radiation-based therapy of resistant tumors.

  20. The Role of Deoxycytidine Kinase (dCK) in Radiation-Induced Cell Death

    PubMed Central

    Zhong, Rui; Xin, Rui; Chen, Zongyan; Liang, Nan; Liu, Yang; Ma, Shumei; Liu, Xiaodong

    2016-01-01

    Deoxycytidine kinase (dCK) is a key enzyme in deoxyribonucleoside salvage and the anti-tumor activity for many nucleoside analogs. dCK is activated in response to ionizing radiation (IR)-induced DNA damage and it is phosphorylated on Serine 74 by the Ataxia-Telangiectasia Mutated (ATM) kinase in order to activate the cell cycle G2/M checkpoint. However, whether dCK plays a role in radiation-induced cell death is less clear. In this study, we genetically modified dCK expression by knocking down or expressing a WT (wild-type), S74A (abrogates phosphorylation) and S74E (mimics phosphorylation) of dCK. We found that dCK could decrease IR-induced total cell death and apoptosis. Moreover, dCK increased IR-induced autophagy and dCK-S74 is required for it. Western blotting showed that the ratio of phospho-Akt/Akt, phospho-mTOR/mTOR, phospho-P70S6K/P70S6K significantly decreased in dCK-WT and dCK-S74E cells than that in dCK-S74A cells following IR treatment. Reciprocal experiment by co-immunoprecipitation showed that mTOR can interact with wild-type dCK. IR increased polyploidy and decreased G2/M arrest in dCK knock-down cells as compared with control cells. Taken together, phosphorylated and activated dCK can inhibit IR-induced cell death including apoptosis and mitotic catastrophe, and promote IR-induced autophagy through PI3K/Akt/mTOR pathway. PMID:27879648

  1. Metformin inhibits the radiation-induced invasive phenotype of esophageal squamous cell carcinoma.

    PubMed

    Nakayama, Akira; Ninomiya, Itasu; Harada, Shinichi; Tsukada, Tomoya; Okamoto, Koichi; Nakanuma, Shinichi; Sakai, Seisho; Makino, Isamu; Kinoshita, Jun; Hayashi, Hironori; Oyama, Katsunobu; Miyashita, Tomoharu; Tajima, Hidehiro; Takamura, Hiroyuki; Fushida, Sachio; Ohta, Tetsuo

    2016-11-01

    Esophageal cancer is one of the most aggressive tumor types because of its invasiveness and metastatic potential. Several reports have described an association between increased invasiveness after ionizing radiation (IR) treatment and epithelial-to-mesenchymal transition (EMT). The biguanide metformin is reported to prevent transforming growth factor-β (TGF-β)-induced EMT and proliferation of cancer. This study examined whether IR induces EMT and promotes the invasive potential of TE-9 esophageal squamous cell carcinoma cells and the effect of metformin on IR-induced EMT. After IR exposure, TE-9 cells showed a spindle-shaped morphology and lost cell-cell adhesion. Immunoblotting showed that IR induced expression of mesenchymal markers (vimentin and N-cadherin), transcription factors (Slug, Snail, and Twist), and matrix metalloproteinases. A scratch wound assay and Matrigel invasion assay showed that IR enhanced the invasive potential and migratory capacity of TE-9 cells. Expression of hypoxia-related factor-1α and TGF-β was increased after IR. IR also induced phosphorylation of Smad2 and Smad3. Metformin inhibited radiation-induced EMT-like morphological changes, and enhanced invasion and migration of TE-9 cells. Metformin inhibited IR-induced phosphorylation of Smad2 and Smad3. Although phosphorylation of AMP-activated protein kinase was enhanced by IR and metformin, phosphorylation of mammalian target of rapamycin was enhanced by IR and suppressed by metformin. These results indicated that metformin suppressed IR-induced EMT via suppression of the TGF-β-Smad phosphorylation pathway, and a part of the non-Smad pathway. Metformin might be useful to prevent IR-induced invasion and metastasis of esophageal squamous cell carcinoma.

  2. The Role of Deoxycytidine Kinase (dCK) in Radiation-Induced Cell Death.

    PubMed

    Zhong, Rui; Xin, Rui; Chen, Zongyan; Liang, Nan; Liu, Yang; Ma, Shumei; Liu, Xiaodong

    2016-11-21

    Deoxycytidine kinase (dCK) is a key enzyme in deoxyribonucleoside salvage and the anti-tumor activity for many nucleoside analogs. dCK is activated in response to ionizing radiation (IR)-induced DNA damage and it is phosphorylated on Serine 74 by the Ataxia-Telangiectasia Mutated (ATM) kinase in order to activate the cell cycle G2/M checkpoint. However, whether dCK plays a role in radiation-induced cell death is less clear. In this study, we genetically modified dCK expression by knocking down or expressing a WT (wild-type), S74A (abrogates phosphorylation) and S74E (mimics phosphorylation) of dCK. We found that dCK could decrease IR-induced total cell death and apoptosis. Moreover, dCK increased IR-induced autophagy and dCK-S74 is required for it. Western blotting showed that the ratio of phospho-Akt/Akt, phospho-mTOR/mTOR, phospho-P70S6K/P70S6K significantly decreased in dCK-WT and dCK-S74E cells than that in dCK-S74A cells following IR treatment. Reciprocal experiment by co-immunoprecipitation showed that mTOR can interact with wild-type dCK. IR increased polyploidy and decreased G2/M arrest in dCK knock-down cells as compared with control cells. Taken together, phosphorylated and activated dCK can inhibit IR-induced cell death including apoptosis and mitotic catastrophe, and promote IR-induced autophagy through PI3K/Akt/mTOR pathway.

  3. A role for TRAIL/TRAIL-R2 in radiation-induced apoptosis and radiation-induced bystander response of human neural stem cells.

    PubMed

    Ivanov, Vladimir N; Hei, Tom K

    2014-03-01

    Adult neurons, which are terminally differentiated cells, demonstrate substantial radioresistance. In contrast, human neural stem cells (NSC), which have a significant proliferative capacity, are highly sensitive to ionizing radiation. Cranial irradiation that is widely used for treatment of brain tumors may induce death of NSC and further cause substantial cognitive deficits such as impairing learning and memory. The main goal of our study was to determine a mechanism of NSC radiosensitivity. We observed a constitutive high-level expression of TRAIL-R2 in human NSC. On the other hand, ionizing radiation through generation of reactive oxygen species targeted cell signaling pathways and dramatically changed the pattern of gene expression, including upregulation of TRAIL. A significant increase of endogenous expression and secretion of TRAIL could induce autocrine/paracrine stimulation of the TRAIL-R2-mediated signaling cascade with activation of caspase-3-driven apoptosis. Furthermore, paracrine stimulation could initiate bystander response of non-targeted NSC that is driven by death ligands produced by directly irradiated NSC. Experiments with media transfer from directly irradiated NSC to non-targeted (bystander) NSC confirmed a role of secreted TRAIL for induction of a death signaling cascade in non-targeted NSC. Subsequently, TRAIL production through elimination of bystander TRAIL-R-positive NSC might substantially restrict a final yield of differentiating young neurons. Radiation-induced TRAIL-mediated apoptosis could be partially suppressed by anti-TRAIL antibody added to the cell media. Interestingly, direct gamma-irradiation of SK-N-SH human neuroblastoma cells using clinical doses (2-5 Gy) resulted in low levels of apoptosis in cancer cells that was accompanied however by induction of a strong bystander response in non-targeted NSC. Numerous protective mechanisms were involved in the maintenance of radioresistance of neuroblastoma cells, including

  4. A role for TRAIL/TRAIL-R2 in radiation-induced apoptosis and radiation-induced bystander response of human neural stem cells

    PubMed Central

    Ivanov, Vladimir N.; Hei, Tom K.

    2013-01-01

    Adult neurons, which are terminally differentiated cells, demonstrate substantial radioresistance. In contrast, human neural stem cells (NSC), which have a significant proliferative capacity, are highly sensitive to ionizing radiation. Cranial irradiation that is widely used for treatment of brain tumors may induce death of NSC and further cause substantial cognitive deficits such as impairing learning and memory. The main goal of our study was to determine a mechanism of NSC radiosensitivity. We observed a constitutive high-level expression of TRAIL-R2 in human NSC. On the other hand, ionizing radiation through generation of reactive oxygen species targeted cell signaling pathways and dramatically changed the pattern of gene expression, including upregulation of TRAIL. A significant increase of endogenous expression and secretion of TRAIL could induce autocrine/paracrine stimulation of the TRAIL-R2-mediated signaling cascade with activation of caspase-3-driven apoptosis. Furthermore, paracrine stimulation could initiate bystander response of non-targeted NSC that is driven by death ligands produced by directly irradiated NSC. Experiments with media transfer from directly irradiated NSC to non-targeted (bystander) NSC confirmed a role of secreted TRAIL for induction of a death signaling cascade in non-targeted NSC. Subsequently, TRAIL production through elimination of bystander TRAIL-R-positive NSC might substantially restrict a final yield of differentiating young neurons. Radiation-induced TRAIL-mediated apoptosis could be partially suppressed by anti-TRAIL antibody added to the cell media. Interestingly, direct gamma-irradiation of SKN-SH human neuroblastoma cells using clinical doses (2–5 Gy) resulted in low levels of apoptosis in cancer cells that was accompanied however by induction of a strong bystander response in non-targeted NSC. Numerous protective mechanisms were involved in the maintenance of radioresistance of neuroblastoma cells, including

  5. X-Radiation Induces Non-Small-Cell Lung Cancer Apoptosis by Upregulation of Axin Expression

    SciTech Connect

    Han Yang; Wang Yan; Xu Hongtao; Yang Lianhe; Wei Qiang; Liu Yang; Zhang Yong; Zhao Yue; Dai Shundong; Miao Yuan; Yu Juanhan; Zhang Junyi; Li, Guang; Yuan Ximing; Wang Enhua

    2009-10-01

    Purpose: Axis inhibition (Axin) is an important negative regulator of the Wnt pathway. This study investigated the relationship between Axin expression and sensitivity to X-rays in non-small-cell lung cancer (NSCLC) to find a useful indicator of radiosensitivity. Methods and Materials: Tissue from NSCLC patients, A549 cells, and BE1 cells expressing Axin were exposed to 1-Gy of X-radiation. Axin and p53 expression levels were detected by immunohistochemistry and reverse transcription-PCR. Apoptosis was determined by TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling) assay and FACS (fluorescence-activate cell sorter) analysis. Caspase-3 activity was determined by Western blotting. Phospho-JNK expression was determined by immunofluorescence. Results: The expression of Axin was significantly lower in NSCLC tissues than in normal lung tissues (p < 0.05). Axin expression correlates with differentiation, TNM staging, and lymph node metastasis of NSCLC (p < 0.05). Its expression negatively correlates with the expression of p53(mt) (p=0.000) and positively correlates with apoptosis (p=0.002). The prognosis of patients with high expression of Axin was better than those with low expression. X-radiation increases Axin expression in NSCLC tissue, and caspase-3 is significantly higher in samples in which Axin is increased (p < 0.05). Both X-radiation and Axin induce apoptosis of A549 and BE1 cells; however, the combination of the two enhances the apoptotic effect (p < 0.05). In A549 cells, inhibition of p53 blocks Axin-induced apoptosis, whereas in BE1 cells, the JNK pathway is required. Conclusions: Axin induces the p53 apoptotic pathway in cells where this pathway is intact; however, in cells expressing p53(mt), Axin induces apoptosis via the JNK pathway. Elevated Axin expression following X-ray exposure is a reliable indicator for determining the radiosensitivity of NSCLC.

  6. Relief of delayed oxidative stress by ascorbic acid can suppress radiation-induced cellular senescence in mammalian fibroblast cells.

    PubMed

    Kobashigawa, Shinko; Kashino, Genro; Mori, Hiromu; Watanabe, Masami

    2015-03-01

    Ionizing radiation-induced cellular senescence is thought to be caused by nuclear DNA damage that cannot be repaired. However, here we found that radiation induces delayed increase of intracellular oxidative stress after irradiation. We investigated whether the relief of delayed oxidative stress by ascorbic acid would suppress the radiation-induced cellular senescence in Syrian golden hamster embryo (SHE) cells. We observed that the level of oxidative stress was drastically increased soon after irradiation, then declined to the level in non-irradiated cells, and increased again with a peak on day 3 after irradiation. We found that the inductions of cellular senescence after X-irradiation were reduced along with suppression of the delayed induction of oxidative stress by treatment with ascorbic acid, but not when oxidative stress occurred immediately after irradiation. Moreover, treatment of ascorbic acid inhibited p53 accumulation at 3 days after irradiation. Our data suggested a delayed increase of intracellular oxidative stress levels plays an important role in the process of radiation-induced cellular senescence by p53 accumulation.

  7. A human esophageal epithelial cell model for study of radiation induced cancer and DNA repair

    NASA Astrophysics Data System (ADS)

    Huff, Janice; Patel, Zarana; Hada, Megumi; Cucinotta, Francis A.

    For cancer risk assessment in astronauts and for countermeasure development, it is essential to understand the molecular mechanisms of radiation carcinogenesis and how these mechanisms are influenced by exposure to the types of radiation found in space. We are developing an in vitro model system for the study of radiation-induced initiation and progression of esophageal carcinoma. Development of squamous cell carcinoma of the esophagus is associated with radiation exposure, as revealed by the significant enhanced in incidence rates for this type of cancer in the survivors of the atomic bomb detonations in Japan. It is also associated with poor nutritional status and micronutrient deficiencies, which are also important issues for long duration spaceflight. The possible synergies between nutritional issues and radiation exposure are unknown. Here we present the results of preliminary characterization of both normal and hTERT-immortalized esophageal epithelial cells grown in 2-dimensional culture. We analyzed DNA repair capacity by measuring the kinetics of formation and loss of gamma-H2AX foci following radiation exposure. Additionally, we analyzed induction of chromosomal aberrations using 3-color fluorescence in situ hybridization (FISH). Data were generated using both low LET (gamma rays) and high LET ions (1000 MeV/nucleon iron.

  8. Directional delivery of RSPO1 by mesenchymal stem cells ameliorates radiation-induced intestinal injury.

    PubMed

    Chen, Wei; Ju, Songwen; Lu, Ting; Xu, Yongfang; Zheng, Xiaocui; Wang, Haiyan; Ge, Yan; Ju, Songguang

    2017-02-16

    Radiation-induced intestinal injury (RIII) commonly occurs in patients who received radiotherapy for pelvic or abdominal cancer, or who suffered from whole-body irradiation during a nuclear accident. RIII can lead to intestinal disorders and even death given its integrity damage that results from intestinal stem cell (ISC) loss. Recovery from RIII relies on the intensity of supportive treatment, which can attenuate lethal infection and give surviving stem cells an opportunity to regenerate. It has been reported that RSPO1 is a cytokine with potent and specific proliferative effects on intestinal crypt cells. MSCs have multiple RIII-healing effects, including anti-inflammatory and anti-irradiation injury properties, due to its negative immune regulation and its homing ability to the damaged intestinal epithelia. To combine the comprehensive anti-injury potential of MSCs, and the potent ability of RSPO1 as a mitogenic factor for ISCs, we constructed RSPO1-modified C3H10 T1/2 cells and expected that RSPO1, the ISC-proliferative cytokine, could be delivered to the site of injury in a targeted manner. In this study, we transferred C3H10/RSPO1 intravenously via the retro-orbital sinus into mice suffering from abdominal irradiation at lethal dosages. Our findings demonstrated that C3H10/RSPO1 cells are able to directionally migrate to the injury site; enhance ISC survival, proliferation, and differentiation; and effectively repair the radiation-damaged intestinal epithelial cells. This study suggests that the directional delivery of RSPO1 by MSCs is a promising strategy to ameliorate, and even cure, RIII.

  9. From radiation-induced chromosome damage to cell death: modelling basic mechanisms and applications to boron neutron capture therapy.

    PubMed

    Ballarini, F; Bortolussi, S; Clerici, A M; Ferrari, C; Protti, N; Altieri, S

    2011-02-01

    Cell death is a crucial endpoint in radiation-induced biological damage: on one side, cell death is a reference endpoint to characterise the action of radiation in biological targets; on the other side, any cancer therapy aims to kill tumour cells. Starting from Lea's target theory, many models have been proposed to interpret radiation-induced cell killing; after briefly discussing some of these models, in this paper, a mechanistic approach based on an experimentally observed link between chromosome aberrations and cell death was presented. More specifically, a model and a Monte Carlo code originally developed for chromosome aberrations were extended to simulate radiation-induced cell death applying an experimentally observed one-to-one relationship between the average number of 'lethal aberrations' (dicentrics, rings and deletions) per cell and -ln S, S being the fraction of surviving cells. Although such observation was related to X rays, in the present work, the approach was also applied to protons and alpha particles. A good agreement between simulation outcomes and literature data provided a model validation for different radiation types. The same approach was then successfully applied to simulate the survival of cells enriched with boron and irradiated with thermal neutrons at the Triga Mark II reactor in Pavia, to mimic a typical treatment for boron neutron capture therapy.

  10. Sam68 is cleaved by caspases under apoptotic cell death induced by ionizing radiation.

    PubMed

    Cho, Seong-Jun; Choi, Moo Hyun; Nam, Seon Young; Kim, Ji Young; Kim, Cha Soon; Pyo, Suhkneung; Yang, Kwang Hee

    2015-03-01

    The RNA-binding protein Sam68, a mitotic substrate of tyrosine kinases, has been reported to participate in the cell cycle, apoptosis, and signaling. In particular, overexpression of Sam68 protein is known to suppress cell growth and cell cycle progression in NIH3T3 cells. Although Sam68 is involved in many cellular activities, the function of Sam68, especially in response to apoptotic stimulation, is not well understood. In this study, we found that Sam68 protein is cleaved in immune cells undergoing apoptosis induced by γ-radiation. Moreover, we found that Sam68 cleavage was induced by apoptotic stimuli containing γ-radiation in a caspase-dependent manner. In particular, we showed that activated casepase-3, 7, 8 and 9 can directly cleave Sam68 protein through in vitro protease cleavage assay. Finally, we found that the knockdown of Sam68 attenuated γ-radiation-induced cell death and growth suppression. Conclusively, the cleavage of Sam68 is a new indicator for the cell damaging effects of ionizing radiation. © The Author 2015. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.

  11. Silibinin attenuates ionizing radiation-induced pro-angiogenic response and EMT in prostate cancer cells

    SciTech Connect

    Nambiar, Dhanya K.; Rajamani, Paulraj; Singh, Rana P.

    2015-01-02

    Graphical abstract: Potential model showing mechanism of silibinin-mediated attenuation of IR-induced angiogenic phenotype and EMT in tumor cells. Silibinin counters radiation induced invasive and migratory phenotype of cancer cells by down-regulating mitogenic pathways activated by IR, leading to inhibition of molecules including VEGF, iNOS, MMPs and N-cadherin. Silibinin also reverses IR mediated E-cadherin down-regulation, inhibiting EMT in tumor cells. Silibinin also radiosensitizes endothelial cells, reduces capillary tube formation by targeting various pro-angiogenic molecules. Further, silibinin may inhibit autocrine and paracrine signaling between tumor and endothelial cells by decreasing the levels of VEGF and other signaling molecules activated in response to IR. - Highlights: • Silibinin radiosensitizes endothelial cells. • Silibinin targets ionization radiation (IR)-induced EMT in PCa cells. • Silibinin is in phase II clinical trial in PCa patients, hence clinically relevant. - Abstract: Radiotherapy of is well established and frequently utilized in prostate cancer (PCa) patients. However, recurrence following therapy and distant metastases are commonly encountered problems. Previous studies underline that, in addition to its therapeutic effects, ionizing radiation (IR) increases the vascularity and invasiveness of surviving radioresistant cancer cells. This invasive phenotype of radioresistant cells is an upshot of IR-induced pro-survival and mitogenic signaling in cancer as well as endothelial cells. Here, we demonstrate that a plant flavonoid, silibinin can radiosensitize endothelial cells by inhibiting expression of pro-angiogenic factors. Combining silibinin with IR not only strongly down-regulated endothelial cell proliferation, clonogenicity and tube formation ability rather it strongly (p < 0.001) reduced migratory and invasive properties of PCa cells which were otherwise marginally affected by IR treatment alone. Most of the pro

  12. Circulating IgM Requires Plasma Membrane Disruption to Bind Apoptotic and Non-Apoptotic Nucleated Cells and Erythrocytes.

    PubMed

    Hesketh, Emily E; Dransfield, Ian; Kluth, David C; Hughes, Jeremy

    2015-01-01

    Autoimmunity is associated with defective phagocytic clearance of apoptotic cells. IgM deficient mice exhibit an autoimmune phenotype consistent with a role for circulating IgM antibodies in apoptotic cell clearance. We have extensively characterised IgM binding to non-apoptotic and apoptotic mouse thymocytes and human Jurkat cells using flow cytometry, confocal imaging and electron microscopy. We demonstrate strong specific IgM binding to a subset of Annexin-V (AnnV)+PI (Propidium Iodide)+ apoptotic cells with disrupted cell membranes. Electron microscopy studies indicated that IgM+AnnV+PI+ apoptotic cells exhibited morphologically advanced apoptosis with marked plasma membrane disruption compared to IgM-AnnV+PI+ apoptotic cells, suggesting that access to intracellular epitopes is required for IgM to bind. Strong and comparable binding of IgM to permeabilised non-apoptotic and apoptotic cells suggests that IgM bound epitopes are 'apoptosis independent' such that IgM may bind any cell with profound disruption of cell plasma membrane integrity. In addition, permeabilised erythrocytes exhibited significant IgM binding thus supporting the importance of cell membrane epitopes. These data suggest that IgM may recognize and tag damaged nucleated cells or erythrocytes that exhibit significant cell membrane disruption. The role of IgM in vivo in conditions characterized by severe cell damage such as ischemic injury, sepsis and thrombotic microangiopathies merits further exploration.

  13. Radiation Induced Apoptosis of Murine Bone Marrow Cells Is Independent of Early Growth Response 1 (EGR1)

    PubMed Central

    Oben, Karine Z.; Gachuki, Beth W.; Alhakeem, Sara S.; McKenna, Mary K.; Liang, Ying; St. Clair, Daret K.; Rangnekar, Vivek M.; Bondada, Subbarao

    2017-01-01

    An understanding of how each individual 5q chromosome critical deleted region (CDR) gene contributes to malignant transformation would foster the development of much needed targeted therapies for the treatment of therapy related myeloid neoplasms (t-MNs). Early Growth Response 1 (EGR1) is a key transcriptional regulator of myeloid differentiation located within the 5q chromosome CDR that has been shown to regulate HSC (hematopoietic stem cell) quiescence as well as the master regulator of apoptosis—p53. Since resistance to apoptosis is a hallmark of malignant transformation, we investigated the role of EGR1 in apoptosis of bone marrow cells; a cell population from which myeloid malignancies arise. We evaluated radiation induced apoptosis of Egr1+/+ and Egr1-/- bone marrow cells in vitro and in vivo. EGR1 is not required for radiation induced apoptosis of murine bone marrow cells. Neither p53 mRNA (messenger RNA) nor protein expression is regulated by EGR1 in these cells. Radiation induced apoptosis of bone marrow cells by double strand DNA breaks induced p53 activation. These results suggest EGR1 dependent signaling mechanisms do not contribute to aberrant apoptosis of malignant cells in myeloid malignancies. PMID:28081176

  14. Signaling pathway for phagocyte priming upon encounter with apoptotic cells.

    PubMed

    Nonaka, Saori; Ando, Yuki; Kanetani, Takuto; Hoshi, Chiharu; Nakai, Yuji; Nainu, Firzan; Nagaosa, Kaz; Shiratsuchi, Akiko; Nakanishi, Yoshinobu

    2017-05-12

    The phagocytic elimination of cells undergoing apoptosis is an evolutionarily conserved innate immune mechanism for eliminating unnecessary cells. Previous studies showed an increase in the level of engulfment receptors in phagocytes after the phagocytosis of apoptotic cells, which leads to the enhancement of their phagocytic activity. However, precise mechanisms underlying this phenomenon require further clarification. We found that the pre-incubation of a Drosophila phagocyte cell line with the fragments of apoptotic cells enhanced the subsequent phagocytosis of apoptotic cells, accompanied by an augmented expression of the engulfment receptors Draper and integrin αPS3. The DNA-binding activity of the transcription repressor Tailless was transiently raised in those phagocytes, depending on two partially overlapping signal-transduction pathways for the induction of phagocytosis as well as the occurrence of engulfment. The RNAi knockdown of tailless in phagocytes abrogated the enhancement of both phagocytosis and engulfment receptor expression. Furthermore, the hemocyte-specific RNAi of tailless reduced apoptotic cell clearance in Drosophila embryos. Taken together, we propose the following mechanism for the activation of Drosophila phagocytes after an encounter with apoptotic cells: two partially overlapping signal-transduction pathways for phagocytosis are initiated; transcription repressor Tailless is activated; expression of engulfment receptors is stimulated; and phagocytic activity is enhanced. This phenomenon most likely ensures the phagocytic elimination of apoptotic cells by stimulated phagocytes and is thus considered as a mechanism to prime phagocytes in innate immunity. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Mesenchymal stem cells inhibit cutaneous radiation-induced fibrosis by suppressing chronic inflammation.

    PubMed

    Horton, Jason A; Hudak, Kathryn E; Chung, Eun Joo; White, Ayla O; Scroggins, Bradley T; Burkeen, Jeffrey F; Citrin, Deborah E

    2013-10-01

    Exposure to ionizing radiation (IR) can result in the development of cutaneous fibrosis, for which few therapeutic options exist. We tested the hypothesis that bone marrow-derived mesenchymal stem cells (BMSC) would favorably alter the progression of IR-induced fibrosis. We found that a systemic infusion of BMSC from syngeneic or allogeneic donors reduced skin contracture, thickening, and collagen deposition in a murine model. Transcriptional profiling with a fibrosis-targeted assay demonstrated increased expression of interleukin-10 (IL-10) and decreased expression of IL-1β in the irradiated skin of mice 14 days after receiving BMSC. Similarly, immunoassay studies demonstrated durable alteration of these and several additional inflammatory mediators. Immunohistochemical studies revealed a reduction in infiltration of proinflammatory classically activated CD80(+) macrophages and increased numbers of anti-inflammatory regulatory CD163(+) macrophages in irradiated skin of BMSC-treated mice. In vitro coculture experiments confirmed that BMSC induce expression of IL-10 by activated macrophages, suggesting polarization toward a regulatory phenotype. Furthermore, we demonstrated that tumor necrosis factor-receptor 2 (TNF-R2) mediates IL-10 production and transition toward a regulatory phenotype during coculture with BMSC. Taken together, these data demonstrate that systemic infusion of BMSC can durably alter the progression of radiation-induced fibrosis by altering macrophage phenotype and suppressing local inflammation in a TNF-R2-dependent fashion. © AlphaMed Press.

  16. Intra- and extracellular plasminogen activator inhibitor-1 regulate effect of vitronectin against radiation-induced endothelial cell death.

    PubMed

    Hazawa, Masaharu; Yasuda, Takeshi; Saotome-Nakamura, Ai; Tomiyama, Kenichi; Obara, Chizuka; Goto, Takaya; Tajima, Katsushi

    2016-12-01

    Plasminogen activator inhibitor-1 (PAI-1) is induced by radiation resulting in endothelial cell impairment, potentially leading to multiple organ failure. Vitronectin (VN) is a 75-kDa glycoprotein (VN75) cleaved into two forms (VN75 or VN65/10) by furin, which is regulated by intracellular PAI-1. VN protects against radiation-induced endothelial cell death, but the mechanisms involved in VN processing and its interactions with intra- and extracellular PAI-1 remain unclear. We examined these processes in cells in vitro using recombinant proteins or overexpression of VN and PAI-1 genes, including furin-susceptible (T(381)) and furin-resistant VN (A(381)). VN processing was analyzed using a mutant PAI-1 with relatively weaker binding to VN. VN function was evaluated by survival of radiation-damaged endothelial cells. Wild-type, but not mutant PAI-1 inhibited furin-dependent VN processing. Gene transfer revealed that furin-susceptible VN was processed more than the furin-resistant form, but processing of both was inhibited by PAI-1 overexpression. Intracellular PAI-1 formed a complex with VN75 (T(381)) in cells and media, and the VN75 form was secreted preferentially. Only VN75 protected against radiation-induced endothelial cell death, in which its effect was abolished by wild-type but not mutant PAI-1. These findings indicate that intracellular PAI-1 inhibits VN processing and protects against radiation-induced endothelial cell death. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Human CD14 mediates recognition and phagocytosis of apoptotic cells.

    PubMed

    Devitt, A; Moffatt, O D; Raykundalia, C; Capra, J D; Simmons, D L; Gregory, C D

    1998-04-02

    Cells undergoing programmed cell death (apoptosis) are cleared rapidly in vivo by phagocytes without inducing inflammation. Here we show that the glycosylphosphatidylinositol-linked plasma-membrane glycoprotein CD14 on the surface of human macrophages is important for the recognition and clearance of apoptotic cells. CD14 can also act as a receptor that binds bacterial lipopolysaccharide (LPS), triggering inflammatory responses. Overstimulation of CD14 by LPS can cause the often fatal toxic-shock syndrome. Here we show that apoptotic cells interact with CD14, triggering phagocytosis of the apoptotic cells. This interaction depends on a region of CD14 that is identical to, or at least closely associated with, a region known to bind LPS. However, apoptotic cells, unlike LPS, do not provoke the release of pro-inflammatory cytokines from macrophages. These results indicate that clearance of apoptotic cells is mediated by a receptor whose interactions with 'non-self' components (LPS) and 'self' components (apoptotic cells) produce distinct macrophage responses.

  18. Interplay of CREB and ATF2 in Ionizing Radiation-Induced Neuroendocrine Differentiation of Prostate Cancer Cells

    DTIC Science & Technology

    2012-06-01

    that contributes to IR- induced phos- phorylation of CREB. It will be interesting to determine whether this effect is independent of or dependent on...S. (2010) Growth factor stimulation induces cell survival by c- Jun. ATF2- dependent activation of Bcl-XL. J. Biol. Chem. 285, 23096–23104 7. Shimizu...08-1-0394 TITLE: Interplay of CREB and ATF2 in Ionizing Radiation- Induced Neuroendocrine Differentiation of Prostate Cancer Cells

  19. Caspase-independent cell death mediated by apoptosis-inducing factor (AIF) nuclear translocation is involved in ionizing radiation induced HepG2 cell death

    SciTech Connect

    Sun, Hengwen; Yang, Shana; Li, Jianhua; Zhang, Yajie; Gao, Dongsheng; Zhao, Shenting

    2016-03-25

    Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. The aim of radiotherapy is to eradicate cancer cells with ionizing radiation. Except for the caspase-dependent mechanism, several lines of evidence demonstrated that caspase-independent mechanism is directly involved in the cell death responding to irradiation. For this reason, defining the contribution of caspase-independent molecular mechanisms represents the main goal in radiotherapy. In this study, we focused on the role of apoptosis-inducing factor (AIF), the caspase-independent molecular, in ionizing radiation induced hepatocellular carcinoma cell line (HepG2) cell death. We found that ionizing radiation has no function on AIF expression in HepG2 cells, but could induce AIF release from the mitochondria and translocate into nuclei. Inhibition of AIF could reduce ionizing radiation induced HepG2 cell death. These studies strongly support a direct relationship between AIF nuclear translocation and radiation induced cell death. What's more, AIF nuclear translocation is caspase-independent manner, but not caspase-dependent manner, in this process. These new findings add a further attractive point of investigation to better define the complex interplay between caspase-independent cell death and radiation therapy. - Highlights: • AIF nuclear translocation is involved in ionizing radiation induced hepatocellular carcinoma cell line HepG2 cell death. • AIF mediated cell death induced by ionizing radiation is caspase-independent. • Caspase-independent pathway is involved in ionzing radiation induced HepG2 cell death.

  20. Diallyl disulfide enhances carbon ion beams-induced apoptotic cell death in cervical cancer cells through regulating Tap73 /ΔNp73.

    PubMed

    Di, Cuixia; Sun, Chao; Li, Hongyan; Si, Jing; Zhang, Hong; Han, Lu; Zhao, Qiuyue; Liu, Yang; Liu, Bin; Miao, Guoying; Gan, Lu; Liu, Yuanyuan

    2015-01-01

    Diallyl disulfide (DADS), extracted from crushed garlic by steam-distillation, has been reported to provide the anticancer activity in several cancer types. However, the effect of DADS on high-LET carbon beams - induced cell death remains unknown. Therefore, we used human cervical cancer cells to elucidate the molecular effects of this diallyl sulfide. Radiotherapy remains the mainstay of treatment, especially in advanced cervical cancer and there is still space to improve the radiosensitivity to reduce radiation dosage. In this study, we found that radiation effects evoked by high-LET carbon beam was marked by inhibition of cell viability, cell cycle arrest, significant rise of apoptotic cells, regulation of transcription factor, such as p73, as well as alterations of crucial mediator of the apoptosis pathway. We further demonstrated that pretreatment of 10 µM DADS in HeLa cells exposed to radiation resulted in decrease in cell viability and increased radiosensitivity. Additionally, cells pretreated with DADS obviously inhibited the radiation-induced G2/M phase arrest, but promoted radiation-induced apoptosis. Moreover, combination DADS and the radiation exacerbated the activation of apoptosis pathways through up-regulated ration of pro-apoptotic Tap73 to anti-apoptotic ΔNp73, and its downstream proteins, such as FASLG, and APAF1. Taken together, these results suggest that DADS is a potential candidate as radio sensitive agent for cervical cancer.

  1. Gene expression profiling in MOLT-4 cells during gamma-radiation-induced apoptosis.

    PubMed

    Lindgren, Theres; Stigbrand, Torgny; Riklund, Katrine; Johansson, Lennart; Eriksson, David

    2012-06-01

    This study aims to identify the temporal changes in gene expression in MOLT-4, a leukemia cell line, in response to radiation and to present a comprehensive description of the pathways and processes that most significantly relate to the cellular biological responses. A global gene expression profile of 24,500 genes was performed on MOLT-4 tumor cells following exposure to 5 Gy of ionizing radiation ((60)Co) using a bead chip array (Illumina). Signaling pathways and processes significantly altered following irradiation were explored using MetaCore. Cellular viability [3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide], activation of cell cycle checkpoints [fluorescence activated cell sorting (FACS)], and induction of apoptosis (FACS, caspase assays) were evaluated to correlate these biological responses to the gene expression changes. Totally, 698 different genes displayed a significantly altered expression following radiation, and out of these transcripts, all but one showed increased expression. One hour following irradiation, the expression was changed only for a few genes. Striking changes appeared at later time-points. From 3 to 24 h post-irradiation, a significant fraction of the genes with altered expression were found to be involved in cell cycle checkpoints and their regulation (CDKN1A), DNA repair (GADD45A, DDB2, XPC), apoptosis induction (DR5, FasR, Apo-2L, Bax), and T-cell activation/proliferation (CD70, OX40L). Irradiated MOLT-4 cells were arrested at the G2-checkpoint, followed by a decrease in cell viability, most pronounced 48 h after exposure. The cell death was executed by induced apoptosis and was visualized by an increase in subG1 cells and an increased activation of initiator (caspase-8 and caspase-9) and execution (caspase-3) caspases. Activation of cell cycle arrest and apoptosis correlated well in time with the changes in gene expression of those genes important for these biological processes. Activation of the apoptotic signaling

  2. Cell Cycle Regulators Guide Mitochondrial Activity in Radiation-Induced Adaptive Response

    PubMed Central

    Alexandrou, Aris T.

    2014-01-01

    Abstract Significance: There are accruing concerns on potential genotoxic agents present in the environment including low-dose ionizing radiation (LDIR) that naturally exists on earth's surface and atmosphere and is frequently used in medical diagnosis and nuclear industry. Although its long-term health risk is being evaluated and remains controversial, LDIR is shown to induce temporary but significant adaptive responses in mammalian cells and animals. The mechanisms guiding the mitochondrial function in LDIR-induced adaptive response represent a unique communication between DNA damage and cellular metabolism. Elucidation of the LDIR-regulated mitochondrial activity may reveal new mechanisms adjusting cellular function to cope with hazardous environmental stress. Recent Advances: Key cell cycle regulators, including Cyclin D1/CDK4 and Cyclin B1/cyclin-dependent kinase 1 (CDK1) complexes, are actively involved in the regulation of mitochondrial functions via phosphorylation of their mitochondrial targets. Accumulating new evidence supports a concept that the Cyclin B1/CDK1 complex acts as a mediator in the cross talk between radiation-induced DNA damage and mitochondrial functions to coordinate cellular responses to low-level genotoxic stresses. Critical Issues: The LDIR-mediated mitochondrial activity via Cyclin B1/CDK1 regulation is an irreplaceable network that is able to harmonize vital cellular functions with adjusted mitochondrial metabolism to enhance cellular homeostasis. Future Directions: Further investigation of the coordinative mechanism that regulates mitochondrial activities in sublethal stress conditions, including LDIR, will reveal new insights of how cells cope with genotoxic injury and will be vital for future targeted therapeutic interventions that reduce environmental injury and cancer risk. Antioxid. Redox Signal. 20, 1463–1480. PMID:24180340

  3. Cell cycle regulators guide mitochondrial activity in radiation-induced adaptive response.

    PubMed

    Alexandrou, Aris T; Li, Jian Jian

    2014-03-20

    There are accruing concerns on potential genotoxic agents present in the environment including low-dose ionizing radiation (LDIR) that naturally exists on earth's surface and atmosphere and is frequently used in medical diagnosis and nuclear industry. Although its long-term health risk is being evaluated and remains controversial, LDIR is shown to induce temporary but significant adaptive responses in mammalian cells and animals. The mechanisms guiding the mitochondrial function in LDIR-induced adaptive response represent a unique communication between DNA damage and cellular metabolism. Elucidation of the LDIR-regulated mitochondrial activity may reveal new mechanisms adjusting cellular function to cope with hazardous environmental stress. Key cell cycle regulators, including Cyclin D1/CDK4 and Cyclin B1/cyclin-dependent kinase 1 (CDK1) complexes, are actively involved in the regulation of mitochondrial functions via phosphorylation of their mitochondrial targets. Accumulating new evidence supports a concept that the Cyclin B1/CDK1 complex acts as a mediator in the cross talk between radiation-induced DNA damage and mitochondrial functions to coordinate cellular responses to low-level genotoxic stresses. The LDIR-mediated mitochondrial activity via Cyclin B1/CDK1 regulation is an irreplaceable network that is able to harmonize vital cellular functions with adjusted mitochondrial metabolism to enhance cellular homeostasis. Further investigation of the coordinative mechanism that regulates mitochondrial activities in sublethal stress conditions, including LDIR, will reveal new insights of how cells cope with genotoxic injury and will be vital for future targeted therapeutic interventions that reduce environmental injury and cancer risk.

  4. Radiation-Induced Bioradicals

    NASA Astrophysics Data System (ADS)

    Mondelaers, Win; Lahorte, Philippe

    This chapter is part one of a review in which the production and application of radiation-induced bioradicals is discussed. Bioradicals play a pivotal role in the complex chain of processes starting with the absorption of radiation in biological materials and ending with the radiation-induced biological after-effects. The general aspects of the four consecutive stages (physical, physicochemical, chemical and biological) are discussed from an interdisciplinary point of view. The close relationship between radiation dose and track structure, induced DNA damage and cell survival or killing is treated in detail. The repair mechanisms that cells employ, to insure DNA stability following irradiation, are described. Because of their great biomedical importance tumour suppressor genes involved in radiation-induced DNA repair and in checkpoint activation will be treated briefly, together with the molecular genetics of radiosensitivity. Part two of this review will deal with modern theoretical methods and experimental instrumentation for quantitative studies in this research field. Also an extensive overview of the applications of radiation-induced bioradicals will be given. A comprehensive list of references allows further exploration of this research field, characterised in the last decade by a substantial advance, both in fundamental knowledge and in range of applications.

  5. Radiation induced esophageal adenocarcinoma in a woman previously treated for breast cancer and renal cell carcinoma

    PubMed Central

    2012-01-01

    Background Secondary radiation-induced cancers are rare but well-documented as long-term side effects of radiation in large populations of breast cancer survivors. Multiple neoplasms are rare. We report a case of esophageal adenocarcinoma in a patient treated previously for breast cancer and clear cell carcinoma of the kidney. Case presentation A 56 year-old non smoking woman, with no alcohol intake and no familial history of cancer; followed in the National Institute of Oncology of Rabat Morocco since 1999 for breast carcinoma, presented on consultation on January 2011 with dysphagia. Breast cancer was treated with modified radical mastectomy, 6 courses of chemotherapy based on CMF regimen and radiotherapy to breast, inner mammary chain and to pelvis as castration. Less than a year later, a renal right mass was discovered incidentally. Enlarged nephrectomy realized and showed renal cell carcinoma. A local and metastatic breast cancer recurrence occurred in 2007. Patient had 2 lines of chemotherapy and 2 lines of hormonotherapy with Letrozole and Tamoxifen assuring a stable disease. On January 2011, the patient presented dysphagia. Oesogastric endoscopy showed middle esophagus stenosing mass. Biopsy revealed adenocarcinoma. No evidence of metastasis was noticed on computed tomography and breast disease was controlled. Palliative brachytherapy to esophagus was delivered. Patient presented dysphagia due to progressive disease 4 months later. Jejunostomy was proposed but the patient refused any treatment. She died on July 2011. Conclusion We present here a multiple neoplasm in a patient with no known family history of cancers. Esophageal carcinoma is most likely induced by radiation. However the presence of a third malignancy suggests the presence of genetic disorders. PMID:22873795

  6. Gamma tocotrienol, a potent radioprotector, preferentially upregulates expression of anti-apoptotic genes to promote intestinal cell survival.

    PubMed

    Suman, Shubhankar; Datta, Kamal; Chakraborty, Kushal; Kulkarni, Shilpa S; Doiron, Kathryn; Fornace, Albert J; Sree Kumar, K; Hauer-Jensen, Martin; Ghosh, Sanchita P

    2013-10-01

    Gamma tocotrienol (GT3) has been reported as a potent ameliorator of radiation-induced gastrointestinal (GI) toxicity when administered prophylactically. This study aimed to evaluate the role of GT3 mediated pro- and anti-apoptotic gene regulation in protecting mice from radiation-induced GI damage. Male 10- to 12-weeks-old CD2F1 mice were administered with a single dose of 200 mg/kg of GT3 or equal volume of vehicle (5% Tween-80) 24 h before exposure to 11 Gy of whole-body γ-radiation. Mouse jejunum was surgically removed 4 and 24h after radiation exposure, and was used for PCR array, histology, immunohistochemistry, and immunoblot analysis. Results were compared among vehicle pre-treated no radiation, vehicle pre-treated irradiated, and GT3 pre-treated irradiated groups. GT3 pretreated irradiated groups, both 4h and 24h after radiation, showed greater upregulation of anti-apoptotic gene expression than vehicle pretreated irradiated groups. TUNEL staining and intestinal crypt analysis showed protection of jejunum after GT3 pre-treatment and immunoblot results were supportive of PCR data. Our study demonstrated that GT3-mediated protection of intestinal cells from a GI-toxic dose of radiation occurred via upregulation of antiapoptotic and downregulation of pro-apoptotic factors, both at the transcript as well as at the protein levels.

  7. Radiation-induced glioma following CyberKnife® treatment of metastatic renal cell carcinoma: a case report

    PubMed Central

    2012-01-01

    Introduction Post-stereotactic radiation-induced neoplasms, although relatively rare, have raised the question of benefit regarding CyberKnife® treatments versus the risk of a secondary malignancy. The incidence of such neoplasms arising in the nervous system is thought to be low, given the paucity of case reports regarding such secondary lesions. Case presentation Here we describe a case of a 43-year-old Middle Eastern woman with primary clear cell renal cell carcinoma and a metastatic focus to the left brain parenchyma who presented with focal neurologic deficits. Following post-surgical stereotactic radiation in the region of the brain metastasis, the patient developed a secondary high-grade astrocytoma nearly 5 years after the initial treatment. Conclusion Although the benefit of CyberKnife® radiotherapy treatments continues to outweigh the relatively low risk of a radiation-induced secondary malignancy, knowledge of such risks and a review of the literature are warranted. PMID:22943305

  8. Role of autophagy in high linear energy transfer radiation-induced cytotoxicity to tumor cells

    PubMed Central

    Jin, Xiaodong; Liu, Yan; Ye, Fei; Liu, Xiongxiong; Furusawa, Yoshiya; Wu, Qingfeng; Li, Feifei; Zheng, Xiaogang; Dai, Zhongying; Li, Qiang

    2014-01-01

    Heavy-ion radiotherapy has a potential advantage over conventional radiotherapy due to improved dose distribution and a higher biological effectiveness in cancer therapy. However, there is a little information currently available on the cellular and molecular basis for heavy-ion irradiation-induced cell death. Autophagy, as a novel important target to improve anticancer therapy, has recently attracted considerable attention. In this study, the effect of autophagy induced by high linear energy transfer (LET) carbon ions was examined in various tumor cell lines. To our knowledge, our study is the first to reveal that high-LET carbon ions could induce autophagy in various tumor cells effectively, and the autophagic level in the irradiated cells increased in a dose- and LET-dependent manner. The ability of carbon ions to inhibit the activation of the PI3K/Akt pathway rose with increasing their LET. Moreover, modulation of autophagy in tumor cells could modify their sensitivity to high-LET radiation, and inhibiting autophagy accelerated apoptotic cell death, resulting in an increase in radiosensitivity. Our data imply that targeting autophagy might enhance the effectiveness of heavy-ion radiotherapy. PMID:24731006

  9. Glioblastoma cells block radiation-induced programmed cell death of endothelial cells.

    PubMed

    Brown, Charles K; Khodarev, Nikolai N; Yu, Jianqing; Moo-Young, Tricia; Labay, Edwardine; Darga, Thomas E; Posner, Mitchell C; Weichselbaum, Ralph R; Mauceri, Helena J

    2004-05-07

    We demonstrate that human umbilical vein endothelial cells (HUVEC) grown in co-culture (CC) with U87 glioblastoma cells transfected with green fluorescent protein (GFP-U87) exhibit resistance to radiation-mediated apoptosis. cDNA macroarray analysis reveals increases in the accumulation of RNAs for HUVEC genes encoding cell adhesion molecules, growth factor-related proteins, and cell cycle regulatory/DNA repair proteins. An increase in protein expression of integrin alphav, integrin beta1, MAPK(p42), Rad51, DNA-PK(CS), and ataxia telangiectasia gene (ATM) was detected in HUVEC grown in CC with GFP-U87 cells compared with HUVEC grown in mono-culture. Treatment with anti-VEGF antibody decreases the expression of integrin alphav, integrin beta1, DNA-PK(CS) and ATM with a corresponding increase in ionizing radiation (IR)-induced apoptosis. These data support the concept that endothelial cells growing in the tumor microenvironment may develop resistance to cytotoxic therapies due to the up-regulation by tumor cells of endothelial cells genes associated with survival.

  10. Surface code—biophysical signals for apoptotic cell clearance

    NASA Astrophysics Data System (ADS)

    Biermann, Mona; Maueröder, Christian; Brauner, Jan M.; Chaurio, Ricardo; Janko, Christina; Herrmann, Martin; Muñoz, Luis E.

    2013-12-01

    Apoptotic cell death and the clearance of dying cells play an important and physiological role in embryonic development and normal tissue turnover. In contrast to necrosis, apoptosis proceeds in an anti-inflammatory manner. It is orchestrated by the timed release and/or exposure of so-called ‘find-me’, ‘eat me’ and ‘tolerate me’ signals. Mononuclear phagocytes are attracted by various ‘find-me’ signals, including proteins, nucleotides, and phospholipids released by the dying cell, whereas the involvement of granulocytes is prevented via ‘stay away’ signals. The exposure of anionic phospholipids like phosphatidylserine (PS) by apoptotic cells on the outer leaflet of the plasma membrane is one of the main ‘eat me’ signals. PS is recognized by a number of innate receptors as well as by soluble bridging molecules on the surface of phagocytes. Importantly, phagocytes are able to discriminate between viable and apoptotic cells both exposing PS. Due to cytoskeleton remodeling PS has a higher lateral mobility on the surfaces of apoptotic cells thereby promoting receptor clustering on the phagocyte. PS not only plays an important role in the engulfment process, but also acts as ‘tolerate me’ signal inducing the release of anti-inflammatory cytokines by phagocytes. An efficient and fast clearance of apoptotic cells is required to prevent secondary necrosis and leakage of intracellular danger signals into the surrounding tissue. Failure or prolongation of the clearance process leads to the release of intracellular antigens into the periphery provoking inflammation and development of systemic inflammatory autoimmune disease like systemic lupus erythematosus. Here we review the current findings concerning apoptosis-inducing pathways, important players of apoptotic cell recognition and clearance as well as the role of membrane remodeling in the engulfment of apoptotic cells by phagocytes.

  11. Endoplasmic reticulum stress protects human thyroid carcinoma cell lines against ionizing radiation-induced apoptosis.

    PubMed

    Wu, Xin-Yu; Fan, Rui-Tai; Yan, Xin-Hui; Cui, Jing; Xu, Jun-Ling; Gu, Hao; Gao, Yong-Ju

    2015-03-01

    Radiotherapy is one of the most effective forms of cancer treatment, used in the treatment of a number of malignant tumors. However, the resistance of tumor cells to ionizing radiation remains a major therapeutic problem and the critical mechanisms determining radiation resistance are poorly defined. In the present study, a cellular endoplasmic reticulum (ER) stress microenvironment was established through the pretreatment of cultured thyroid cancer cells with tunicamycin (TM) and thapsigargin (TG), in order to mimic the ER stress response in a tumor microenvironment. This microenviroment was confirmed through the X‑box binding protein 1 splice process, glucose‑regulated protein 78 kD and ER degradation‑enhancing α‑mannosidase‑like mRNA expression. A clonogenic assay was used to measure cancer cell resistance to 60Co‑γ following TM pretreatment; in addition, human C/EBP homologous protein (CHOP) mRNA expression was determined and apoptosis assays were performed. The results showed that TM or TG pretreatment inhibited CHOP expression and reduced the apoptotic rate of cells. Furthermore, the results demonstrated that the induced ER stress response rendered cancer cells more resistant to ionizing radiation‑induced apoptosis. Therefore, the ER stress pathway may be a potential therapeutic target in order to improve the clinical efficiency of radiotherapy.

  12. The probabilities of one- and multi-track events for modeling radiation-induced cell kill.

    PubMed

    Schneider, Uwe; Vasi, Fabiano; Besserer, Jürgen

    2017-08-01

    In view of the clinical importance of hypofractionated radiotherapy, track models which are based on multi-hit events are currently reinvestigated. These models are often criticized, because it is believed that the probability of multi-track hits is negligible. In this work, the probabilities for one- and multi-track events are determined for different biological targets. The obtained probabilities can be used with nano-dosimetric cluster size distributions to obtain the parameters of track models. We quantitatively determined the probabilities for one- and multi-track events for 100, 500 and 1000 keV electrons, respectively. It is assumed that the single tracks are statistically independent and follow a Poisson distribution. Three different biological targets were investigated: (1) a DNA strand (2 nm scale); (2) two adjacent chromatin fibers (60 nm); and (3) fiber loops (300 nm). It was shown that the probabilities for one- and multi-track events are increasing with energy, size of the sensitive target structure, and dose. For a 2 × 2 × 2 nm(3) target, one-track events are around 10,000 times more frequent than multi-track events. If the size of the sensitive structure is increased to 100-300 nm, the probabilities for one- and multi-track events are of the same order of magnitude. It was shown that target theories can play a role for describing radiation-induced cell death if the targets are of the size of two adjacent chromatin fibers or fiber loops. The obtained probabilities can be used together with the nano-dosimetric cluster size distributions to determine model parameters for target theories.

  13. Suppression of radiation-induced migration of non-small cell lung cancer through inhibition of Nrf2-Notch Axis.

    PubMed

    Zhao, Qiuyue; Mao, Aihong; Guo, Ruoshui; Zhang, Liping; Yan, Jiawei; Sun, Chao; Tang, Jinzhou; Ye, Yancheng; Zhang, Yanshan; Zhang, Hong

    2017-03-28

    Nuclear factor E2 related factor 2 (Nrf2) is a transcription factor that is associated with tumor growth and resistance to radiation. The canonical Notch signaling pathway is also crucial for maintaining non-small cell lung cancer (NSCLC). Aberrant Nrf2 and Notch signaling has repeatedly been showed to facilitate metastasis of NSCLC. Here, we show that radiation induce Nrf2 and Notch1 expression in NSCLC. Knockdown of Nrf2 enhanced radiosensitivity of NSCLC and reduced epithelial-to-mesenchymal transition. Importantly, we found that knockdown of Nrf2 dramatically decreased radiation-induced NSCLC invasion and significantly increased E-cadherin, but reduced N-cadherin and matrix metalloproteinase (MMP)-2/9 expression. We found that Notch1 knockdown also upregulated E-cadherin and suppressed N-cadherin expression. Nrf2 contributes to NSCLC cell metastatic properties and this inhibition correlated with reduced Notch1 expression. These results establish that Nrf2 and Notch1 downregulation synergistically inhibit radiation-induced migratory and invasive properties of NSCLC cells.

  14. Marrow-derived stromal cell delivery on fibrin microbeads can correct radiation-induced wound-healing deficits.

    PubMed

    Xie, Michael W; Gorodetsky, Raphael; Micewicz, Ewa D; Micevicz, Ewa D; Mackenzie, Natalia C; Gaberman, Elena; Levdansky, Lilia; McBride, William H

    2013-02-01

    Skin that is exposed to radiation has an impaired ability to heal wounds. This is especially true for whole-body irradiation, where even moderate nonlethal doses can result in wound-healing deficits. Our previous attempts to administer dermal cells locally to wounds to correct radiation-induced deficits were hampered by poor cell retention. Here we improve the outcome by using biodegradable fibrin microbeads (FMBs) to isolate a population of mesenchymal marrow-derived stromal cells (MSCs) from murine bone marrow by their specific binding to the fibrin matrix, culture them to high density in vitro, and deliver them as MSCs on FMBs at the wound site. MSCs are retained locally, proliferate in site, and assist wounds in gaining tensile strength in whole-body irradiated mice with or without additional skin-only exposure. MSC-FMBs were effective in two different mouse strains but were ineffective across a major histocompatability barrier. Remarkably, irradiated mice whose wounds were treated with MSC-FMBs showed enhanced hair regrowth, suggesting indirect effect on the correction of radiation-induced follicular damage. Further studies showed that additional wound-healing benefit could be gained by administration of granulocyte colony-stimulating factor and AMD3100. Collagen strips coated with haptides and MSCs were also highly effective in correcting radiation-induced wound-healing deficits.

  15. Inhibition of p38 mitogen-activated protein kinase ameliorates radiation-induced ototoxicity in zebrafish and cochlea-derived cell lines.

    PubMed

    Shin, Yoo Seob; Hwang, Hye Sook; Kang, Sung Un; Chang, Jae Won; Oh, Young-Taek; Kim, Chul-Ho

    2014-01-01

    Radiation is a widely used treatment for head and neck cancers, and one of its most severe side effects is ototoxicity. Radiation-induced ototoxicity has been demonstrated to be linked to the increased production of ROS and MAPK. We intended to investigate the effect of p38 inhibition on radiation-induced ototoxicity in cochlea-derived HEI-OC1 cells and in a zebrafish model. The otoprotective effect of p38 inhibition against radiation was tested in vitro in the organ of Corti-derived cell line, HEI-OC1, and in vivo in a zebrafish model. Radiation-induced apoptosis, mitochondrial dysfunction, and an increase of intracellular NO generation were demonstrated in HEI-OC1 cells. The p38-specific inhibitor, SB203580, ameliorated radiation-induced apoptosis and mitochondrial injury in HEI-OC1 cells. p38 inhibition reduced radiation-induced activation of JNK, p38, cytochrome c, and cleavage of caspase-3 and PARP in HEI-OC1 cells. Scanning electron micrography showed that SB203580 prevented radiation-induced destruction of kinocilium and stereocilia in zebrafish neuromasts. The results of this study suggest that p38 plays an important role in mediating radiation-induced ototoxicity and inhibition of p38 could be a plausible option for preventing radiation ototoxicity. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. Carbocisteine promotes phagocytosis of apoptotic cells by alveolar macrophages.

    PubMed

    Inoue, Masako; Ishibashi, Yuji; Nogawa, Hisashi; Yasue, Tokutaro

    2012-02-29

    Clearance of apoptotic cells, so-called efferocytosis, by alveolar macrophages (AMs) is important for lung homeostasis and is impaired in pulmonary inflammatory diseases, such as chronic obstructive pulmonary disease and asthma. Carbocisteine, a mucoregulatory drug, corrects the contents of fucose in airway mucus and has anti-inflammatory properties in airway inflammation. Thus, we conducted the present study to better understand the anti-inflammatory properties of carbocisteine. First, we induced airway inflammation in mice with lipopolysaccharide intratracheally. Carbocisteine significantly decreased neutrophil numbers in bronchoalveolar lavage fluid at the resolution phase of inflammation, implying the promotion of neutrophil clearance. Then, we investigated whether carbocisteine would enhance the efferocytosis by AMs isolated from mice and found that this drug promoted not only the phagocytosis but also the binding of apoptotic cells to AMs in vitro. Furthermore, carbocisteine decreased the fucose residues stained with fluorescent fucose-binding lectin, Lens culinaris agglutinin, on the cell surface of AMs. We found here that removing fucose residues from cell surfaces of AMs by fucosidase markedly enhanced both the binding and phagocytosis of apoptotic cells. Finally, AMs from mice orally given carbocisteine also promoted both the binding and phagocytosis ex vivo similarly to in vitro. These results suggest that carbocisteine could promote the clearance of apoptotic cells by AMs in airway. In addition, the present findings suggest that the binding and phagocytosis of apoptotic cells may be modulated by fucose residues on the cell surface of AMs.

  17. Phagocytosis Assay for Apoptotic Cells in Drosophila Embryos.

    PubMed

    Nonaka, Saori; Hori, Aki; Nakanishi, Yoshinobu; Kuraishi, Takayuki

    2017-08-03

    The molecular mechanisms underlying the phagocytosis of apoptotic cells need to be elucidated in more detail because of its role in immune and inflammatory intractable diseases. We herein developed an experimental method to investigate phagocytosis quantitatively using the fruit fly Drosophila, in which the gene network controlling engulfment reactions is evolutionally conserved from mammals. In order to accurately detect and count engulfing and un-engulfing phagocytes using whole animals, Drosophila embryos were homogenized to obtain dispersed cells including phagocytes and apoptotic cells. The use of dispersed embryonic cells enables us to measure in vivo phagocytosis levels as if we performed an in vitro phagocytosis assay in which it is possible to observe all phagocytes and apoptotic cells in whole embryos and precisely quantify the level of phagocytosis. We confirmed that this method reproduces those of previous studies that identified the genes required for the phagocytosis of apoptotic cells. This method allows the engulfment of dead cells to be analyzed, and when combined with the powerful genetics of Drosophila, will reveal the complex phagocytic reactions comprised of the migration, recognition, engulfment, and degradation of apoptotic cells by phagocytes.

  18. Innate and Adaptive Immune Response to Apoptotic Cells

    PubMed Central

    Peng, YuFeng; Martin, David A; Kenkel, Justin; Zhang, Kang; Ogden, Carol Anne; Elkon, Keith B.

    2007-01-01

    The immune system is constantly exposed to dying cells, most of which arise during central tolerance and from effete circulating immune cells. Under homeostatic conditions, phagocytes (predominantly macrophages and dendritic cells) belonging to the innate immune system, rapidly ingest cells and their debris. Apoptotic cell removal requires recognition of altered self on the apoptotic membrane, a process which is facilitated by natural antibodies and serum opsonins. Recognition, may be site and context specific. Uptake and ingestion of apoptotic cells promotes an immunosuppressive environment that avoids inflammatory responses to self antigens. However, it does not preclude a T cell response and it is likely that constant exposure to self antigen, particularly by immature dendritic cells, leads to T cell tolerance. Tolerance occurs by several different mechanisms including anergy and deletion (for CD8+ T cells) and induction of T regulatory cells (for CD4+ T cells). Failed apoptotic cell clearance promotes immune responses to self antigens, especially when the cellular contents are leaked from the cell (necrosis). Inflammatory responses may be induced by nucleic acid stimulation of toll like receptors and other immune sensors, specific intracellular proteins and non protein (uric acid) stimulation of inflammasomes. PMID:17888627

  19. Ribosome Synthesis and MAPK Activity Modulate Ionizing Radiation-Induced Germ Cell Apoptosis in Caenorhabditis elegans

    PubMed Central

    Eberhard, Ralf; Stergiou, Lilli; Hofmann, E. Randal; Hofmann, Jen; Haenni, Simon; Teo, Youjin; Furger, André; Hengartner, Michael O.

    2013-01-01

    Synthesis of ribosomal RNA by RNA polymerase I (RNA pol I) is an elemental biological process and is key for cellular homeostasis. In a forward genetic screen in C. elegans designed to identify DNA damage-response factors, we isolated a point mutation of RNA pol I, rpoa-2(op259), that leads to altered rRNA synthesis and a concomitant resistance to ionizing radiation (IR)-induced germ cell apoptosis. This weak apoptotic IR response could be phenocopied when interfering with other factors of ribosome synthesis. Surprisingly, despite their resistance to DNA damage, rpoa-2(op259) mutants present a normal CEP-1/p53 response to IR and increased basal CEP-1 activity under normal growth conditions. In parallel, rpoa-2(op259) leads to reduced Ras/MAPK pathway activity, which is required for germ cell progression and physiological germ cell death. Ras/MAPK gain-of-function conditions could rescue the IR response defect in rpoa-2(op259), pointing to a function for Ras/MAPK in modulating DNA damage-induced apoptosis downstream of CEP-1. Our data demonstrate that a single point mutation in an RNA pol I subunit can interfere with multiple key signalling pathways. Ribosome synthesis and growth-factor signalling are perturbed in many cancer cells; such an interplay between basic cellular processes and signalling might be critical for how tumours evolve or respond to treatment. PMID:24278030

  20. Apoptotic cell death in rat epididymis following epichlorohydrin treatment.

    PubMed

    Lee, I-C; Kim, K-H; Kim, S-H; Baek, H-S; Moon, C; Kim, S-H; Yun, W-K; Nam, K-H; Kim, H-C; Kim, J-C

    2013-06-01

    Epichlorohydrin (ECH) is an antifertility agent that acts both as an epididymal toxicant and an agent capable of directly affecting sperm motility. This study identified the time course of apoptotic cell death in rat epididymides after ECH treatment. Rats were administrated with a single oral dose of ECH (50 mg/kg). ECH-induced apoptotic changes were evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and its related mechanism was confirmed by Western blot analysis and colorimetric assay. The TUNEL assay showed that the number of apoptotic cells increased at 8 h, reached a maximum level at 12 h, and then decreased progressively. The Western blot analysis demonstrated no significant changes in proapoptotic Bcl-2-associated X (Bax) and anti-apoptotic Bcl-2 expression during the time course of the study. However, phospho-p38 mitogen-activated protein kinase (p-p38 MAPK) and phospho-c-Jun amino-terminal kinase (p-JNK) expression increased at 8-24 h. Caspase-3 and caspase-8 activities also increased at 8-48 h and 12-48 h, respectively, in the same manner as p-p38 MAPK and p-JNK expression. These results indicate that ECH induced apoptotic changes in rat epididymides and that the apoptotic cell death may be related more to the MAPK pathway than to the mitochondrial pathway.

  1. Dose-dependency and reversibility of radiation-induced injury in cardiac explant-derived cells of mice

    PubMed Central

    Luo, Lan; Yan, Chen; Urata, Yoshishige; Hasan, Al Shaimaa; Goto, Shinji; Guo, Chang-Ying; Zhang, Shouhua; Li, Tao-Sheng

    2017-01-01

    We evaluated the dose-dependency and reversibility of radiation-induced injury in cardiac explant-derived cells (CDCs), a mixed cell population grown from heart tissues. Adult C57BL/6 mice were exposed to 0, 10, 50 and 250 mGy γ-rays for 7 days and atrial tissues were collected for experiments 24 hours after last exposure. The number of CDCs was significantly decreased by daily exposure to over 250 mGy. Interestingly, daily exposure to over 50 mGy significantly decreased the c-kit expression and telomerase activity, increased 53BP1 foci in the nuclei of CDCs. However, CD90 expression and growth factors production in CDCs were not significantly changed even after daily exposure to 250 mGy. We further evaluated the reversibility of radiation-induced injury in CDCs at 1 week and 3 weeks after a single exposure to 3 Gy γ-rays. The number and growth factors production of CDCs were soon recovered at 1 week. However, the increased expression of CD90 were retained at 1 week, but recovered at 3 weeks. Moreover, the decreased expression of c-kit, impaired telomerase activity, and increased 53BP1 foci were poorly recovered even at 3 weeks. These data may help us to find the most sensitive and reliable bio-parameter(s) for evaluating radiation-induced injury in CDCs. PMID:28098222

  2. Effects of Caffeine on Radiation-Induced Phenomena Associated with Cell-Cycle Traverse of Mammalian Cells

    PubMed Central

    Walters, Ronald A.; Gurley, Lawrence R.; Tobey, Robert A.

    1974-01-01

    Caffeine induced a state of G1 arrest when added to an exponentially growing culture of Chinese hamster cells (line CHO). In addition to its effect on cell-cycle traverse, caffeine ameliorated a number of the responses of cells to ionizing radiation. The duration of the division delay period following X-irradiation of caffeine-treated cells was reduced, and the magnitude of reduction was dependent on caffeine concentration. Cells irradiated during the DNA synthetic phase in the presence of caffeine were delayed less in their exit from S, measured autoradiographically, and the radiation-induced reduction of radioactive thymidine incorporation into DNA was lessened. Cells synchronized by isoleucine deprivation, while being generally less sensitive to the effects of ionizing radiation than mitotically synchronized cells, were equally responsive to the effects of caffeine. The X-ray-induced reduction of phosphorylation of lysine-rich histone F1 was less in caffeine-treated cells than in untreated cells. Finally, survival after irradiation was only slightly reduced in caffeine-treated cells. A possible role of cyclic AMP in cell-cycle traverse of irradiated cells is discussed. PMID:4360269

  3. Visualizing the effect of tumor microenvironments on radiation-induced cell kinetics in multicellular spheroids consisting of HeLa cells

    SciTech Connect

    Kaida, Atsushi; Miura, Masahiko

    2013-10-04

    Highlights: •We visualized radiation-induced cell kinetics in spheroids. •HeLa-Fucci cells were used for detection of cell-cycle changes. •Radiation-induced G2 arrest was prolonged in the spheroid. •The inner and outer cell fractions behaved differently. -- Abstract: In this study, we visualized the effect of tumor microenvironments on radiation-induced tumor cell kinetics. For this purpose, we utilized a multicellular spheroid model, with a diameter of ∼500 μm, consisting of HeLa cells expressing the fluorescent ubiquitination-based cell-cycle indicator (Fucci). In live spheroids, a confocal laser scanning microscope allowed us to clearly monitor cell kinetics at depths of up to 60 μm. Surprisingly, a remarkable prolongation of G2 arrest was observed in the outer region of the spheroid relative to monolayer-cultured cells. Scale, an aqueous reagent that renders tissues optically transparent, allowed visualization deeper inside spheroids. About 16 h after irradiation, a red fluorescent cell fraction, presumably a quiescent G0 cell fraction, became distinct from the outer fraction consisting of proliferating cells, most of which exhibited green fluorescence indicative of G2 arrest. Thereafter, the red cell fraction began to emit green fluorescence and remained in prolonged G2 arrest. Thus, for the first time, we visualized the prolongation of radiation-induced G2 arrest in spheroids and the differences in cell kinetics between the outer and inner fractions.

  4. Profiling mitochondrial proteins in radiation-induced genome-unstable cell lines with persistent oxidative stress by mass spectrometry

    SciTech Connect

    Miller, John H.; Jin, Shuangshuang; Morgan, William F.; Yang, Austin; Wan, Yunhu; Aypar, Umut; Peters, Jonathan S.; Springer, David L.

    2008-06-01

    Radiation-induced genome instability (RIGI) is a response to radiation exposure in which the progeny of surviving cells exhibit increased frequency of chromosomal changes many generations after the initial insult. Persistently elevated oxidative stress accompanying RIGI and the ability of free-radical scavengers, given before irradiation, to reduce the incidence of instability suggest that radiation induced alterations to mitochondrial function likely play a role in RIGI. To further elucidate this mechanism, we performed high-throughput quantitative mass spectrometry on samples enriched in mitochondrial proteins from three chromosomally-unstable GM10115 Chinese-hamster-ovary cell lines and their stable parental cell line. Out of several hundred identified proteins, sufficient data were collected on 74 mitochondrial proteins to test for statistically significant differences in their abundance between unstable and stable cell lines. Each of the unstable cell lines showed a distinct profile of statistically-significant differential abundant mitochondrial proteins. The LS-12 cell line was characterized by 8 downregulated proteins, whereas the CS-9 cell line exhibited 5 distinct up-regulated proteins. The unstable 115 cell line had two down-regulated proteins, one of which was also downregulated in LS-12, and one up-regulated protein relative to stable parental cells. The mitochondrial protein profiles for LS-12 and C-9 provide further evidence that mitochondrial dysfunction is involved in the genome instability of these cell lines.

  5. Rho kinase regulates fragmentation and phagocytosis of apoptotic cells

    SciTech Connect

    Orlando, Kelly A.; Stone, Nicole L.; Pittman, Randall N. . E-mail: pittman@pharm.med.upenn.edu

    2006-01-01

    During the execution phase of apoptosis, a cell undergoes cytoplasmic and nuclear changes that prepare it for death and phagocytosis. The end-point of the execution phase is condensation into a single apoptotic body or fragmentation into multiple apoptotic bodies. Fragmentation is thought to facilitate phagocytosis; however, mechanisms regulating fragmentation are unknown. An isoform of Rho kinase, ROCK-I, drives membrane blebbing through its activation of actin-myosin contraction; this raises the possibility that ROCK-I may regulate other execution phase events, such as cellular fragmentation. Here, we show that COS-7 cells fragment into a number of small apoptotic bodies during apoptosis; treating with ROCK inhibitors (Y-27632 or H-1152) prevents fragmentation. Latrunculin B and blebbistatin, drugs that interfere with actin-myosin contraction, also inhibit fragmentation. During apoptosis, ROCK-I is cleaved and activated by caspases, while ROCK-II is not activated, but rather translocates to a cytoskeletal fraction. siRNA knock-down of ROCK-I but not ROCK-II inhibits fragmentation of dying cells, consistent with ROCK-I being required for apoptotic fragmentation. Finally, cells dying in the presence of the ROCK inhibitor Y-27632 are not efficiently phagocytized. These data show that ROCK plays an essential role in fragmentation and phagocytosis of apoptotic cells.

  6. Amelioration of radiation-induced hematopoietic syndrome by an antioxidant chlorophyllin through increased stem cell activity and modulation of hematopoiesis.

    PubMed

    Suryavanshi, Shweta; Sharma, Deepak; Checker, Rahul; Thoh, Maikho; Gota, Vikram; Sandur, Santosh K; Sainis, Krishna B

    2015-08-01

    Hematopoietic stem cells and progenitor cells (HSPC) are low in abundance and exhibit high radiosensitivity and their ability to divide dramatically decreases following exposure to ionizing radiation. Our earlier studies have shown antiapoptotic, immune-stimulatory, and antioxidant effects of chlorophyllin, a constituent of the over the counter drug derifil. Here we describe the beneficial effects of chlorophyllin against radiation-induced hematopoietic syndrome. Chlorophyllin administration significantly enhanced the abundance of HSPC in vivo. It induced a transient cell cycle arrest in lineage-negative cells in the bone marrow. However, the chlorophyllin-treated mice exposed to whole body irradiation (WBI) had a significantly higher proportion of actively dividing HSPC in the bone marrow as compared to only WBI-exposed mice. It significantly increased the number of colony forming units (CFUs) by bone marrow cells in vitro and spleen CFUs in irradiated mice in vivo. Pharmacokinetic study showed that chlorophyllin had a serum half-life of 141.8 min in mice. Chlorophyllin upregulated antiapoptotic genes and antioxidant machinery via activation of prosurvival transcription factors Nrf-2 and NF-κB and increased the survival and recovery of bone marrow cells in mice exposed to WBI. Chlorophyllin stimulated granulocyte production in bone marrow and increased the abundance of peripheral blood neutrophils by enhancing serum levels of granulocyte-colony stimulation factor (GCSF). Most importantly, prophylactic treatment of mice with chlorophyllin significantly abrogated radiation-induced mortality. Chlorophyllin mitigates radiation-induced hematopoietic syndrome by increasing the abundance of hematopoietic stem cells, enhancing granulopoiesis, and stimulating prosurvival pathways in bone marrow cells and lymphocytes.

  7. The Sound of Silence: Signaling by Apoptotic Cells

    PubMed Central

    Fogarty, Caitlin E.; Bergmann, Andreas

    2016-01-01

    Apoptosis is a carefully choreographed process of cellular self-destruction in the absence of inflammation. During the death process, apoptotic cells actively communicate with their environment, signaling to both their immediate neighbors as well as distant sentinels. Some of these signals direct the anti-inflammatory immune response, instructing specific subsets of phagocytes to participate in the limited and careful clearance of dying cellular debris. These immunomodulatory signals can also regulate the activation state of the engulfing phagocytes. Other signals derived from apoptotic cells contribute to tissue growth control with the common goal of maintaining tissue integrity. Derangements in these growth control signals during prolonged apoptosis can lead to excessive cell loss or proliferation. Here, we highlight some of the most intriguing signals produced by apoptotic cells during the course of normal development as well as during physiological disturbances such as atherosclerosis and cancer. PMID:26431570

  8. Lack of evidence for low-LET radiation induced bystander response in normal human fibroblasts and colon carcinoma cells

    SciTech Connect

    Marianne B. Sowa; Wilfried Goetz; Janet E. Baulch; Dinah N. Pyles; Jaroslaw Dziegielewski; Susannah Yovino; Andrew R. Snyder; Sonia M. de Toledo; Edouard I. Azzam; William F. Morgan

    2008-06-30

    Purpose: To investigate radiation induced bystander responses and to determine the role of gap junction intercellular communication and the radiation environment in propagating this response. Materials and Methods: We use medium transfer and targeted irradiation to examine radiation induced bystander effects in primary human fibroblast (AG1522) and human colon carcinoma (RKO36) cells. We examined the effect of variables such as gap junction intercellular communication, linear energy transfer (LET), and the role of the radiation environment in non-targeted responses. Endpoints included clonogenic survival, micronucleus formation and foci formation at histone 2AX over doses ranging from 10 to 100 cGy. Results: The results show no evidence of a low-LET radiation induced bystander response for the endpoints of clonogenic survival and induction of DNA damage. Nor do we see evidence of a high-LET, Fe ion radiation (1 GeV/n) induced bystander effect. However, direct comparison for 3.2 MeV α-particle exposures showed a statistically significant medium transfer bystander effect for this high-LET radiation. Conclusions: From our results, it is evident that there are many confounding factors influencing bystander responses as reported in the literature. Our observations reflect the inherent variability in biological systems and the difficulties in extrapolating from in vitro models to radiation risks in humans.

  9. Bone marrow mesenchymal stem cell transplantation improves radiation-induced heart injury through DNA damage repair in rat model.

    PubMed

    Gao, Song; Zhao, Zhiying; Wu, Rong; Zeng, Yuecan; Zhang, Zhenyong; Miao, Jianing; Yuan, Zhengwei

    2017-03-01

    Radiotherapy is an effective form of therapy for most thoracic malignant tumors. However, myocardial injury resulting from the high doses of radiation is a severe complication. Here we aimed to study the possibility of reducing radiation-induced myocardial injury with mesenchymal stem cell (MSC) transplantation. We used MSCs extracted from bone marrow (BMSCs) to transplant via the tail vein into a radiation-induced heart injury (RIHI) rat model. The rats were divided into six groups: a Sham group, an IRR (irradiation) group, and four IRR + BMSCs transplantation groups obtained at different time points. After irradiation, BMSC transplantation significantly enhanced the cardiac function in rats. By analyzing the expression of PPAR-α, PPAR-γ, TGF-β, IL-6, and IL-8, we found that BMSC transplantation alleviated radiation-induced myocardial fibrosis and decreased the inflammatory reaction. Furthermore, we found that expression of γ-H2AX, XRCC4, DNA ligase4, and TP53BP1, which are associated with DNA repair, was up-regulated, along with increased secretion of growth factors SDF-1, CXCR4, VEGF, and IGF in rat myocardium in the IRR + BMSCs transplantation groups compared with the IRR group. Thus, BMSC transplantation has the potential to improve RIHI via DNA repair and be a new therapeutic approach for patients with myocardial injury.

  10. Different Roles of CHOP and JNK in Mediating Radiation-Induced Autophagy and Apoptosis in Breast Cancer Cells.

    PubMed

    Li, Feifei; Zheng, Xiaogang; Liu, Yan; Li, Ping; Liu, Xiongxiong; Ye, Fei; Zhao, Ting; Wu, Qingfeng; Jin, Xiaodong; Li, Qiang

    2016-05-01

    Unfolded protein response (UPR) is comprised of complex and conserved stress pathways that function as a short-term adaptive mechanism to reduce levels of unfolded or misfolded proteins and maintain homeostasis in the endoplasmic reticulum (ER). UPR can be triggered by prolonged or persistent ER stress under many physiological or pathological conditions, including radiation exposure. Radiation-induced ER stress elicits autophagy and apoptosis in cancer cells, where C/EBP homologous protein (CHOP) and c-Jun NH2-terminal kinase (JNK) may play crucial roles. However, the specific mechanisms that regulate autophagy and apoptosis through CHOP and JNK after radiation exposure and how the balance of these activities determines the cellular radiosensitivity remain largely unclear. In this study, we found that exposure to X-ray radiation induced ER stress, UPR and high expression of CHOP and JNK. Furthermore, autophagy and apoptosis occurred in sequential order when breast cancer MDA-MB-231 and MCF-7 cells were exposed to X-ray radiation. CHOP gene knockdown with RNA interference inhibited autophagy and enhanced radiosensitivity in MDA-MB-231 cells, while impacting apoptosis and subsequently increasing radioresistance in MCF-7 cells. However, treatment with JNK inhibitor decreased autophagy while promoting apoptosis, thereby leading to radiosensitivity in both cell lines. Our results indicate that CHOP mediates radiation-induced autophagy and apoptosis in a cellular environment. Importantly, the functional consistency of regulating apoptosis and autophagy in these two irradiated breast cancer cell lines suggests that JNK may be more useful as a potential target for maximizing the efficacy of radiation therapy for breast cancers.

  11. Radiation damage and repair in cells and cell components. Radiation-induced repair. Progress report, 1981-1982

    SciTech Connect

    Not Available

    1982-01-01

    Progress in research on the description and interpretation of radiation-induced repair in cells is reported. It has been found that for the p-recA data induction seems to follow a model of fractional site occupancy rather than being all-or-none. Other areas investigated include: (1) the induction of the RecA-gene product; (2) the effect of uv-phage lambda infection on Rec-A protein synthesis; (3) induced uv radioresistance; (4) cold-shock effects; (5) lambda-prophage induction by x-rays and uv; (6) photoreactivation of uv-induced dimers; and (7) a comparative study of S.O.S. phenomena in various strains of E. coli. (ACR)

  12. Dragon's blood extracts reduce radiation-induced peripheral blood injury and protects human megakaryocyte cells from GM-CSF withdraw-induced apoptosis.

    PubMed

    Ran, Yuanyuan; Xu, Bing; Wang, Ran; Gao, Qian; Jia, Qiutian; Hasan, Murtaza; Shan, Shuangquan; Ma, Hong; Dai, Rongji; Deng, Yulin; Qing, Hong

    2016-01-01

    Dragon's blood (DB), a Chinese traditional herb, was shown to have certain protective effects on radiation-induced bone marrow injury due to the presence of several phenolic compounds. The 50% ethanol extracts (DBE) were separated from DB by the methods of alcohol extracting-water precipitating. The protective effects of DBE on hematopoiesis were studied, particularly on megakaryocytes. In this study, we investigated the in vivo radioprotective effects of DBE on hematopoiesis and pathological changes using an irradiated-mouse model. Moreover, the protective effects and potential molecular mechanisms of DBE on megakaryocytopoiesis in vitro were explored in GM-CSF depletion-induced Mo7e cell model. DBE significantly promoted the recovery of peripheral blood cells in irradiated mice. Histology bone marrow confirmed the protective effect of DBE, as shown by an increased number of hematopoietic cells and a reduction of apoptosis. In a megakaryocytic apoptotic model, DBE (50 µg/mL) markedly alleviated GM-CSF withdrawal-induced apoptosis and cell-cycle arrest of Mo7e cells. DBE (50 µg/mL) also significantly decreased the ratio of Bax to Bcl-2 expression, inhibited the active caspase-3 expression. In addition, DBE could induce ERK1/2 phosphorylation in GM-CSF-depleted Mo7e cell, but not Akt. Our data demonstrated that DBE could effectively accelerate the recovery of peripheral blood cells, especially platelet. DBE attenuated cell apoptosis and cell cycle arrest through the decrease of Bax/Bcl-2 ratio and the reduction of active caspase-3 expression. The effect of DBE on Mo7e cells survival and proliferation is likely associated with the activation of ERK, but not Akt. Copyright © 2015 Associazione Italiana di Fisica Medica. Published by Elsevier Ltd. All rights reserved.

  13. Monitoring circulating apoptotic cells by in-vivo flow cytometry

    NASA Astrophysics Data System (ADS)

    Wei, Xunbin; Tan, Yuan; Chen, Yun; Zhang, Li; Li, Yan; Liu, Guangda; Wu, Bin; Wang, Chen

    2008-02-01

    Chemotherapies currently constitute one main venue of cancer treatment. For a large number of adult and elderly patients, however, treatment options are poor. These patients may suffer from disease that is resistant to conventional chemotherapy or may not be candidates for curative therapies because of advanced age or poor medical conditions. To control disease in these patients, new therapies must be developed that are selectively targeted to unique characteristics of tumor cell growth and metastasis. A reliable early evaluation and prediction of response to the chemotherapy is critical to its success. Chemotherapies induce apoptosis in tumor cells and a portion of such apoptotic cancer cells may be present in the circulation. However, the fate of circulating tumor cells is difficult to assess with conventional methods that require blood sampling. We report the in situ measurement of circulating apoptotic cells in live animals using in vivo flow cytometry, a novel method that enables real-time detection and quantification of circulating cells without blood extraction. Apoptotic cells are rapidly cleared from the circulation with a half-life of ~10 minutes. Real-time monitoring of circulating apoptotic cells can be useful for detecting early changes in disease processes, as well as for monitoring response to therapeutic intervention.

  14. Effects of N-acetylcysteine amide (NACA), a thiol antioxidant on radiation-induced cytotoxicity in Chinese hamster ovary cells.

    PubMed

    Wu, Wei; Abraham, Linu; Ogony, Joshua; Matthews, Richard; Goldstein, Glenn; Ercal, Nuran

    2008-05-23

    Ionizing radiation is known to cause tissue damage in biological systems, mainly due to its ability to produce reactive oxygen species (ROS) in cells. Many thiol antioxidants have been used previously as radioprotectors, but their application has been limited by their toxicity. In this investigation, we have explored the possible radioprotective effects of a newly synthesized thiol antioxidant, N-acetylcysteine amide (NACA), in comparison with N-acetylcysteine (NAC), a commonly used antioxidant. Protective effects of NACA and NAC were assessed using Chinese hamster ovary (CHO) cells, irradiated with 6 gray (Gy) radiation. Oxidative stress parameters, including levels of reduced glutathione (GSH), cysteine, malondialdehyde (MDA), and activities of antioxidant enzymes like glutathione peroxidase, glutathione reductase, and catalase, were measured. Results indicate that NACA was capable of restoring GSH levels in irradiated cells in a dose dependent manner. In addition, NACA prevented radiation-induced loss in cell viability. NACA further restored levels of malondialdehyde, caspase-3 activity, and antioxidant enzyme activities to control levels. Although NAC affected cells in a similar manner to NACA, its effects were not as significant. Further, NAC was also found to be cytotoxic to cells at higher concentrations, whereas NACA was non-toxic at similar concentrations. These results suggest that NACA may be able to attenuate radiation-induced cytotoxicity, possibly by its ability to provide thiols to cells.

  15. Millimeter wave radiation induces apoptosis via affecting the ratio of Bax/Bcl-2 in SW1353 human chondrosarcoma cells.

    PubMed

    Li, Xihai; Ye, Hongzhi; Cai, Liangliang; Yu, Fangrong; Chen, Wenlie; Lin, Ruhui; Zheng, Chunsong; Xu, Huifeng; Ye, Jinxia; Wu, Guangwen; Liu, Xianxiang

    2012-03-01

    The efficacy and safety of millimeter wave radiation has been proven for various types of malignant tumors. However, the mechanisms underlying effects of millimeter wave radiation on apoptosis are still unclear. The present study was undertaken to examine the effects of millimeter wave radiation on cell apoptosis and mitochondrial membrane potential, and to determine the molecular mechanism of millimeter wave radiation-induced apoptosis by investigating the expression of Bcl-2 family proteins (Bcl-2, Bax), caspase-9 and caspase-3 in SW1353 cells. We found that millimeter wave radiation suppressed the viability of SW1353 cells, demonstrating that millimeter wave radiation induced cell apoptosis and reduced cell viability in a time-dependent manner. Furthermore, we observed that treatment of cells with millimeter wave radiation significantly induced loss of mitochondrial membrane potential, upregulated proapoptotic Bax, caspase-9 and caspase-3, but did not significantly change levels of antiapoptotic Bcl-2. These data suggested that millimeter wave radiation may induce apoptosis via affecting the ratio of Bax/Bcl-2 in SW1353 cells.

  16. Apoptotic lymphocytes induce progenitor cell mobilization after exercise.

    PubMed

    Mooren, Frank C; Krüger, Karsten

    2015-07-15

    There is evidence that apoptotic cells and their components have immunmodulatory properties and signaling function. The present study investigated first whether exercise-induced apoptosis and exercise-induced mobilization of progenitor cells are similarly affected by subjects' training status and, second, whether the appearance of dying cells in the circulation might mobilize progenitor cells. CD1 SWISS mice were subjected to a 10-wk endurance training using free wheel running or served as untrained controls. Mice of both groups performed an intensive exercise test after the training period at a velocity corresponding to 80% maximal oxygen uptake for 30 min. Cells from blood and bone marrow were analyzed, and apoptosis and number of progenitor cells determined via flow cytometry. In a second experiment, apoptotic cells were transferred into recipient mice, and mobilization of progenitor cells was analyzed while vital cells served as controls. In untrained animals, the exhaustive exercise was followed by an enhanced rate of annexin V positive CD3(+) cells in blood and bone marrow (P < 0.05), whereas no increase was found in trained mice. Similarly, exercise mobilized Sca-1(+)/c-kit(+) and Sca-1(+)/Flk(+) cells in untrained (P < 0.05) but not trained mice. Furthermore, application of apoptotic cells and their supernatant mobilized Sca-1(+)/c-kit(+) cells into the blood (P < 0.05), whereas Sca-1(+)/Flk(+) cells were not affected. The present study demonstrated that both lymphocyte apoptosis, as well as mobilization of progenitor cells are similarly related to training status. Furthermore, apoptotic cells seem to induce signals that effectively mobilize hematopoietic progenitor cells. The relevance of this effect for the adaptation to exercise stimuli remains to be shown. Copyright © 2015 the American Physiological Society.

  17. Radiation induced secretion of surfactant from cell cultures of type II pneumocytes: an in vitro model of radiation toxicity

    SciTech Connect

    Shapiro, D.L.; Finkelstein, J.N.; Rubin, P.; Penney, D.P.; Siemann, D.W.

    1984-03-01

    The pathogenesis of pneumonitis and fibrosis secondary to lung irradiation is incompletely understood. The role of the type II alveolar epithelial pneumocyte in these processes has been under investigation. The type II pneumocyte has been shown in vivo to respond to radiation induced injury with release of pulmonary surfactant. The effect of irradiation on cell cultures of type II pneumocytes was studied to determine if this could be reproduced in vitro. Type II pneumocytes were found to release surfactant material with a threshold of radiation dose between 1000 and 1500 rad. This is similar to the dosage range over which the same effect has been demonstrated in vivo. Experimental results support the concept that the release of surfactant is not due to either cell disruption or non-specific release of phospholipid from cell membranes. Irradiation appears to trigger membrane receptor mediated surfactant release. In addition, irradiation abolishes the ability of cells to subsequently respond to a physiologic agonist, suggesting radiation induced damage to the secretory mechanism. These studies establish that surfactant release in response to irradiation in vivo is a direct effect on type II pneumocytes. Cell cultures of type II pneumocytes can serve as a laboratory model of lung cell radiation toxicity.

  18. Radiation-induced interphase death observed in human T-cell lymphoma cells established as a nude mouse tumor line

    SciTech Connect

    Igarashi, T.; Yoshida, S.; Miyamoto, T. )

    1990-08-01

    Interphase death of cells occurs physiologically in healthy animal tissues as well as in tissues pathologically injured by radiation or drugs. An active self-destruction process has been found to play a major role in the interphase death of highly radiosensitive cells. However, the mechanism of this radiation-induced interphase death in human lymphoma has not yet been studied in detail. In the present study, we examined a lymphoma derived from a child lymphoblastic lymphoma bearing CD1, CD4, and CD8 antigens and established in nude mice. Low-dose x-irradiation of this lymphoma induced interphase cell death with characteristic morphological and biological changes of an active self-destruction process, i.e., changes in cell surface appearance seen using scanning electron microscopy and nuclear fragmentation accompanied with an increase in free DNA. The process was proved to require protein synthesis. It was concluded that the radiosensitivity of this T-cell lymphoma of common thymic type is mainly due to the occurrence of the active self-destruction process.

  19. The Protective Roles of ROS-Mediated Mitophagy on 125I Seeds Radiation Induced Cell Death in HCT116 Cells

    PubMed Central

    Hu, Lelin; Wang, Hao; Huang, Li; Zhao, Yong

    2016-01-01

    For many unresectable carcinomas and locally recurrent cancers (LRC), 125I seeds brachytherapy is a feasible, effective, and safe treatment. Several studies have shown that 125I seeds radiation exerts anticancer activity by triggering DNA damage. However, recent evidence shows mitochondrial quality to be another crucial determinant of cell fate, with mitophagy playing a central role in this control mechanism. Herein, we found that 125I seeds irradiation injured mitochondria, leading to significantly elevated mitochondrial and intracellular ROS (reactive oxygen species) levels in HCT116 cells. The accumulation of mitochondrial ROS increased the expression of HIF-1α and its target genes BINP3 and NIX (BINP3L), which subsequently triggered mitophagy. Importantly, 125I seeds radiation induced mitophagy promoted cells survival and protected HCT116 cells from apoptosis. These results collectively indicated that 125I seeds radiation triggered mitophagy by upregulating the level of ROS to promote cellular homeostasis and survival. The present study uncovered the critical role of mitophagy in modulating the sensitivity of tumor cells to radiation therapy and suggested that chemotherapy targeting on mitophagy might improve the efficiency of 125I seeds radiation treatment, which might be of clinical significance in tumor therapy. PMID:28119765

  20. The Protective Roles of ROS-Mediated Mitophagy on (125)I Seeds Radiation Induced Cell Death in HCT116 Cells.

    PubMed

    Hu, Lelin; Wang, Hao; Huang, Li; Zhao, Yong; Wang, Junjie

    2016-01-01

    For many unresectable carcinomas and locally recurrent cancers (LRC), (125)I seeds brachytherapy is a feasible, effective, and safe treatment. Several studies have shown that (125)I seeds radiation exerts anticancer activity by triggering DNA damage. However, recent evidence shows mitochondrial quality to be another crucial determinant of cell fate, with mitophagy playing a central role in this control mechanism. Herein, we found that (125)I seeds irradiation injured mitochondria, leading to significantly elevated mitochondrial and intracellular ROS (reactive oxygen species) levels in HCT116 cells. The accumulation of mitochondrial ROS increased the expression of HIF-1α and its target genes BINP3 and NIX (BINP3L), which subsequently triggered mitophagy. Importantly, (125)I seeds radiation induced mitophagy promoted cells survival and protected HCT116 cells from apoptosis. These results collectively indicated that (125)I seeds radiation triggered mitophagy by upregulating the level of ROS to promote cellular homeostasis and survival. The present study uncovered the critical role of mitophagy in modulating the sensitivity of tumor cells to radiation therapy and suggested that chemotherapy targeting on mitophagy might improve the efficiency of (125)I seeds radiation treatment, which might be of clinical significance in tumor therapy.

  1. Modelling radiation-induced cell death and tumour re-oxygenation: local versus global and instant versus delayed cell death

    NASA Astrophysics Data System (ADS)

    Gago-Arias, Araceli; Aguiar, Pablo; Espinoza, Ignacio; Sánchez-Nieto, Beatriz; Pardo-Montero, Juan

    2016-02-01

    The resistance of hypoxic cells to radiation, due to the oxygen dependence of radiosensitivity, is well known and must be taken into account to accurately calculate the radiation induced cell death. A proper modelling of the response of tumours to radiation requires deriving the distribution of oxygen at a microscopic scale. This usually involves solving the reaction-diffusion equation in tumour voxels using a vascularization distribution model. Moreover, re-oxygenation arises during the course of radiotherapy, one reason being the increase of available oxygen caused by cell killing, which can turn hypoxic tumours into oxic. In this work we study the effect of cell death kinetics in tumour oxygenation modelling, analysing how it affects the timing of re-oxygenation, surviving fraction and tumour control. Two models of cell death are compared, an instantaneous cell killing, mimicking early apoptosis, and a delayed cell death scenario in which cells can die shortly after being damaged, as well as long after irradiation. For each of these scenarios, the decrease in oxygen consumption due to cell death can be computed globally (macroscopic voxel average) or locally (microscopic). A re-oxygenation model already used in the literature, the so called full re-oxygenation, is also considered. The impact of cell death kinetics and re-oxygenation on tumour responses is illustrated for two radiotherapy fractionation schemes: a conventional schedule, and a hypofractionated treatment. The results show large differences in the doses needed to achieve 50% tumour control for the investigated cell death models. Moreover, the models affect the tumour responses differently depending on the treatment schedule. This corroborates the complex nature of re-oxygenation, showing the need to take into account the kinetics of cell death in radiation response models.

  2. Modelling radiation-induced cell death and tumour re-oxygenation: local versus global and instant versus delayed cell death.

    PubMed

    Gago-Arias, Araceli; Aguiar, Pablo; Espinoza, Ignacio; Sánchez-Nieto, Beatriz; Pardo-Montero, Juan

    2016-02-07

    The resistance of hypoxic cells to radiation, due to the oxygen dependence of radiosensitivity, is well known and must be taken into account to accurately calculate the radiation induced cell death. A proper modelling of the response of tumours to radiation requires deriving the distribution of oxygen at a microscopic scale. This usually involves solving the reaction-diffusion equation in tumour voxels using a vascularization distribution model. Moreover, re-oxygenation arises during the course of radiotherapy, one reason being the increase of available oxygen caused by cell killing, which can turn hypoxic tumours into oxic. In this work we study the effect of cell death kinetics in tumour oxygenation modelling, analysing how it affects the timing of re-oxygenation, surviving fraction and tumour control. Two models of cell death are compared, an instantaneous cell killing, mimicking early apoptosis, and a delayed cell death scenario in which cells can die shortly after being damaged, as well as long after irradiation. For each of these scenarios, the decrease in oxygen consumption due to cell death can be computed globally (macroscopic voxel average) or locally (microscopic). A re-oxygenation model already used in the literature, the so called full re-oxygenation, is also considered. The impact of cell death kinetics and re-oxygenation on tumour responses is illustrated for two radiotherapy fractionation schemes: a conventional schedule, and a hypofractionated treatment. The results show large differences in the doses needed to achieve 50% tumour control for the investigated cell death models. Moreover, the models affect the tumour responses differently depending on the treatment schedule. This corroborates the complex nature of re-oxygenation, showing the need to take into account the kinetics of cell death in radiation response models.

  3. M-BAND Study of Radiation-Induced Chromosome Aberrations in Human Epithelial Cells: Radiation Quality and Dose Rate Effects

    NASA Technical Reports Server (NTRS)

    Hada, Megumi; Cucinotta, Francis; Wu, Honglu

    2009-01-01

    The advantage of the multicolor banding in situ hybridization (mBAND) technique is its ability to identify both inter- (translocation to unpainted chromosomes) and intra- (inversions and deletions within a single painted chromosome) chromosome aberrations simultaneously. To study the detailed rearrangement of low- and high-LET radiation induced chromosome aberrations in human epithelial cells (CH184B5F5/M10) in vitro, we performed a series of experiments with Cs-137 gamma rays of both low and high dose rates, neutrons of low dose rate and 600 MeV/u Fe ions of high dose rate, with chromosome 3 painted with multi-binding colors. We also compared the chromosome aberrations in both 2- and 3-dimensional cell cultures. Results of these experiments revealed the highest chromosome aberration frequencies after low dose rate neutron exposures. However, detailed analysis of the radiation induced inversions revealed that all three radiation types induced a low incidence of simple inversions. Most of the inversions in gamma-ray irradiated samples were accompanied by other types of intra-chromosomal aberrations but few inversions were accompanied by inter-chromosomal aberrations. In contrast, neutrons and Fe ions induced a significant fraction of inversions that involved complex rearrangements of both inter- and intrachromosomal exchanges. The location of the breaks involved in chromosome exchanges was analyzed along the painted chromosome. The breakpoint distribution was found to be randomly localized on chromosome 3 after neutron or Fe ion exposure, whereas non-random distribution with clustering breakpoints was observed after -ray exposure. Our comparison of chromosome aberration yields between 2- and 3-dimensional cell cultures indicated a significant difference for gamma exposures, but not for Fe ion exposures. These experimental results indicated that the track structure of the radiation and the cellular/chromosome structure can both affect radiation-induced chromosome

  4. Radiation-Induced c-Jun Activation Depends on MEK1-ERK1/2 Signaling Pathway in Microglial Cells

    PubMed Central

    Deng, Zhiyong; Sui, Guangchao; Rosa, Paulo Mottin; Zhao, Weiling

    2012-01-01

    Radiation-induced normal brain injury is associated with acute and/or chronic inflammatory responses, and has been a major concern in radiotherapy. Recent studies suggest that microglial activation is a potential contributor to chronic inflammatory responses following irradiation; however, the molecular mechanism underlying the response of microglia to radiation is poorly understood. c-Jun, a component of AP-1 transcription factors, potentially regulates neural cell death and neuroinflammation. We observed a rapid increase in phosphorylation of N-terminal c-Jun (on serine 63 and 73) and MAPK kinases ERK1/2, but not JNKs, in irradiated murine microglial BV2 cells. Radiation-induced c-Jun phosphorylation is dependent on the canonical MEK-ERK signaling pathway and required for both ERK1 and ERK2 function. ERK1/2 directly interact with c-Jun in vitro and in cells; meanwhile, the JNK binding domain on c-Jun is not required for its interaction with ERK kinases. Radiation-induced reactive oxygen species (ROS) potentially contribute to c-Jun phosphorylation through activating the ERK pathway. Radiation stimulates c-Jun transcriptional activity and upregulates c-Jun-regulated proinflammatory genes, such as tumor necrosis factor-α, interleukin-1β, and cyclooxygenase-2. Pharmacologic blockade of the ERK signaling pathway interferes with c-Jun activity and inhibits radiation-stimulated expression of c-Jun target genes. Overall, our study reveals that the MEK-ERK1/2 signaling pathway, but not the JNK pathway, contributes to the c-Jun-dependent microglial inflammatory response following irradiation. PMID:22606284

  5. Radiation-induced c-Jun activation depends on MEK1-ERK1/2 signaling pathway in microglial cells.

    PubMed

    Deng, Zhiyong; Sui, Guangchao; Rosa, Paulo Mottin; Zhao, Weiling

    2012-01-01

    Radiation-induced normal brain injury is associated with acute and/or chronic inflammatory responses, and has been a major concern in radiotherapy. Recent studies suggest that microglial activation is a potential contributor to chronic inflammatory responses following irradiation; however, the molecular mechanism underlying the response of microglia to radiation is poorly understood. c-Jun, a component of AP-1 transcription factors, potentially regulates neural cell death and neuroinflammation. We observed a rapid increase in phosphorylation of N-terminal c-Jun (on serine 63 and 73) and MAPK kinases ERK1/2, but not JNKs, in irradiated murine microglial BV2 cells. Radiation-induced c-Jun phosphorylation is dependent on the canonical MEK-ERK signaling pathway and required for both ERK1 and ERK2 function. ERK1/2 directly interact with c-Jun in vitro and in cells; meanwhile, the JNK binding domain on c-Jun is not required for its interaction with ERK kinases. Radiation-induced reactive oxygen species (ROS) potentially contribute to c-Jun phosphorylation through activating the ERK pathway. Radiation stimulates c-Jun transcriptional activity and upregulates c-Jun-regulated proinflammatory genes, such as tumor necrosis factor-α, interleukin-1β, and cyclooxygenase-2. Pharmacologic blockade of the ERK signaling pathway interferes with c-Jun activity and inhibits radiation-stimulated expression of c-Jun target genes. Overall, our study reveals that the MEK-ERK1/2 signaling pathway, but not the JNK pathway, contributes to the c-Jun-dependent microglial inflammatory response following irradiation.

  6. Protein Kinase CK2 Regulates Cytoskeletal Reorganization during Ionizing Radiation-Induced Senescence of Human Mesenchymal Stem Cells

    SciTech Connect

    Wang, Daojing; Jang, Deok-Jin

    2009-08-21

    Human mesenchymal stem cells (hMSC) are critical for tissue regeneration. How hMSC respond to genotoxic stresses and potentially contribute to aging and cancer remain underexplored. We demonstrated that ionizing radiation induced cellular senescence of hMSC over a period of 10 days, showing a critical transition between day 3 and day 6. This was confirmed by senescence-associated beta-galactosidase (SA-{beta}-gal) staining, protein expression profiles of key cell cycle regulators (retinoblastoma (Rb) protein, p53, p21{sup waf1/Cip1}, and p16{sup INK4A}), and senescence-associated secretory phenotypes (SASPs) (IL-8, IL-12, GRO, and MDC). We observed dramatic cytoskeletal reorganization of hMSC through reduction of myosin-10, redistribution of myosin-9, and secretion of profilin-1. Using a SILAC-based phosphoproteomics method, we detected significant reduction of myosin-9 phosphorylation at Ser1943, coinciding with its redistribution. Importantly, through treatment with cell permeable inhibitors (4,5,6,7-tetrabromo-1H-benzotriazole (TBB) and 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole (DMAT)), and gene knockdown using RNA interference, we identified CK2, a kinase responsible for myosin-9 phosphorylation at Ser1943, as a key factor contributing to the radiation-induced senescence of hMSC. We showed that individual knockdown of CK2 catalytic subunits CK2{alpha} and CK2{alpha}{prime} induced hMSC senescence. However, only knockdown of CK2{alpha} resulted in morphological phenotypes resembling those of radiation-induced senescence. These results suggest that CK2{alpha} and CK2{alpha}{prime} play differential roles in hMSC senescence progression, and their relative expression might represent a novel regulatory mechanism for CK2 activity.

  7. M-BAND Study of Radiation-Induced Chromosome Aberrations in Human Epithelial Cells: Radiation Quality and Dose Rate Effects

    NASA Technical Reports Server (NTRS)

    Hada, Megumi; Cucinotta, Francis; Wu, Honglu

    2009-01-01

    The advantage of the multicolor banding in situ hybridization (mBAND) technique is its ability to identify both inter- (translocation to unpainted chromosomes) and intra- (inversions and deletions within a single painted chromosome) chromosome aberrations simultaneously. To study the detailed rearrangement of low- and high-LET radiation induced chromosome aberrations in human epithelial cells (CH184B5F5/M10) in vitro, we performed a series of experiments with Cs-137 gamma rays of both low and high dose rates, neutrons of low dose rate and 600 MeV/u Fe ions of high dose rate, with chromosome 3 painted with multi-binding colors. We also compared the chromosome aberrations in both 2- and 3-dimensional cell cultures. Results of these experiments revealed the highest chromosome aberration frequencies after low dose rate neutron exposures. However, detailed analysis of the radiation induced inversions revealed that all three radiation types induced a low incidence of simple inversions. Most of the inversions in gamma-ray irradiated samples were accompanied by other types of intra-chromosomal aberrations but few inversions were accompanied by inter-chromosomal aberrations. In contrast, neutrons and Fe ions induced a significant fraction of inversions that involved complex rearrangements of both inter- and intrachromosomal exchanges. The location of the breaks involved in chromosome exchanges was analyzed along the painted chromosome. The breakpoint distribution was found to be randomly localized on chromosome 3 after neutron or Fe ion exposure, whereas non-random distribution with clustering breakpoints was observed after -ray exposure. Our comparison of chromosome aberration yields between 2- and 3-dimensional cell cultures indicated a significant difference for gamma exposures, but not for Fe ion exposures. These experimental results indicated that the track structure of the radiation and the cellular/chromosome structure can both affect radiation-induced chromosome

  8. Modulation of macrophage antitumor potential by apoptotic lymphoma cells.

    PubMed

    Voss, Jorine J L P; Ford, Catriona A; Petrova, Sofia; Melville, Lynsey; Paterson, Margaret; Pound, John D; Holland, Pam; Giotti, Bruno; Freeman, Tom C; Gregory, Christopher D

    2017-06-01

    In aggressive non-Hodgkin's lymphoma (NHL), constitutive apoptosis of a proportion of the tumor cell population can promote net tumor growth. This is associated with the accumulation of tumor-associated macrophages (TAMs) that clear apoptotic cells and exhibit pro-oncogenic transcriptional activation profiles characteristic of reparatory, anti-inflammatory and angiogenic programs. Here we consider further the activation status of these TAMs. We compare their transcriptomic profile with that of a range of other macrophage types from various tissues noting especially their expression of classically activated (IFN-γ and LPS) gene clusters - typically antitumor - in addition to their previously described protumor phenotype. To understand the impact of apoptotic cells on the macrophage activation state, we cocultured apoptotic lymphoma cells with classically activated macrophages (M(IFN-γ/LPS), also known as M1, macrophages). Although untreated and M(IFN-γ/LPS) macrophages were able to bind apoptotic lymphoma cells equally well, M(IFN-γ/LPS) macrophages displayed enhanced ability to phagocytose them. We found that direct exposure of M(IFN-γ/LPS) macrophages to apoptotic lymphoma cells caused switching towards a protumor activation state (often referred to as M2-like) with concomitant inhibition of antitumor activity that was a characteristic feature of M(IFN-γ/LPS) macrophages. Indeed, M(IFN-γ/LPS) macrophages exposed to apoptotic lymphoma cells displayed increased lymphoma growth-promoting activities. Antilymphoma activity by M(IFN-γ/LPS) macrophages was mediated, in part, by galectin-3, a pleiotropic glycoprotein involved in apoptotic cell clearance that is strongly expressed by lymphoma TAMs but not lymphoma cells. Intriguingly, aggressive lymphoma growth was markedly impaired in mice deficient in galectin-3, suggesting either that host galectin-3-mediated antilymphoma activity is required to sustain net tumor growth or that additional functions of galectin-3

  9. Modulation of macrophage antitumor potential by apoptotic lymphoma cells

    PubMed Central

    Voss, Jorine J L P; Ford, Catriona A; Petrova, Sofia; Melville, Lynsey; Paterson, Margaret; Pound, John D; Holland, Pam; Giotti, Bruno; Freeman, Tom C; Gregory, Christopher D

    2017-01-01

    In aggressive non-Hodgkin's lymphoma (NHL), constitutive apoptosis of a proportion of the tumor cell population can promote net tumor growth. This is associated with the accumulation of tumor-associated macrophages (TAMs) that clear apoptotic cells and exhibit pro-oncogenic transcriptional activation profiles characteristic of reparatory, anti-inflammatory and angiogenic programs. Here we consider further the activation status of these TAMs. We compare their transcriptomic profile with that of a range of other macrophage types from various tissues noting especially their expression of classically activated (IFN-γ and LPS) gene clusters – typically antitumor – in addition to their previously described protumor phenotype. To understand the impact of apoptotic cells on the macrophage activation state, we cocultured apoptotic lymphoma cells with classically activated macrophages (M(IFN-γ/LPS), also known as M1, macrophages). Although untreated and M(IFN-γ/LPS) macrophages were able to bind apoptotic lymphoma cells equally well, M(IFN-γ/LPS) macrophages displayed enhanced ability to phagocytose them. We found that direct exposure of M(IFN-γ/LPS) macrophages to apoptotic lymphoma cells caused switching towards a protumor activation state (often referred to as M2-like) with concomitant inhibition of antitumor activity that was a characteristic feature of M(IFN-γ/LPS) macrophages. Indeed, M(IFN-γ/LPS) macrophages exposed to apoptotic lymphoma cells displayed increased lymphoma growth-promoting activities. Antilymphoma activity by M(IFN-γ/LPS) macrophages was mediated, in part, by galectin-3, a pleiotropic glycoprotein involved in apoptotic cell clearance that is strongly expressed by lymphoma TAMs but not lymphoma cells. Intriguingly, aggressive lymphoma growth was markedly impaired in mice deficient in galectin-3, suggesting either that host galectin-3-mediated antilymphoma activity is required to sustain net tumor growth or that additional functions of

  10. Radiation-induced delayed cell death in a hypomorphic Artemis cell line.

    PubMed

    Evans, Paul M; Woodbine, Lisa; Riballo, Enriquetta; Gennery, Andrew R; Hubank, Michael; Jeggo, Penny A

    2006-04-15

    Null mutations in Artemis confer a condition described as RS-SCID, in which patients display radiosensitivity combined with severe combined immunodeficiency. Here, we characterize the defect in Artemis in a patient who displayed progressive combined immunodeficiency (CID) and elevated lymphocyte apoptosis. The patient is a compound heterozygote with novel mutations in both alleles, resulting in Artemis proteins with either L70 deletion or G126D substitution. Both mutational changes impact upon Artemis function and a fibroblast cell line derived from the patient (F96-224) has greatly reduced Artemis protein. In contrast to Artemis null cell lines, which fail to repair a subset of DNA double strand breaks (DSBs) induced by ionizing radiation, F96-224 cells show slow but residual DSB rejoining. Despite showing intermediate cellular and clinical features, F96-224 cells are as radiosensitive as Artemis null cell lines. We developed a FACS-based assay to examine cell division and cellular characteristics for 10 days following exposure to ionizing radiation (2 and 4 Gy). This analysis demonstrated that F96-224 cells show delayed cell death when compared with rapid growth arrest of an Artemis null cell line, and the emergence of a cycling population shown by a control line. F96-224 cells also display elevated chromosome aberrations when compared with control cells. F96-224 therefore represents a novel phenotype for a hypomorphic cell line. We suggest that delayed cell death contributes to the progressive CID phenotype of the Artemis patient.

  11. Porphyromonas gingivalis gingipains cause defective macrophage migration towards apoptotic cells and inhibit phagocytosis of primary apoptotic neutrophils.

    PubMed

    Castro, Sowmya A; Collighan, Russell; Lambert, Peter A; Dias, Irundika Hk; Chauhan, Parbata; Bland, Charlotte E; Milic, Ivana; Milward, Michael R; Cooper, Paul R; Devitt, Andrew

    2017-03-02

    Periodontal disease is a prevalent chronic inflammatory condition characterised by an aberrant host response to a pathogenic plaque biofilm resulting in local tissue damage and frustrated healing that can result in tooth loss. Cysteine proteases (gingipains) from the key periodontal pathogen Porphyromonas gingivalis have been implicated in periodontal disease pathogenesis by inhibiting inflammation resolution and are linked with systemic chronic inflammatory conditions such as rheumatoid arthritis. Efficient clearance of apoptotic cells is essential for the resolution of inflammation and tissue restoration. Here we sought to characterise the innate immune clearance of apoptotic cells and its modulation by gingipains. We examined the capacity of gingipain-treated macrophages to migrate towards and phagocytose apoptotic cells. Lysine gingipain treatment of macrophages impaired macrophage migration towards apoptotic neutrophils. Furthermore, lysine gingipain treatment reduced surface expression levels of CD14, a key macrophage receptor for apoptotic cells, which resulted in reduced macrophage interactions with apoptotic cells. Additionally, while apoptotic cells and their derived secretome were shown to inhibit TNF-α-induced expression by P. gingivalis lipopolysaccharide, we demonstrated that gingipain preparations induced a rapid inflammatory response in macrophages that was resistant to the anti-inflammatory effects of apoptotic cells or their secretome. Taken together, these data indicate that P. gingivalis may promote the chronic inflammation seen in periodontal disease patients by multiple mechanisms, including rapid, potent gingipain-mediated inflammation, coupled with receptor cleavage leading to defective clearance of apoptotic cells and reduced anti-inflammatory responses. Thus, gingipains represent a potential therapeutic target for intervention in the management of chronic periodontal disease.

  12. The BIANCA model/code of radiation-induced cell death: application to human cells exposed to different radiation types.

    PubMed

    Ballarini, Francesca; Altieri, Saverio; Bortolussi, Silva; Carante, Mario; Giroletti, Elio; Protti, Nicoletta

    2014-08-01

    This paper presents a biophysical model of radiation-induced cell death, implemented as a Monte Carlo code called BIophysical ANalysis of Cell death and chromosome Aberrations (BIANCA), based on the assumption that some chromosome aberrations (dicentrics, rings, and large deletions, called ‘‘lethal aberrations’’) lead to clonogenic inactivation. In turn, chromosome aberrations are assumed to derive from clustered, and thus severe, DNA lesions (called ‘‘cluster lesions,’’ or CL) interacting at the micrometer scale; the CL yield and the threshold distance governing CL interaction are the only model parameters. After a pilot study on V79 hamster cells exposed to protons and carbon ions, in the present work the model was extended and applied to AG1522 human cells exposed to photons, He ions, and heavier ions including carbon and neon. The agreement with experimental survival data taken from the literature supported the assumptions. In particular, the inactivation of AG1522 cells was explained by lethal aberrations not only for X-rays, as already reported by others, but also for the aforementioned radiation types. Furthermore, the results are consistent with the hypothesis that the critical initial lesions leading to cell death are DNA cluster lesions having yields in the order of *2 CL Gy-1 cell-1 at low LET and*20 CL Gy-1 cell-1 at high LET, and that the processing of these lesions is modulated by proximity effects at the micrometer scale related to interphase chromatin organization. The model was then applied to calculate the fraction of inactivated cells, as well as the yields of lethal aberrations and cluster lesions, as a function of LET; the results showed a maximum around 130 keV/lm, and such maximum was much higher for cluster lesions and lethal aberrations than for cell inactivation.

  13. Involvement of DNA-PK and ATM in radiation- and heat-induced DNA damage recognition and apoptotic cell death.

    PubMed

    Tomita, Masanori

    2010-01-01

    Exposure to ionizing radiation and hyperthermia results in important biological consequences, e.g. cell death, chromosomal aberrations, mutations, and DNA strand breaks. There is good evidence that the nucleus, specifically cellular DNA, is the principal target for radiation-induced cell lethality. DNA double-strand breaks (DSBs) are considered to be the most serious type of DNA damage induced by ionizing radiation. On the other hand, verifiable mechanisms which can lead to heat-induced cell death are damage to the plasma membrane and/or inactivation of heat-labile proteins caused by protein denaturation and subsequent aggregation. Recently, several reports have suggested that DSBs can be induced after hyperthermia because heat-induced phosphorylated histone H2AX (γ-H2AX) foci formation can be observed in several mammalian cell lines. In mammalian cells, DSBs are repaired primarily through two distinct and complementary mechanisms: non-homologous end joining (NHEJ), and homologous recombination (HR) or homology-directed repair (HDR). DNA-dependent protein kinase (DNA-PK) and ataxia-telangiectasia mutated (ATM) are key players in the initiation of DSB repair and phosphorylate and/or activate many substrates, including themselves. These phosphorylated substrates have important roles in the functioning of cell cycle checkpoints and in cell death, as well as in DSB repair. Apoptotic cell death is a crucial cell suicide mechanism during development and in the defense of homeostasis. If DSBs are unrepaired or misrepaired, apoptosis is a very important system which can protect an organism against carcinogenesis. This paper reviews recently obtained results and current topics concerning the role of DNA-PK and ATM in heat- or radiation-induced apoptotic cell death.

  14. Radiation-induced CXCL16 release by breast cancer cells attracts effector T cells1

    PubMed Central

    Matsumura, Satoko; Wang, Baomei; Kawashima, Noriko; Braunstein, Steve; Badura, Michelle; Cameron, Thomas O.; Babb, James S.; Schneider, Robert J.; Formenti, Silvia C.; Dustin, Michael L.; Demaria, Sandra

    2008-01-01

    Recruitment of effector T cells to inflamed peripheral tissues is regulated by chemokines and their receptors, but the factors regulating recruitment to tumors remain largely undefined. Ionizing radiation (IR) therapy is a common treatment modality for breast and other cancers. Used as a cytocidal agent for proliferating cancer cells, IR in combination with immunotherapy has been shown to promote immune-mediated tumor destruction in pre-clinical studies. Here we demonstrate that IR markedly enhanced the secretion by mouse and human breast cancer cells of CXCL16, a chemokine that binds to CXCR6 on Th1 and activated CD8 effector T cells, and plays an important role in their recruitment to sites of inflammation. Employing a poorly immunogenic mouse model of breast cancer, we found that irradiation increased the migration of CD8+CXCR6+ activated T cells to tumors in vitro and in vivo. CXCR6-deficient mice showed reduced infiltration of tumors by activated CD8 T cells and impaired tumor regression following treatment with local IR to the tumor and antibodies blocking the negative regulator of T cell activation CTLA-4. These results provide the first evidence that IR can induce the secretion by cancer cells of pro-inflammatory chemotactic factors that recruit anti-tumor effector T cells. The ability of IR to convert tumors into “inflamed” peripheral tissues could be exploited to overcome obstacles at the effector phase of the anti-tumor immune response and improve the therapeutic efficacy of immunotherapy. PMID:18713980

  15. Harnessing the apoptotic programs in cancer stem-like cells.

    PubMed

    Wang, Ying-Hua; Scadden, David T

    2015-09-01

    Elimination of malignant cells is an unmet challenge for most human cancer types even with therapies targeting specific driver mutations. Therefore, a multi-pronged strategy to alter cancer cell biology on multiple levels is increasingly recognized as essential for cancer cure. One such aspect of cancer cell biology is the relative apoptosis resistance of tumor-initiating cells. Here, we provide an overview of the mechanisms affecting the apoptotic process in tumor cells emphasizing the differences in the tumor-initiating or stem-like cells of cancer. Further, we summarize efforts to exploit these differences to design therapies targeting that important cancer cell population. © 2015 The Authors.

  16. Harnessing the apoptotic programs in cancer stem-like cells

    PubMed Central

    Wang, Ying-Hua; Scadden, David T

    2015-01-01

    Elimination of malignant cells is an unmet challenge for most human cancer types even with therapies targeting specific driver mutations. Therefore, a multi-pronged strategy to alter cancer cell biology on multiple levels is increasingly recognized as essential for cancer cure. One such aspect of cancer cell biology is the relative apoptosis resistance of tumor-initiating cells. Here, we provide an overview of the mechanisms affecting the apoptotic process in tumor cells emphasizing the differences in the tumor-initiating or stem-like cells of cancer. Further, we summarize efforts to exploit these differences to design therapies targeting that important cancer cell population. PMID:26253117

  17. Different mechanisms of radiation-induced loss of heterozygosity in two human lymphoid cell lines from a single donor

    NASA Technical Reports Server (NTRS)

    Wiese, C.; Gauny, S. S.; Liu, W. C.; Cherbonnel-Lasserre, C. L.; Kronenberg, A.

    2001-01-01

    Allelic loss is an important mutational mechanism in human carcinogenesis. Loss of heterozygosity (LOH) at an autosomal locus is one outcome of the repair of DNA double-strand breaks (DSBs) and can occur by deletion or by mitotic recombination. We report that mitotic recombination between homologous chromosomes occurred in human lymphoid cells exposed to densely ionizing radiation. We used cells derived from the same donor that express either normal TP53 (TK6 cells) or homozygous mutant TP53 (WTK1 cells) to assess the influence of TP53 on radiation-induced mutagenesis. Expression of mutant TP53 (Met 237 Ile) was associated with a small increase in mutation frequencies at the hemizygous HPRT (hypoxanthine phosphoribosyl transferase) locus, but the mutation spectra were unaffected at this locus. In contrast, WTK1 cells (mutant TP53) were 30-fold more susceptible than TK6 cells (wild-type TP53) to radiation-induced mutagenesis at the TK1 (thymidine kinase) locus. Gene dosage analysis combined with microsatellite marker analysis showed that the increase in TK1 mutagenesis in WTK1 cells could be attributed, in part, to mitotic recombination. The microsatellite marker analysis over a 64-cM region on chromosome 17q indicated that the recombinational events could initiate at different positions between the TK1 locus and the centromere. Virtually all of the recombinational LOH events extended beyond the TK1 locus to the most telomeric marker. In general, longer LOH tracts were observed in mutants from WTK1 cells than in mutants from TK6 cells. Taken together, the results demonstrate that the incidence of radi-ation-induced mutations is dependent on the genetic background of the cell at risk, on the locus examined, and on the mechanisms for mutation available at the locus of interest.

  18. Different mechanisms of radiation-induced loss of heterozygosity in two human lymphoid cell lines from a single donor

    NASA Technical Reports Server (NTRS)

    Wiese, C.; Gauny, S. S.; Liu, W. C.; Cherbonnel-Lasserre, C. L.; Kronenberg, A.

    2001-01-01

    Allelic loss is an important mutational mechanism in human carcinogenesis. Loss of heterozygosity (LOH) at an autosomal locus is one outcome of the repair of DNA double-strand breaks (DSBs) and can occur by deletion or by mitotic recombination. We report that mitotic recombination between homologous chromosomes occurred in human lymphoid cells exposed to densely ionizing radiation. We used cells derived from the same donor that express either normal TP53 (TK6 cells) or homozygous mutant TP53 (WTK1 cells) to assess the influence of TP53 on radiation-induced mutagenesis. Expression of mutant TP53 (Met 237 Ile) was associated with a small increase in mutation frequencies at the hemizygous HPRT (hypoxanthine phosphoribosyl transferase) locus, but the mutation spectra were unaffected at this locus. In contrast, WTK1 cells (mutant TP53) were 30-fold more susceptible than TK6 cells (wild-type TP53) to radiation-induced mutagenesis at the TK1 (thymidine kinase) locus. Gene dosage analysis combined with microsatellite marker analysis showed that the increase in TK1 mutagenesis in WTK1 cells could be attributed, in part, to mitotic recombination. The microsatellite marker analysis over a 64-cM region on chromosome 17q indicated that the recombinational events could initiate at different positions between the TK1 locus and the centromere. Virtually all of the recombinational LOH events extended beyond the TK1 locus to the most telomeric marker. In general, longer LOH tracts were observed in mutants from WTK1 cells than in mutants from TK6 cells. Taken together, the results demonstrate that the incidence of radi-ation-induced mutations is dependent on the genetic background of the cell at risk, on the locus examined, and on the mechanisms for mutation available at the locus of interest.

  19. Enhancement of radiation-induced cell kill by platinum complexes (carboplatin and iproplatin) in V79 cells

    SciTech Connect

    O'Hara, J.A.; Douple, E.B.; Richmond, R.C.

    1986-08-01

    Two second generation platinum complexes currently undergoing clinical chemotherapeutic trials, carboplatin (CBDCA) and iproplatin (CHIP), were evaluated for their ability to alter the survival of cultured Chinese hamster V79 cells following irradiation. Two protocols were employed. In the first, the drug was added to preplated cells, some of which were subsequently made hypoxic with nitrogen gas. These hypoxic cells were irradiated following 1 hour exposure to drug and survival was assessed by standard colony forming unit (CFU) methods. Enhancement ratios (ER) of approximately 1.4 were obtained for irradiation under hypoxic conditions, if the cells were exposed to equitoxic doses of CBDCA (500 microM) CHIP (50 microM). In the second series of experiments, cells were treated with 10 Gy in air and then incubated for various times prior to trypsinization and serial dilution of single cell suspensions. Six hours after irradiation, cells treated with X rays alone had recovered to produce a surviving fraction twice that of cells trypsinized immediately after irradiation (not held). Post-irradiation administration of CBDCA (50 microM) or CHIP (20 microM), at a time when free radical-mediated radiosensitization would not be possible, operationally inhibited this recovery from radiation-induced potentially lethal damage (PLD). Inhibition, expressed as recovery inhibition factor (RIF) after 6 hr with drug, was 2.0 for CBDCA and 1.2 for CHIP. These results suggest that the rationale for designing clinical trials to exploit interactions between cisplatin and radiation might also extend to include combined modality therapy using radiation with either of these two platinum complexes.

  20. Cycloheximide suppresses radiation-induced apoptosis in MOLT-4 cells with Arg72 variant of p53 through translational inhibition of p53 accumulation.

    PubMed

    Ito, Azusa; Morita, Akinori; Ohya, Soichiro; Yamamoto, Shinichi; Enomoto, Atsushi; Ikekita, Masahiko

    2011-01-01

    The human T-cell leukemia cell line MOLT-4 is highly radiosensitive, and thus it is often used as a model of p53-dependent radiation-induced apoptosis. Two branches of the p53-mediated apoptotic pathway are reported: "transcription-dependent" and "transcription-independent." However, the relative contribution of each in different types of cells is not yet clearly defined. Moreover, recent studies have shown that the codon 72 polymorphic variants of p53 show different sensitivities to apoptosis signals. The Arg72 variant has a more potent apoptosis-inducing activity in mitochondria than the Pro72 variant. Here, we initially investigated the codon 72 polymorphism of p53 in MOLT-4 cells. Analysis of the p53 exon 4 genomic DNA sequence, which includes codon 72, revealed that MOLT-4 cells are homozygous for the allele encoding Arg72. We next investigated the involvement of the transcription-independent function of p53 using an RNA synthesis inhibitor, actinomycin D (ActD), and a protein synthesis inhibitor, cycloheximide (CHX), and found that the apoptosis was suppressed by CHX but not by ActD. We also revealed that the suppressive effect of CHX on apoptosis was specifically mediated by p53, using a p53-knockdown MOLT-4 transfectant. Furthermore, the suppressive effect of CHX on apoptosis was highly correlated with the suppression of p53 protein accumulation, and less correlated with the suppression of p53 target genes expression. These results indicated that p53 transactivation is not necessary to induce apoptosis, and that p53 protein accumulation itself is both necessary and sufficient to do so.

  1. Necdin modulates proliferative cell survival of human cells in response to radiation-induced genotoxic stress

    PubMed Central

    2012-01-01

    Background The finite replicative lifespan of cells, termed cellular senescence, has been proposed as a protective mechanism against the proliferation of oncogenically damaged cells, that fuel cancer. This concept is further supported by the induction of premature senescence, a process which is activated when an oncogene is expressed in normal primary cells as well as following intense genotoxic stresses. Thus, deregulation of genes that control this process, like the tumor suppressor p53, may contribute to promoting cancer by allowing cells to bypass senescence. A better understanding of the genes that contribute to the establishment of senescence is therefore warranted. Necdin interacts with p53 and is also a p53 target gene, although the importance of Necdin in the p53 response is not clearly understood. Methods In this study, we first investigated Necdin protein expression during replicative senescence and premature senescence induced by gamma irradiation and by the overexpression of oncogenic RasV12. Gain and loss of function experiments were used to evaluate the contribution of Necdin during the senescence process. Results Necdin expression declined during replicative aging of IMR90 primary human fibroblasts or following induction of premature senescence. Decrease in Necdin expression seemed to be a consequence of the establishment of senescence since the depletion of Necdin in human cells did not induce a senescence-like growth arrest nor a flat morphology or SA-β-galactosidase activity normally associated with senescence. Similarly, overexpression of Necdin did not affect the life span of IMR90 cells. However, we demonstrate that in normal human cells, Necdin expression mimicked the effect of p53 inactivation by increasing radioresistance. Conclusion This result suggests that Necdin potentially attenuate p53 signaling in response to genotoxic stress in human cells and supports similar results describing an inhibitory function of Necdin over p53-dependent

  2. Middle infrared radiation induces G2/M cell cycle arrest in A549 lung cancer cells.

    PubMed

    Chang, Hsin-Yi; Shih, Meng-Her; Huang, Hsuan-Cheng; Tsai, Shang-Ru; Juan, Hsueh-Fen; Lee, Si-Chen

    2013-01-01

    There were studies investigating the effects of broadband infrared radiation (IR) on cancer cell, while the influences of middle-infrared radiation (MIR) are still unknown. In this study, a MIR emitter with emission wavelength band in the 3-5 µm region was developed to irradiate A549 lung adenocarcinoma cells. It was found that MIR exposure inhibited cell proliferation and induced morphological changes by altering the cellular distribution of cytoskeletal components. Using quantitative PCR, we found that MIR promoted the expression levels of ATM (ataxia telangiectasia mutated), ATR (ataxia-telangiectasia and Rad3-related and Rad3-related), TP53 (tumor protein p53), p21 (CDKN1A, cyclin-dependent kinase inhibitor 1A) and GADD45 (growth arrest and DNA-damage inducible), but decreased the expression levels of cyclin B coding genes, CCNB1 and CCNB2, as well as CDK1 (Cyclin-dependent kinase 1). The reduction of protein expression levels of CDC25C, cyclin B1 and the phosphorylation of CDK1 at Thr-161 altogether suggest G(2)/M arrest occurred in A549 cells by MIR. DNA repair foci formation of DNA double-strand breaks (DSB) marker γ-H2AX and sensor 53BP1 was induced by MIR treatment, it implies the MIR induced G(2)/M cell cycle arrest resulted from DSB. This study illustrates a potential role for the use of MIR in lung cancer therapy by initiating DSB and blocking cell cycle progression.

  3. Middle Infrared Radiation Induces G2/M Cell Cycle Arrest in A549 Lung Cancer Cells

    PubMed Central

    Huang, Hsuan-Cheng; Tsai, Shang-Ru; Juan, Hsueh-Fen; Lee, Si-Chen

    2013-01-01

    There were studies investigating the effects of broadband infrared radiation (IR) on cancer cell, while the influences of middle-infrared radiation (MIR) are still unknown. In this study, a MIR emitter with emission wavelength band in the 3–5 µm region was developed to irradiate A549 lung adenocarcinoma cells. It was found that MIR exposure inhibited cell proliferation and induced morphological changes by altering the cellular distribution of cytoskeletal components. Using quantitative PCR, we found that MIR promoted the expression levels of ATM (ataxia telangiectasia mutated), ATR (ataxia-telangiectasia and Rad3-related and Rad3-related), TP53 (tumor protein p53), p21 (CDKN1A, cyclin-dependent kinase inhibitor 1A) and GADD45 (growth arrest and DNA-damage inducible), but decreased the expression levels of cyclin B coding genes, CCNB1 and CCNB2, as well as CDK1 (Cyclin-dependent kinase 1). The reduction of protein expression levels of CDC25C, cyclin B1 and the phosphorylation of CDK1 at Thr-161 altogether suggest G2/M arrest occurred in A549 cells by MIR. DNA repair foci formation of DNA double-strand breaks (DSB) marker γ-H2AX and sensor 53BP1 was induced by MIR treatment, it implies the MIR induced G2/M cell cycle arrest resulted from DSB. This study illustrates a potential role for the use of MIR in lung cancer therapy by initiating DSB and blocking cell cycle progression. PMID:23335992

  4. Autophagy Promotes the Repair of Radiation-Induced DNA Damage in Bone Marrow Hematopoietic Cells via Enhanced STAT3 Signaling.

    PubMed

    Xu, Fei; Li, Xin; Yan, Lili; Yuan, Na; Fang, Yixuan; Cao, Yan; Xu, Li; Zhang, Xiaoying; Xu, Lan; Ge, Chaorong; An, Ni; Jiang, Gaoyue; Xie, Jialing; Zhang, Han; Jiang, Jiayi; Li, Xiaotian; Yao, Lei; Zhang, Suping; Zhou, Daohong; Wang, Jianrong

    2017-03-01

    Autophagy protects hematopoietic cells from radiation damage in part by promoting DNA damage repair. However, the molecular mechanisms by which autophagy regulates DNA damage repair remain largely elusive. Here, we report that this radioprotective effect of autophagy depends on STAT3 signaling in murine bone marrow mononuclear cells (BM-MNCs). Specifically, we found that STAT3 activation and nuclear translocation in BM-MNCs were increased by activation of autophagy with an mTOR inhibitor and decreased by knockout of the autophagy gene Atg7. The autophagic regulation of STAT3 activation is likely mediated by induction of KAP1 degradation, because we showed that KAP1 directly interacted with STAT3 in the cytoplasm and knockdown of KAP1 increased the phosphorylation and nuclear translocation of STAT3. Subsequently, activated STAT3 transcriptionally upregulated the expression of BRCA1, which increased the ability of BM-MNCs to repair radiation-induced DNA damage. This novel finding that activation of autophagy can promote DNA damage repair in BM-MNCs via the ATG-KAP1-STAT3-BRCA1 pathway suggests that autophagy plays an important role in maintaining genomic integrity of BM-MNCs and its activation may confer protection of BM-MNCs against radiation-induced genotoxic stress.

  5. Progressive behavioral changes during the maturation of rats with early radiation-induced hypoplasia of fascia dentata granule cells

    SciTech Connect

    Mickley, G.A.; Ferguson, J.L.; Mulvihill, M.A.; Nemeth, T.J.

    1989-01-01

    Localized exposure of the neonatal rat brain to x rays produces neuronal hypoplasia specific to the granule cell layer of the hippocampal dentate gyrus. This brain damage causes locomotor hyperactivity, slowed acquisition of passive avoidance tasks and long bouts of spontaneous turning (without reversals) in a bowl apparatus. The authors report here how these behavioral deficits change as a function of subject aging and behavioral test replications. Portions of the neonatal rat cerebral hemispheres were X-irradiated in order to selectively damage the granule cells of the dentate gyrus. Rats between the ages of 71-462 days were tested 3 separate times on each of the following 3 behavioral tests: (1) spontaneous locomotion, (2) passive avoidance acquisition, and (3) spontaneous circling in a large plastic hemisphere. Rats with radiation-induced damage to the fascia dentata exhibited long bouts of slow turns without reversals. Once they began, irradiated subjects perseverated in turning to an extent significantly greater than sham-irradiated control subjects. The hyperactivity of the irradiated animals decreased significantly as they matured. These data suggest that radiation-induced damage to the fascia dentata produces task-dependent behavioral deficits that change as a function of subject age and/or behavioral testing.

  6. Evidence for apoptotic cell death in Alzheimer's disease.

    PubMed

    Smale, G; Nichols, N R; Brady, D R; Finch, C E; Horton, W E

    1995-06-01

    We provide evidence for apoptosis in Alzheimer's disease using the in situ labeling technique TUNEL (terminal transferase-mediated dUTP-biotin nick end labeling). The technique specifically detects apoptotic cells by utilizing terminal transferase to incorporate biotinylated nucleotides into the fragmented DNA of apoptotic cells. The labeled cells are visualized by reaction with avidin peroxidase and a suitable substrate. Sections from the hippocampus of Alzheimer-diseased (AD) brains and non-AD brains were examined for apoptosis. While considerable variation in the quantity of apoptotic cells was observed among individual samples, the incidence of apoptosis in AD brains was elevated in comparison to age-matched, non-AD brains in specific regions of the hippocampal formation. Immunostaining indicated that both neurons and astrocytes were undergoing apoptosis, although the majority of the TUNEL-positive cells appeared to be glial, based on the location of the stained cells. These data suggest that apoptosis may be involved in both the primary neuronal cell loss and in the glial response that is a component of AD.

  7. PDT-apoptotic tumor cells induce macrophage immune response

    NASA Astrophysics Data System (ADS)

    Zhou, Fei-fan; Xing, Da; Chen, Wei R.

    2008-02-01

    Photodynamic therapy (PDT) functions as a cancer therapy through two major cell death mechanisms: apoptosis and necrosis. Immunological responses induced by PDT has been mainly associated with necrosis while apoptosis associated immune responses have not fully investigated. Heat shock proteins (HSPs) play an important role in regulating immune responses. In present study, we studied whether apoptotic tumor cells could induce immune response and how the HSP70 regulates immune response. The endocytosis of tumor cells by the activated macrophages was observed at single cell level by LSM. The TNF-α release of macrophages induced by co-incubated with PDT-apoptotic tumor cells was detected by ELISA. We found that apoptotic tumor cells treated by PDT could activate the macrophages, and the immune effect decreased evidently when HSP70 was blocked. These findings not only show that apoptosis can induce immunological responses, but also show HSP70 may serves as a danger signal for immune cells and induce immune responses to regulate the efficacy of PDT.

  8. Apoptotic cell signaling in cancer progression and therapy†

    PubMed Central

    Plati, Jessica; Bucur, Octavian; Khosravi-Far, Roya

    2011-01-01

    Apoptosis is a tightly regulated cell suicide program that plays an essential role in the development and maintenance of tissue homeostasis by eliminating unnecessary or harmful cells. Impairment of this native defense mechanism promotes aberrant cellular proliferation and the accumulation of genetic defects, ultimately resulting in tumorigenesis, and frequently confers drug resistance to cancer cells. The regulation of apoptosis at several levels is essential to maintain the delicate balance between cellular survival and death signaling that is required to prevent disease. Complex networks of signaling pathways act to promote or inhibit apoptosis in response to various cues. Apoptosis can be triggered by signals from within the cell, such as genotoxic stress, or by extrinsic signals, such as the binding of ligands to cell surface death receptors. Various upstream signaling pathways can modulate apoptosis by converging on, and thereby altering the activity of, common central control points within the apoptotic signaling pathways, which involve the BCL-2 family proteins, inhibitor of apoptosis (IAP) proteins, and FLICE-inhibitory protein (c-FLIP). This review highlights the role of these fundamental regulators of apoptosis in the context of both normal apoptotic signaling mechanisms and dysregulated apoptotic pathways that can render cancer cells resistant to cell death. In addition, therapeutic strategies aimed at modulating the activity of BCL-2 family proteins, IAPs, and c-FLIP for the targeted induction of apoptosis are briefly discussed. PMID:21340093

  9. Apoptotic cell signaling in cancer progression and therapy.

    PubMed

    Plati, Jessica; Bucur, Octavian; Khosravi-Far, Roya

    2011-04-01

    Apoptosis is a tightly regulated cell suicide program that plays an essential role in the development and maintenance of tissue homeostasis by eliminating unnecessary or harmful cells. Impairment of this native defense mechanism promotes aberrant cellular proliferation and the accumulation of genetic defects, ultimately resulting in tumorigenesis, and frequently confers drug resistance to cancer cells. The regulation of apoptosis at several levels is essential to maintain the delicate balance between cellular survival and death signaling that is required to prevent disease. Complex networks of signaling pathways act to promote or inhibit apoptosis in response to various cues. Apoptosis can be triggered by signals from within the cell, such as genotoxic stress, or by extrinsic signals, such as the binding of ligands to cell surface death receptors. Various upstream signaling pathways can modulate apoptosis by converging on, and thereby altering the activity of, common central control points within the apoptotic signaling pathways, which involve the BCL-2 family proteins, inhibitor of apoptosis (IAP) proteins, and FLICE-inhibitory protein (c-FLIP). This review highlights the role of these fundamental regulators of apoptosis in the context of both normal apoptotic signaling mechanisms and dysregulated apoptotic pathways that can render cancer cells resistant to cell death. In addition, therapeutic strategies aimed at modulating the activity of BCL-2 family proteins, IAPs, and c-FLIP for the targeted induction of apoptosis are briefly discussed.

  10. The effect of ultraviolet radiation-induced suppressor cells on T-cell activity.

    PubMed Central

    Ullrich, S E

    1987-01-01

    The suppression of contact hypersensitivity (CHS) after a single exposure to ultraviolet (UV) radiation provides an excellent model system with which to study both the activation and the mode of action of suppressor T cells. Suppression of CHS after UV radiation is mediated by hapten-specific suppressor T cells (UVTs). These cells have a broad range of activity: CHS and antibody production in vivo and the generation of cytolytic T lymphocytes (CTL) and T-cell proliferative responses in vitro are suppressed by UVTs. The present study is concerned with determining the target of UVTs. The UVTs could suppress the response to hapten-modified T-dependent antigens, such as trinitrophenyl (TNP)-modified sheep erythrocytes (TNP-SRBC) or TNP-conjugated bovine serum albumin (TNP-BSA), but had no suppressive effect on the response to a T-independent antigen, TNP-conjugated lipopolysaccharide (TNP-LPS). The UVTs also suppressed the generation of interleukin-2 (IL-2) in vitro. The suppression of CTL generation in vitro and CHS in vivo could be overcome by the addition of exogenous IL-2. These data suggest that UVTs suppress the immune response by affecting T-helper cell function. PMID:2952584

  11. The role of hypoxia-inducible factor-1α in radiation-induced autophagic cell death in breast cancer cells.

    PubMed

    Zhong, Rui; Xu, Huiying; Chen, Ge; Zhao, Gang; Gao, Yan; Liu, Xiaodong; Ma, Shumei; Dong, Lihua

    2015-09-01

    Hypoxia-inducible factor-1α (HIF-1α) is a major effector in cell survival response to hypoxia, while the roles of HIF-1α in radiation-induced autophagy are still unclear in breast cancer cells. Human breast cancer carcinoma MCF-7 cells were stably transfected with pSUPER-shRNA against human HIF-1α or a scrambled sequence with no homology to mammalian genes, named as pSUPER-HIF-1α and pSUPER-SC, respectively. Cell Counting Kit-8 (CCK-8) assay and colony formation assay were used to detect cell viability, Western blot was used to detect protein expression, monodansylcadaverine (MDC) staining was used to analyze autophagy, and Hoechts/PI staining was used to assess apoptosis. Ionizing radiation (IR) and cobalt chloride (CoCl2) could induce HIF-1α expression and increase the microtubule-associated protein 1 light chain 3 (MAPLC3)-II/MAPLC3-I ratio, especially in radiation + CoCl2 group. After the silencing of HIF-1α, the radiosensitivity of MCF-7 cells increased and the autophagy level decreased in response to DNA damage induced by ionizing radiation, but there was no influence on IR-induced apoptosis. HIF-1α silencing also increased the expression of phospho-Akt, mTOR, and P70S6K and activated the mTOR signals significantly. Hypoxia can induce autophagy and also improve the IR-induced autophagy via the suppression of Akt/mTOR/P70S6K pathway, which consequently lead to radioresistance.

  12. Lipophilic triphenylphosphonium derivatives enhance radiation-induced cell killing via inhibition of mitochondrial energy metabolism in tumor cells.

    PubMed

    Yasui, Hironobu; Yamamoto, Kumiko; Suzuki, Motofumi; Sakai, Yuri; Bo, Tomoki; Nagane, Masaki; Nishimura, Eri; Yamamori, Tohru; Yamasaki, Toshihide; Yamada, Ken-Ichi; Inanami, Osamu

    2017-04-01

    It has recently been reported that radiation enhances mitochondrial energy metabolism in various tumor cell lines. To examine how this radiation-induced alteration in mitochondrial function influences tumor cell viability, various lipophilic triphenylphosphonium (TPP(+)) cation derivatives and related compounds such as 4-hydroxy-2,2,6,6-tetramethyl-1-oxy-piperidin (Tempol) with TPP(+) (named "Mito-") were designed to inhibit the mitochondrial electron transport chain. Mito-(CH2)10-Tempol (M10T) and its derivatives, Mito-(CH2)5-Tempol (M5T), Mito-(CH2)10-Tempol-Methyl (M10T-Me), Mito-C10H21 (M10), and C10H21-Tempol (10T), were prepared. In HeLa human cervical adenocarcinoma cells and A549 human lung carcinoma cells, the fractional uptake of the compound into mitochondria was highest among the TTP(+) analogs conjugated with Tempol (M10T, M5T, and 10T). M10T, M10T-Me, and M10 exhibited strong cytotoxicity and enhanced X-irradiation-induced reproductive cell death, while 10T and M5T did not. Furthermore, M10T, M10T-Me, and M10 decreased basal mitochondrial membrane potential and intracellular ATP. M10T treatment inhibited X-ray-induced increases in ATP production. These results indicate that the TPP cation and a long hydrocarbon linker are essential for radiosensitization of tumor cells. The reduction in intracellular ATP by lipophilic TPP(+) is partly responsible for the observed radiosensitization.

  13. Mechanisms of Ionizing Radiation-Induced Cell Death in Primary Lung Cells

    DTIC Science & Technology

    2013-03-05

    Zhai M, et al. 2007. Protein oxidation implicated as the primary determinant of bacterial radioresistance. PLoS Biol 5:e92 43. Demidenko ZN...oxidative stress induces senescence in retinal pigment epithelial cells via TGF-beta release. Investigative ophthalmology & visual science 50:926- 35

  14. Diallyl disulfide enhances carbon ion beams–induced apoptotic cell death in cervical cancer cells through regulating Tap73 /ΔNp73

    PubMed Central

    Di, Cuixia; Sun, Chao; Li, Hongyan; Si, Jing; Zhang, Hong; Han, Lu; Zhao, Qiuyue; Liu, Yang; Liu, Bin; Miao, Guoying; Gan, Lu; Liu, Yuanyuan

    2015-01-01

    Diallyl disulfide (DADS), extracted from crushed garlic by steam-distillation, has been reported to provide the anticancer activity in several cancer types. However, the effect of DADS on high-LET carbon beams - induced cell death remains unknown. Therefore, we used human cervical cancer cells to elucidate the molecular effects of this dallyl sulfide. Radiotherapy remains the mainstay of treatment, especially in advanced cervical cancer and there is still space to improve the radiosensitivity to reduce radiation dosage. In this study, we found that radiation effects evoked by high-LET carbon beam was marked by inhibition of cell viability, cell cycle arrest, significant rise of apoptotic cells, regulation of transcription factor, such as p73, as well as alterations of crucial mediator of the apoptosis pathway. We further demonstrated that pretreatment of 10 µM DADS in HeLa cells exposed to radiation resulted in decrease in cell viability and increased radiosensitivity. Additionally, cells pretreated with DADS obviously inhibited the radiation-induced G2/M phase arrest, but promoted radiation-induced apoptosis.  Moreover, combination DADS and the radiation exacerbated the activation of apoptosis pathways through up-regulated ration of pro-apoptotic Tap73 to anti-apoptotic ΔNp73, and its downstream proteins, such as FASLG, and APAF1. Taken together, these results suggest that DADS is a potential candidate as radio sensitive agent for cervical cancer. PMID:26505313

  15. Radiation-induced xerostomia: pathophysiology, clinical course and supportive treatment.

    PubMed

    Guchelaar, H J; Vermes, A; Meerwaldt, J H

    1997-07-01

    Xerostomia, or oral dryness, is one of the most common complaints experienced by patients who have had radiotherapy of the oral cavity and neck region. The hallmarks of radiation-induced damage are acinar atrophy and chronic inflammation of the salivary glands. The early response, resulting in atrophy of the secretory cells without inflammation might be due to radiation-induced apoptosis. In contrast, the late response with inflammation could be a result of radiation-induced necrosis. The subjective complaint of a dry mouth appears to be poorly correlated with objective findings of salivary gland dysfunction. Xerostomia, with secondary symptoms of increased dental caries, difficulty in chewing, swallowing and speaking, and an increased incidence of oral candidiasis, can have a significant effect on the quality of life. At present there is no causal treatment for radiation-induced xerostomia. Temporary symptomatic relief can be offered by moistening agents and saliva substitutes, and is the only option for patients without residual salivary function. In patients with residual salivary function, oral administration of pilocarpine 5-10 mg three times a day is effective in increasing salivary flow and improving the symptoms of xerostomia, and this therapy should be considered as the treatment of choice. Effectiveness of sialogogue treatment requires residual salivary function, which emphasizes the potential benefit from sparing normal tissue during irradiation. The hypothesis concerning the existence of early apoptotic and late necrotic effects of irradiation on the salivary glands theoretically offers a way of achieving this goal.

  16. Spatially Fractionated Radiation Induces Cytotoxicity and Changes in Gene Expression in Bystander and Radiation Adjacent Murine Carcinoma Cells

    PubMed Central

    Asur, Rajalakshmi S.; Sharma, Sunil; Chang, Ching-Wei; Penagaricano, Jose; Kommuru, Indira M.; Moros, Eduardo G.; Corry, Peter M.; Griffin, Robert J.

    2012-01-01

    Radiation-induced bystander effects have been extensively studied at low doses, since evidence of bystander induced cell killing and other effects on unirradiated cells were found to be predominant at doses up to 0.5 Gy. Therefore, few studies have examined bystander effects induced by exposure to higher doses of radiation, such as spatially fractionated radiation (GRID) treatment. In the present study, we evaluate the ability of GRID treatment to induce changes in GRID adjacent (bystander) regions, in two different murine carcinoma cell lines following exposure to a single irradiation dose of 10 Gy. Murine SCK mammary carcinoma cells and SCCVII squamous carcinoma cells were irradiated using a brass collimator to create a GRID pattern of nine circular fields 12 mm in diameter with a center-to-center distance of 18 mm. Similar to the typical clinical implementation of GRID, this is approximately a 50:50 ratio of direct and bystander exposure. We also performed experiments by irradiating separate cultures and transferring the medium to unirradiated bystander cultures. Clonogenic survival was evaluated in both cell lines to determine the occurrence of radiation-induced bystander effects. For the purpose of our study, we have defined bystander cells as GRID adjacent cells that received approximately 1 Gy scatter dose or unirradiated cells receiving conditioned medium from irradiated cells. We observed significant bystander killing of cells adjacent to the GRID irradiated regions compared to sham treated controls. We also observed bystander killing of SCK and SCCVII cells cultured in conditioned medium obtained from cells irradiated with 10 Gy. Therefore, our results confirm the occurrence of bystander effects following exposure to a high-dose of radiation and suggest that cell-to-cell contact is not required for these effects. In addition, the gene expression profile for DNA damage and cellular stress response signaling in SCCVII cells after GRID exposure was studied

  17. Sensitivity and dose dependency of radiation-induced injury in hematopoietic stem/progenitor cells in mice

    PubMed Central

    Guo, Chang-Ying; Luo, Lan; Urata, Yoshishige; Goto, Shinji; Huang, Wen-Jing; Takamura, Syu; Hayashi, Fumiko; Doi, Hanako; Kitajima, Yuriko; Ono, Yusuke; Ogi, Tomoo; Li, Tao-Sheng

    2015-01-01

    We evaluated the sensitivity and dose dependency of radiation-induced injury in hematopoietic stem/progenitor cells. Adult C57BL/6 mice were daily exposed to 0, 2, 10, 50, and 250 mGy γ-ray for 1 month in succession, respectively. The damage of hematopoietic stem/progenitor cells in bone marrow were investigated within 2 hours (acute phase) or at 3 months (chronic phase) after the last exposure. Daily exposure to over 10 mGy γ-ray significantly decreased the number and colony-forming capacity of hematopoietic stem/progenitor cells at acute phase, and did not completely recover at chronic phase with 250 mGy exposure. Interestingly, the daily exposure to 10 or 50 mGy γ-ray decreased the formation of mixed types of colonies at chronic phase, but the total number of colonies was comparable to control. Immunostaining analysis showed that the formation of 53BP1 foci in c-kit+ stem/progenitor cells was significantly increased with daily exposure to 50 and 250 mGy at acute phase, and 250 mGy at chronic phase. Many genes involved in toxicity responses were up- or down-regulated with the exposures to all doses. Our data have clearly shown the sensitivity and dose dependency of radiation-induced injury in hematopoietic stem/progenitor cells of mice with daily exposures to 2 ~ 250 mGy γ-ray. PMID:25623887

  18. Dose and time dependent apoptotic response in a human melanoma cell line exposed to accelerated boron ions at four different LET.

    PubMed

    Meijer, A E; Jernberg, A R-M; Heiden, T; Stenerlöw, B; Persson, L M; Tilly, N; Lind, B K; Edgren, M R

    2005-04-01

    The aim was to investigate and compare the influence of linear energy transfer (LET), dose and time on the induction of apoptosis in a human melanoma cell line exposed to accelerated light boron ((10)B) ions and photons. Cells were exposed in vitro to doses up to 6 Gy accelerated boron ions (40, 80, 125 and 160 eV nm(-1)) and up to 12 Gy photons (0.2 eV nm(-1)). The induction of apoptosis was measured up to 9 days after irradiation using morphological characterization of apoptotic cells and bodies. In parallel, measurements of cell-cycle distribution, monitored by DNA flow cytometry, and cell survival based on the clonogenic cell survival assay, were performed. In addition, the induction and repair of DNA double-strand breaks (DSB), using pulsed-field gel electrophoresis (PFGE) were studied. Accelerated boron ions induced a significant increase in apoptosis as compared with photons at all time points studied. At 1-5 h the percentage of radiation-induced apoptotic cells increased with both dose and LET. At the later time points (24-216 h) the apoptotic response was more complex and did not increase in a strictly LET-dependent manner. The early premitotic apoptotic cells disappeared at 24 h following exposure to the highest LET (160 eV nm(-1)). A postmitotic apoptotic response was seen after release of the dose-, time- and LET-dependent G2/M accumulations. The loss of clonogenic ability was dose- and LET-dependent and the fraction of un-rejoined DSB increased with increasing LET. Despite the LET-dependent clonogenic cell killing, it was not possible to measure quantitatively a LET-dependent apoptotic response. This was due to the different time course of appearance and disappearance of apoptotic cells.

  19. Caspase-independent cell death mediated by apoptosis-inducing factor (AIF) nuclear translocation is involved in ionizing radiation induced HepG2 cell death.

    PubMed

    Sun, Hengwen; Yang, Shana; Li, Jianhua; Zhang, Yajie; Gao, Dongsheng; Zhao, Shenting

    2016-03-25

    Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. The aim of radiotherapy is to eradicate cancer cells with ionizing radiation. Except for the caspase-dependent mechanism, several lines of evidence demonstrated that caspase-independent mechanism is directly involved in the cell death responding to irradiation. For this reason, defining the contribution of caspase-independent molecular mechanisms represents the main goal in radiotherapy. In this study, we focused on the role of apoptosis-inducing factor (AIF), the caspase-independent molecular, in ionizing radiation induced hepatocellular carcinoma cell line (HepG2) cell death. We found that ionizing radiation has no function on AIF expression in HepG2 cells, but could induce AIF release from the mitochondria and translocate into nuclei. Inhibition of AIF could reduce ionizing radiation induced HepG2 cell death. These studies strongly support a direct relationship between AIF nuclear translocation and radiation induced cell death. What's more, AIF nuclear translocation is caspase-independent manner, but not caspase-dependent manner, in this process. These new findings add a further attractive point of investigation to better define the complex interplay between caspase-independent cell death and radiation therapy.

  20. Study of radiation induced deep-level defects in proton irradiated AlGaAs-GaAs solar cells

    NASA Technical Reports Server (NTRS)

    Li, S. S.

    1980-01-01

    Radiation induced deep-level defects (both electron and hole traps) in proton irradiated AlGaAs-GaAs p-n junction solar cells are investigated along with the correlation between the measured defect parameters and the solar cell performance parameters. The range of proton energies studied was from 50 KeV to 10 MeV and the proton fluence was varied from 10 to the 10th power to 10 to the 13th power P/sq cm. Experimental tools employed include deep-level transient spectroscopy, capacitance-voltage, current voltage, and SEM-EBIC methods. Defect and recombination parameters such as defect density and energy level, capture cross section, carrier lifetimes and effective hole diffusion lengths in n-GaAs LPE layers were determined from these measurements.

  1. Thymoquinone restores radiation-induced TGF-β expression and abrogates EMT in chemoradiotherapy of breast cancer cells.

    PubMed

    Rajput, Shashi; Kumar, B N Prashanth; Banik, Payel; Parida, Sheetal; Mandal, Mahitosh

    2015-03-01

    Radiotherapy remains a prime approach to adjuvant therapies in patients with early and advanced breast cancer. In spite of therapeutic success, metastatic progression in patients undergoing therapy, limits its application. However, effective therapeutic strategies to understand the cellular and molecular machinery in inhibiting radiation-induced metastatic progression, which is poorly understood so far, need to be strengthened. Ionizing radiation was known to prompt cancer cell's metastatic ability by eliciting Transforming Growth Factor-beta (TGF-β), a key regulator in epithelial-mesenchymal transdifferentiation and radio-resistance. In this viewpoint, we employed thymoquinone as a radiosensitizer to investigate its migration and invasion reversal abilities in irradiated breast cancer cell lines by assessing their respective attributes. The role of metastasis regulatory molecules like TGF-β, E-cadherin, and integrin αV and its downstream molecules were determined using RT-PCR, western blotting, immunofluorescence, and extracellular TGF-β levels affirmed through ELISA assays. These studies affirmed the TGF-β restoring ability of thymoquinone in radiation-driven migration and invasion. Also, results demonstrated that the epithelial markers E-cadherin and cytokeratin 19 were downregulated whereas mesenchymal markers like integrin αV, MMP9, and MMP2 were upregulated by irradiation treatment; however thymoquinone pre-sensitization has reverted the expression of these proteins back to control proteins expression. Here, paclitaxel was chosen as an apoptosis inducer in TGF-β restored cells and confirmed its cytotoxic effects in radiation alone and thymoquinone sensitized irradiated cells. We conclude that this therapeutic modality is effective in preventing radiation-induced epithelial-mesenchymal transdifferentiation and concomitant induction of apoptosis in breast cancer.

  2. Gelam honey attenuated radiation-induced cell death in human diploid fibroblasts by promoting cell cycle progression and inhibiting apoptosis.

    PubMed

    Tengku Ahmad, Tengku Ahbrizal Farizal; Jaafar, Faizul; Jubri, Zakiah; Abdul Rahim, Khairuddin; Rajab, Nor Fadilah; Makpol, Suzana

    2014-03-24

    against gamma-irradiation by attenuating radiation-induced cell death.

  3. Gelam honey attenuated radiation-induced cell death in human diploid fibroblasts by promoting cell cycle progression and inhibiting apoptosis

    PubMed Central

    2014-01-01

    Gelam honey acts a radioprotector against gamma-irradiation by attenuating radiation-induced cell death. PMID:24655584

  4. YThe BigH3 Tumor Suppressor Gene in Radiation-Induced Malignant Transformation of Human Bronchial Epithelial Cells

    NASA Astrophysics Data System (ADS)

    Zhao, Y.; Shao, G.; Piao, C.; Hei, T.

    Carcinogenesis is a multi-stage process with sequences of genetic events governing the phenotypic expression of a series of transformation steps leading to the development of metastatic cancer Previous studies from this laboratory have identified a 7 fold down- regulation of the novel tumor suppressor Big-h3 among radiation induced tumorigenic BEP2D cells Furthermore ectopic re-expression of this gene suppresses tumorigenic phenotype and promotes the sensitivity of these tumor cells to etoposide-induced apoptosis To extend these studies using a genomically more stable bronchial cell line we ectopically expresses the catalytic subunit of telomerase hTERT in primary human small airway epithelial SAE cells and generated several clonal cell lines that have been continuously in culture for more than 250 population doublings and are considered immortal Comparably-treated control SAE cells infected with only the viral vector senesced after less than 10 population doublings The immortalized clones demonstrated anchorage dependent growth and are non-tumorigenic in nude mice These cells show no alteration in the p53 gene but a decrease in p16 expression Exponentially growing SAEh cells were exposed to graded doses of 1 GeV nucleon of 56 Fe ions accelerated at the Brookhaven National Laboratory Irradiated cells underwent gradual phenotypic alterations after extensive in vitro cultivation Transformed cells developed through a series of successive steps before becoming anchorage independent in semisolid medium These findings indicate

  5. Post Treatment With an FGF Chimeric Growth Factor Enhances Epithelial Cell Proliferation to Improve Recovery From Radiation-Induced Intestinal Damage

    SciTech Connect

    Nakayama, Fumiaki; Hagiwara, Akiko; Umeda, Sachiko; Asada, Masahiro; Goto, Megumi; Oki, Junko; Suzuki, Masashi; Imamura, Toru; Akashi, Makoto

    2010-11-01

    Purpose: A fibroblast growth factor (FGF) 1-FGF2 chimera (FGFC) was created previously and showed greater structural stability than FGF1. This chimera was capable of stimulating epithelial cell proliferation much more strongly than FGF1 or FGF2 even without heparin. Therefore FGFC was expected to have greater biologic activity in vivo. This study evaluated and compared the protective activity of FGFC and FGF1 against radiation-induced intestinal injuries. Methods and Materials: We administered FGFC and FGF1 intraperitoneally to BALB/c mice 24 h before or after total-body irradiation (TBI). The numbers of surviving crypts were determined 3.5 days after TBI with gamma rays at doses ranging from 8 to 12 Gy. Results: The effect of FGFC was equal to or slightly superior to FGF1 with heparin. However, FGFC was significantly more effective in promoting crypt survival than FGF1 (p < 0.01) when 10 {mu}g of each FGF was administered without heparin before irradiation. In addition, FGFC was significantly more effective at promoting crypt survival (p < 0.05) than FGF1 even when administered without heparin at 24 h after TBI at 10, 11, or 12 Gy. We found that FGFC post treatment significantly promoted 5-bromo-2'-deoxyuridine incorporation into crypts and increased crypt depth, resulting in more epithelial differentiation. However, the number of apoptotic cells in FGFC-treated mice decreased to almost the same level as that in FGF1-treated mice. Conclusions: These findings suggest that FGFC strongly enhanced radioprotection with the induction of epithelial proliferation without exogenous heparin after irradiation and is useful in clinical applications for both the prevention and post treatment of radiation injuries.

  6. Novel Regenerative Peptide TP508 Mitigates Radiation-Induced Gastrointestinal Damage By Activating Stem Cells and Preserving Crypt Integrity

    PubMed Central

    Kantara, Carla; Moya, Stephanie M.; Houchen, Courtney W.; Umar, Shahid; Ullrich, Robert L.; Singh, Pomila; Carney, Darrell H.

    2015-01-01

    In recent years, increasing threats of radiation exposure and nuclear disasters have become a significant concern for the United States and countries worldwide. Exposure to high doses of radiation triggers a number of potentially lethal effects. Among the most severe is the gastrointestinal (GI) toxicity syndrome caused by the destruction of the intestinal barrier, resulting in bacterial translocation, systemic bacteremia, sepsis and death. The lack of effective radioprotective agents capable of mitigating radiation-induced damage has prompted a search for novel countermeasures that can mitigate the effects of radiation post-exposure, accelerate tissue repair in radiation-exposed individuals, and prevent mortality. We report that a single injection of regenerative peptide TP508 (rusalatide acetate, Chrysalin®) 24h after lethal radiation exposure (9Gy, LD100/15) appears to significantly increase survival and delay mortality by mitigating radiation-induced intestinal and colonic toxicity. TP508 treatment post-exposure prevents the disintegration of gastrointestinal crypts, stimulates the expression of adherens junction protein E-cadherin, activates crypt cell proliferation, and decreases apoptosis. TP508 post-exposure treatment also up-regulates the expression of DCLK1 and LGR5 markers of stem cells that have been shown to be responsible for maintaining and regenerating intestinal crypts. Thus, TP508 appears to mitigate the effects of GI toxicity by activating radioresistant stem cells and increasing the stemness potential of crypts to maintain and restore intestinal integrity. These results suggest that TP508 may be an effective emergency nuclear countermeasure that could be delivered within 24h post-exposure to increase survival and delay mortality, giving victims time to reach clinical sites for advanced medical treatment. PMID:26280221

  7. Novel regenerative peptide TP508 mitigates radiation-induced gastrointestinal damage by activating stem cells and preserving crypt integrity.

    PubMed

    Kantara, Carla; Moya, Stephanie M; Houchen, Courtney W; Umar, Shahid; Ullrich, Robert L; Singh, Pomila; Carney, Darrell H

    2015-11-01

    In recent years, increasing threats of radiation exposure and nuclear disasters have become a significant concern for the United States and countries worldwide. Exposure to high doses of radiation triggers a number of potentially lethal effects. Among the most severe is the gastrointestinal (GI) toxicity syndrome caused by the destruction of the intestinal barrier, resulting in bacterial translocation, systemic bacteremia, sepsis, and death. The lack of effective radioprotective agents capable of mitigating radiation-induced damage has prompted a search for novel countermeasures that can mitigate the effects of radiation post exposure, accelerate tissue repair in radiation-exposed individuals, and prevent mortality. We report that a single injection of regenerative peptide TP508 (rusalatide acetate, Chrysalin) 24 h after lethal radiation exposure (9 Gy, LD100/15) appears to significantly increase survival and delay mortality by mitigating radiation-induced intestinal and colonic toxicity. TP508 treatment post exposure prevents the disintegration of GI crypts, stimulates the expression of adherens junction protein E-cadherin, activates crypt cell proliferation, and decreases apoptosis. TP508 post-exposure treatment also upregulates the expression of DCLK1 and LGR5 markers of stem cells that have been shown to be responsible for maintaining and regenerating intestinal crypts. Thus, TP508 appears to mitigate the effects of GI toxicity by activating radioresistant stem cells and increasing the stemness potential of crypts to maintain and restore intestinal integrity. These results suggest that TP508 may be an effective emergency nuclear countermeasure that could be delivered within 24 h post exposure to increase survival and delay mortality, giving victims time to reach clinical sites for advanced medical treatment.

  8. Ribonuclease binase apoptotic signature in leukemic Kasumi-1 cells.

    PubMed

    Mitkevich, Vladimir A; Kretova, Olga V; Petrushanko, Irina Yu; Burnysheva, Ksenia M; Sosin, Dmitry V; Simonenko, Olga V; Ilinskaya, Olga N; Tchurikov, Nickolai A; Makarov, Alexander A

    2013-06-01

    Cytotoxic exogenous RNases triggering apoptotic response in malignant cells have potential as anticancer drugs; surprisingly, detailed characterization of the RNase-induced apoptosis has not been conducted so far. Here we show that a cytotoxic RNase from Bacillus intermedius (binase) induces extrinsic and intrinsic apoptotic pathways in leukemic Kasumi-1 cells. The experiments were performed using TaqMan Array Human Apoptosis 96-well Plate for gene expression analysis, and flow cytometry. Cytometric studies demonstrated dissipation of the mitochondrial membrane potential, opening of mitochondrial permeability transition pores, activation of caspases, increase of intracellular Ca(2+) and decrease of reactive oxygen species levels. We found that expression of 62 apoptotic genes is up-regulated, including 16 genes that are highly up-regulated, and only one gene was found to be down-regulated. The highest, 16 fold increase of the expression level was observed for TNF gene. Highly up-regulated genes also include the non-canonical NF-κB signaling pathway and inflammatory caspases 1,4. The obtained results suggest that binase induces evolutionary acquired cellular response to a microbial agent and triggers unusual apoptosis pathway. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  9. The complexity of apoptotic cell death in mollusks: An update.

    PubMed

    Romero, A; Novoa, B; Figueras, A

    2015-09-01

    Apoptosis is a type of programmed cell death that produces changes in cell morphology and in biochemical intracellular processes without inflammatory reactions. The components of the apoptotic pathways are conserved throughout evolution. Caspases are key molecules involved in the transduction of the death signal and are responsible for many of the biochemical and morphological changes associated with apoptosis. Nowadays, It is known that caspases are activated through two major apoptotic pathways (the extrinsic or death receptor pathway and the intrinsic or mitochondrial pathway), but there are also evidences of at least other alternative pathway (the perforin/granzyme pathway). Apoptosis in mollusks seems to be similar in complexity to apoptosis in vertebrates but also has unique features maybe related to their recurrent exposure to environmental changes, pollutants, pathogens and also related to the sedentary nature of some stages in the life cycle of mollusks bivalves and gastropods. As in other animals, apoptotic process is involved in the maintenance of tissue homeostasis and also constitutes an important immune response that can be triggered by a variety of stimuli, including cytokines, hormones, toxic insults, viruses, and protozoan parasites. The main goal of this work is to present the current knowledge of the molecular mechanisms of apoptosis in mollusks and to highlight those steps that need further study.

  10. Administration of donor apoptotic cells: an alternative cell-based therapy to induce tolerance?

    PubMed

    Kleinclauss, François; Perruche, Sylvain; Cahn, Jean-Yves; Tiberghien, Pierre; Saas, Philippe

    2003-05-15

    Apoptotic cells are endowed with immunomodulatory properties. The authors propose infusing apoptotic cells as a cell-based therapy product to facilitate allogeneic hematopoietic engraftment after a nonmyeloablative conditioning regimen. Such an approach may be used to obtain macrochimerism in combined hematopoietic cells and solid organ transplantation. In this article, the authors describe the mechanisms of combined hematopoietic and organ allograft transplantation and the potential difficulties. The authors discuss how intravenous apoptotic cell infusion may influence the outcome of combined transplantation. This may prove to be an interesting approach for future development in cell therapy.

  11. Volume increase and spatial shifts of chromosome territories in nuclei of radiation-induced polyploidizing tumour cells.

    PubMed

    Schwarz-Finsterle, Jutta; Scherthan, Harry; Huna, Anda; González, Paula; Mueller, Patrick; Schmitt, Eberhard; Erenpreisa, Jekaterina; Hausmann, Michael

    2013-08-30

    The exposure of tumour cells to high doses of ionizing radiation can induce endopolyploidization as an escape route from cell death. This strategy generally results in mitotic catastrophe during the first few days after irradiation. However, some cells escape mitotic catastrophe, polyploidize and attempt to undergo genome reduction and de-polyploidization in order to create new, viable para-diploid tumour cell sub-clones. In search for the consequences of ionizing radiation induced endopolyploidization, genome and chromosome architecture in nuclei of polyploid tumour cells, and sub-nuclei after division of bi- or multi-nucleated cells were investigated during 7 days following irradiation. Polyploidization was induced in p53-function deficient HeLa cells by exposure to 10Gy of X-irradiation. Chromosome territories #1, #4, #12 and centromeres of chromosomes #6, #10, #X were labelled by FISH and analysed for chromosome numbers, volumes and spatial distribution during 7 days post irradiation. The numbers of interphase chromosome territories or centromeres, respectively, the positions of the most peripherally and centrally located chromosome territories, and the territory volumes were compared to non-irradiated controls over this time course. Nuclei with three copies of several chromosomes (#1, #6, #10, #12, #X) were found in the irradiated as well as non-irradiated specimens. From day 2 to day 5 post irradiation, chromosome territories (#1, #4, #12) shifted towards the nuclear periphery and their volumes increased 16- to 25-fold. Consequently, chromosome territories returned towards the nuclear centre during day 6 and 7 post irradiation. In comparison to non-irradiated cells (∼500μm(3)), the nuclear volume of irradiated cells was increased 8-fold (to ∼4000μm(3)) at day 7 post irradiation. Additionally, smaller cell nuclei with an average volume of about ∼255μm(3) were detected on day 7. The data suggest a radiation-induced generation of large intra

  12. Apoptotic-like programmed cell death in plants.

    PubMed

    Reape, Theresa J; McCabe, Paul F

    2008-01-01

    Programmed cell death (PCD) is now accepted as a fundamental cellular process in plants. It is involved in defence, development and response to stress, and our understanding of these processes would be greatly improved through a greater knowledge of the regulation of plant PCD. However, there may be several types of PCD that operate in plants, and PCD research findings can be confusing if they are not assigned to a specific type of PCD. The various cell-death mechanisms need therefore to be carefully described and defined. This review describes one of these plant cell death processes, namely the apoptotic-like PCD (AL-PCD). We begin by examining the hallmark 'apoptotic-like' features (protoplast condensation, DNA degradation) of the cell's destruction that are characteristic of AL-PCD, and include examples of AL-PCD during the plant life cycle. The review explores the possible cellular 'executioners' (caspase-like molecules; mitochondria; de novo protein synthesis) that are responsible for the hallmark features of the cellular destruction. Finally, senescence is used as a case study to show that a rigorous definition of cell-death processes in plant cells can help to resolve arguments that occur in the scientific literature regarding the timing and control of plant cell death.

  13. Detection of apoptotic cells using propidium iodide staining.

    PubMed

    Newbold, Andrea; Martin, Ben P; Cullinane, Carleen; Bots, Michael

    2014-11-03

    Flow cytometry assays are often used to detect apoptotic cells in in vitro cultures. Depending on the experimental model, these assays can also be useful in evaluating apoptosis in vivo. In this protocol, we describe a propidium iodide (PI) flow cytometry assay to evaluate B-cell lymphomas that have undergone apoptosis in vivo. B-cell lymphoma cells are injected into recipient mice and, on tumor formation, the mice are treated with the apoptosis inducer vorinostat (a histone deacetylase inhibitor). Tumor samples collected from the lymph nodes and/or the spleen are used to prepare a single-cell suspension that is exposed to a hypotonic solution containing the fluorochrome PI. The DNA content of the cells, now labeled with PI, is analyzed by flow cytometry. Nuclear DNA content is lost during apoptosis, resulting in a hypodiploid (or sub-G1) DNA profile during flow cytometry. In contrast, healthy cells display a sharp diploid DNA profile.

  14. Effect of Tinospora cordifolia on the reduction of ultraviolet radiation-induced cytotoxicity and DNA damage in PC12 cells.

    PubMed

    Masuma, Runa; Okuno, Tsutomu; Kabir Choudhuri, Mohammad Shahabuddin; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    The safety of Tinospora cordifolia and its potential to protect against ultraviolet radiation-induced cytotoxicity and DNA damage in PC12 cells were investigated. To evaluate the safety of T. cordifolia, cell viability and agarose gel electrophoresis were carried out using PC12 cells treated with 0 to 100 μg mL(-1) of methanol extract of T. cordifolia. T. cordifolia extracts did not show cytotoxicity ranging 0 to 100 μg mL(-1). In addition, T. cordifolia extracts significantly increased cell viability at 1 ng, 10 ng and 1 μg mL(-1) concentrations in serum-deprived medium compared to control. To confirm the protective role against UV-induced damage, PC12 cells alone or in the presence of 10 ng, 100 ng, or 1 μg mL(-1) of T. cordifolia extract were exposed to 250, 270 and 290 nm of UV radiation, which corresponded to doses of 120, 150 and 300 mJ cm(-2), respectively. Treatment with T. cordifolia extracts significantly increased the cell survival rate irradiated at 290 nm. In addition, T. cordifolia extracts significantly reduced cyclobutane pyrimidine dimer formation induced by UV irradiation at all wavelengths. In conclusion, T. cordifolia is not toxic and safe for cells. Our findings can support its application as phototherapy in the medical sector.

  15. Effect of corticosteroid treatment on cell recovery by lung lavage in acute radiation-induced lung injury

    SciTech Connect

    Wesselius, L.J.; Floreani, A.A.; Kimler, B.F.; Papasian, C.J.; Dixon, A.Y. )

    1989-11-01

    The purpose of this study was to quantitate cell populations recovered by lung lavage up to 6 weeks following thoracic irradiation (24 Gy) as an index of the acute inflammatory response within lung structures. Additionally, rats were treated five times weekly with intraperitoneal saline (0.3 cc) or methylprednisolone (7.5 mg/kg/week). Lung lavage of irradiated rats recovered increased numbers of total cells compared to controls beginning 3 weeks after irradiation (P less than 0.05). The initial increase in number of cells recovered was attributable to an influx of neutrophils (P less than 0.05), and further increases at 4 and 6 weeks were associated with increased numbers of recovered macrophages (P less than 0.05). Lung lavage of steroid-treated rats at 6 weeks after irradiation recovered increased numbers of all cell populations compared to controls (P less than 0.05); however, numbers of recovered total cells, macrophages, neutrophils, and lymphocytes were all significantly decreased compared to saline-treated rats (P less than 0.05). The number of inflammatory cells recovered by lung lavage during acute radiation-induced lung injury is significantly diminished by corticosteroid treatment. Changes in cells recovered by lung lavage can also be correlated with alteration in body weight and respiration rate subsequent to treatment with thoracic irradiation and/or corticosteroids.

  16. Ultraviolet radiation-induced damage to human Langerhans cells in vivo is not reversed by ultraviolet A or visible light.

    PubMed

    Alcalay, J; Goldberg, L H; Wolf, J E; Kripke, M L

    1990-08-01

    Exposure of human skin in vivo to UVB radiation induces pyrimidine dimers in DNA and alters the morphology and function of epidermal Langerhans cells. Cells in human skin have been reported to contain a photoreactivation repair mechanism that, following exposure to UVA or visible light, repairs UVB-induced pyrimidine dimers. The purpose of this study was to determine whether exposure to photoreactivating light would also reverse the UVB-induced morphologic alterations in human Langerhans cells. The skin of eight healthy volunteers was exposed to a low dose of UVB radiation (between 0.75 and 1.5 times the minimal erythema dose), and immediately thereafter exposed to photoreactivating light from either BLB fluorescent lamps (UVA radiation) or incandescent bulbs (visible light). After exposure to UVB radiation, the number of ATPase+ epidermal Langerhans cells was reduced in all subjects to between 21% and 65% of that in unirradiated skin, and the majority of the remaining cells exhibited morphologic alterations. Exposure of the UVB-irradiated skin to photoreactivating light did not reverse or reduce these effects. We conclude that UVB-induced morphologic alterations of human Langerhans cells are not subject to photoreactivation. These results imply either that pyrimidine dimers are not involved in these effects of UVB irradiation, or that photoreactivation does not occur in human Langerhans cells in situ.

  17. Parallel single-cell analysis of active caspase-3/7 in apoptotic and non-apoptotic cells.

    PubMed

    Ledvina, Vojtěch; Janečková, Eva; Matalová, Eva; Klepárník, Karel

    2017-01-01

    Analysing the chemical content of individual cells has already been proven to reveal unique information on various biological processes. Single-cell analysis provides more accurate and reliable results for biology and medicine than analyses of extracts from cell populations, where a natural heterogeneity is averaged. To meet the requirements in the research of important biologically active molecules, such as caspases, we have developed a miniaturized device for simultaneous analyses of individual cells. A stainless steel body with a carousel holder enables high-sensitivity parallel detections in eight microvials. The holder is mounted in front of a photomultiplier tube with cooled photocathode working in photon counting mode. The detection of active caspase-3/7, central effector caspases in apoptosis, in single cells is based on the bioluminescence chemistry commercially available as Caspase-Glo(®) 3/7 reagent developed by Promega. Individual cells were captured from a culture medium under microscope and transferred by micromanipulator into detection microvial filled with the reagent. As a result of testing, the limits of detection and quantification were determined to be 0.27/0.86 of active caspase-3/7 content in an average apoptotic cell and 0.46/2.92 for non-apoptotic cells. Application potential of this technology in laboratory diagnostics and related medical research is discussed. Graphical abstract Miniaturized device for simultaneous analyses of individual cells.

  18. The different radiation response and radiation-induced bystander effects in colorectal carcinoma cells differing in p53 status.

    PubMed

    Widel, Maria; Lalik, Anna; Krzywon, Aleksandra; Poleszczuk, Jan; Fujarewicz, Krzysztof; Rzeszowska-Wolny, Joanna

    2015-08-01

    Radiation-induced bystander effect, appearing as different biological changes in cells that are not directly exposed to ionizing radiation but are under the influence of molecular signals secreted by irradiated neighbors, have recently attracted considerable interest due to their possible implication for radiotherapy. However, various cells present diverse radiosensitivity and bystander responses that depend, inter alia, on genetic status including TP53, the gene controlling the cell cycle, DNA repair and apoptosis. Here we compared the ionizing radiation and bystander responses of human colorectal carcinoma HCT116 cells with wild type or knockout TP53 using a transwell co-culture system. The viability of exposed to X-rays (0-8 Gy) and bystander cells of both lines showed a roughly comparable decline with increasing dose. The frequency of micronuclei was also comparable at lower doses but at higher increased considerably, especially in bystander TP53-/- cells. Moreover, the TP53-/- cells showed a significantly elevated frequency of apoptosis, while TP53+/+ counterparts expressed high level of senescence. The cross-matched experiments where irradiated cells of one line were co-cultured with non-irradiated cells of opposite line show that both cell lines were also able to induce bystander effects in their counterparts, however different endpoints revealed with different strength. Potential mediators of bystander effects, IL-6 and IL-8, were also generated differently in both lines. The knockout cells secreted IL-6 at lower doses whereas wild type cells only at higher doses. Secretion of IL-8 by TP53-/- control cells was many times lower than that by TP53+/+ but increased significantly after irradiation. Transcription of the NFκBIA was induced in irradiated TP53+/+ mainly, but in bystanders a higher level was observed in TP53-/- cells, suggesting that TP53 is required for induction of NFκB pathway after irradiation but another mechanism of activation must operate in

  19. A transcriptome signature of endothelial lymphatic cells coexists with the chronic oxidative stress signature in radiation-induced post-radiotherapy breast angiosarcomas.

    PubMed

    Hadj-Hamou, Nabila-Sandra; Laé, Marick; Almeida, Anna; de la Grange, Pierre; Kirova, Youlia; Sastre-Garau, Xavier; Malfoy, Bernard

    2012-07-01

    Radiation-induced breast angiosarcomas are rare but recognized complication of breast cancer radiotherapy and are of poor prognosis. Little is known about the genetic abnormalities present in these secondary tumors. Herein, we investigated the differences in the genome and in the transcriptome that discriminate these tumors as a function of their etiology. Seven primary breast angiosarcomas and 18 secondary breast angiosarcomas arising in the irradiation field of a radiotherapy were analyzed. Copy number alterations and gene expression were analyzed using Affymetrix SNP 6.0 Array and Affymetrix Exon Arrays, respectively. We showed that two transcriptome signatures of the radiation tumorigenesis coexisted in these tumors. One was histology specific and correctly discriminated 100% of the primary tumors from the radiation-induced tumors. The deregulation of marker genes, including podoplanin (PDPN), prospero homeobox 1 (PROX-1), vascular endothelial growth factor 3 (VEGFR3) and endothelin receptor A (EDNRA), suggests that the radiation-induced breast angiosarcomas developed from radiation-stimulated lymphatic endothelial cells. None of the genes of the histology-specific signature were present in our previously published signature of the radiation tumorigenesis which shows the presence of a chronic oxidative stress in radiation-induced sarcomas of various histologies. Nevertheless, this oxidative stress signature classified correctly 88% of the breast angiosarcomas as a function of the etiology. In contrast, MYC amplification, which is observed in all radiation-induced tumors but also at a low rate in primary tumors, was not a marker of the radiation tumorigenesis.

  20. Low Concentration of Exogenous Carbon Monoxide Modulates Radiation-Induced Bystander Effect in Mammalian Cell Cluster Model.

    PubMed

    Wu, Wenqing; Nie, Lili; Yu, K N; Wu, Lijun; Kong, Peizhong; Bao, Lingzhi; Chen, Guodong; Yang, Haoran; Han, Wei

    2016-12-08

    During radiotherapy procedures, radiation-induced bystander effect (RIBE) can potentially lead to genetic hazards to normal tissues surrounding the targeted regions. Previous studies showed that RIBE intensities in cell cluster models were much higher than those in monolayer cultured cell models. On the other hand, low-concentration carbon monoxide (CO) was previously shown to exert biological functions via binding to the heme domain of proteins and then modulating various signaling pathways. In relation, our previous studies showed that exogenous CO generated by the CO releasing molecule, tricarbonyldichlororuthenium (CORM-2), at a relatively low concentration (20 µM), effectively attenuated the formation of RIBE-induced DNA double-strand breaks (DSB) and micronucleus (MN). In the present work, we further investigated the capability of a low concentration of exogenous CO (CORM-2) of attenuating or inhibiting RIBE in a mixed-cell cluster model. Our results showed that CO (CORM-2) with a low concentration of 30 µM could effectively suppress RIBE-induced DSB (p53 binding protein 1, p53BP1), MN formation and cell proliferation in bystander cells but not irradiated cells via modulating the inducible nitric oxide synthase (iNOS) andcyclooxygenase-2 (COX-2). The results can help mitigate RIBE-induced hazards during radiotherapy procedures.

  1. Low Concentration of Exogenous Carbon Monoxide Modulates Radiation-Induced Bystander Effect in Mammalian Cell Cluster Model

    PubMed Central

    Wu, Wenqing; Nie, Lili; Yu, K. N.; Wu, Lijun; Kong, Peizhong; Bao, Lingzhi; Chen, Guodong; Yang, Haoran; Han, Wei

    2016-01-01

    During radiotherapy procedures, radiation-induced bystander effect (RIBE) can potentially lead to genetic hazards to normal tissues surrounding the targeted regions. Previous studies showed that RIBE intensities in cell cluster models were much higher than those in monolayer cultured cell models. On the other hand, low-concentration carbon monoxide (CO) was previously shown to exert biological functions via binding to the heme domain of proteins and then modulating various signaling pathways. In relation, our previous studies showed that exogenous CO generated by the CO releasing molecule, tricarbonyldichlororuthenium (CORM-2), at a relatively low concentration (20 µM), effectively attenuated the formation of RIBE-induced DNA double-strand breaks (DSB) and micronucleus (MN). In the present work, we further investigated the capability of a low concentration of exogenous CO (CORM-2) of attenuating or inhibiting RIBE in a mixed-cell cluster model. Our results showed that CO (CORM-2) with a low concentration of 30 µM could effectively suppress RIBE-induced DSB (p53 binding protein 1, p53BP1), MN formation and cell proliferation in bystander cells but not irradiated cells via modulating the inducible nitric oxide synthase (iNOS) andcyclooxygenase-2 (COX-2). The results can help mitigate RIBE-induced hazards during radiotherapy procedures. PMID:27941646

  2. Enhanced survival of Leishmania major in neutrophil granulocytes in the presence of apoptotic cells

    PubMed Central

    Hellberg, Lars; Köhl, Jörg; Laskay, Tamás

    2017-01-01

    Neutrophil granulocytes are the first leukocytes that encounter and phagocytose Leishmania major (L. major) parasites in the infected skin. The parasites can nonetheless survive within neutrophils. However, the mechanisms enabling the survival of Leishmania within neutrophils are still elusive. Previous findings indicated that human neutrophils can engulf apoptotic cells. Since apoptotic neutrophils are abundant in infected tissues, we hypothesized that the uptake of apoptotic cells results in diminished anti-leishmanial activity and, consequently, contributes to enhanced survival of the parasites at the site of infection. In the present study, we demonstrated that L. major-infected primary human neutrophils acquire enhanced capacity to engulf apoptotic cells. This was associated with increased expression of the complement receptors 1 and 3 involved in phagocytosis of apoptotic cells. Next, we showed that ingestion of apoptotic cells affects neutrophil antimicrobial functions. We observed that phagocytosis of apoptotic cells by neutrophils downregulates the phosphorylation of p38 MAPK and PKCδ, the kinases involved in activation of NADPH oxidase and hence reactive oxygen species (ROS) production. In line, uptake of apoptotic cells inhibits TNF- and L. major-induced ROS production by neutrophils. Importantly, we found that the survival of Leishmania in neutrophils is strongly enhanced in neutrophils exposed to apoptotic cells. Together, our findings reveal that apoptotic cells promote L. major survival within neutrophils by downregulating critical antimicrobial functions. This suggests that the induction of enhanced uptake of apoptotic cells represents a novel evasion mechanism of the parasites that facilitates their survival in neutrophil granulocytes. PMID:28187163

  3. Reciprocal Paracrine Interactions Between Normal Human Epithelial and Mesenchymal Cells Protect Cellular DNA from Radiation-Induced Damage

    SciTech Connect

    Nakazawa, Yuka; Saenko, Vladimir Rogounovitch, Tatiana; Suzuki, Keiji; Mitsutake, Norisato; Matsuse, Michiko; Yamashita, Shunichi

    2008-06-01

    Purpose: To explore whether interactions between normal epithelial and mesenchymal cells can modulate the extent of radiation-induced DNA damage in one or both types of cells. Methods and Materials: Human primary thyrocytes (PT), diploid fibroblasts BJ, MRC-5, and WI-38, normal human mammary epithelial cells (HMEC), and endothelial human umbilical cord vein endothelial cells (HUV-EC-C), cultured either individually or in co-cultures or after conditioned medium transfer, were irradiated with 0.25 to 5 Gy of {gamma}-rays and assayed for the extent of DNA damage. Results: The number of {gamma}-H2AX foci in co-cultures of PT and BJ fibroblasts was approximately 25% lower than in individual cultures at 1 Gy in both types of cells. Reciprocal conditioned medium transfer to individual cultures before irradiation resulted in approximately a 35% reduction of the number {gamma}-H2AX foci at 1 Gy in both types of cells, demonstrating the role of paracrine soluble factors. The DNA-protected state of cells was achieved within 15 min after conditioned medium transfer; it was reproducible and reciprocal in several lines of epithelial cells and fibroblasts, fibroblasts, and endothelial cells but not in epithelial and endothelial cells. Unlike normal cells, human epithelial cancer cells failed to establish DNA-protected states in fibroblasts and vice versa. Conclusions: The results imply the existence of a network of reciprocal interactions between normal epithelial and some types of mesenchymal cells mediated by soluble factors that act in a paracrine manner to protect DNA from genotoxic stress.

  4. Differential modulation of a radiation-induced bystander effect in glioblastoma cells by pifithrin-α and wortmannin

    NASA Astrophysics Data System (ADS)

    Shao, Chunlin; Zhang, Jianghong; Prise, Kevin M.

    2010-03-01

    The implication of radiation-induced bystander effect (RIBE) for both radiation protection and radiotherapy has attracted significant attention, but a key question is how to modulate the RIBE. The present study found that, when a fraction of glioblastoma cells in T98G population were individually targeted with precise helium particles through their nucleus, micronucleus (MN) were induced and its yield increased non-linearly with radiation dose. After co-culturing with irradiated cells, additional MN could be induced in the non-irradiated bystander cells and its yield was independent of irradiation dose, giving direct evidence of a RIBE. Further results showed that the RIBE could be eliminated by pifithrin-α (p53 inhibitor) but enhanced by wortmannin (PI3K inhibitor). Moreover, it was found that nitric oxide (NO) contributed to this RIBE, and the levels of NO of both irradiated cells and bystander cells could be extensively diminished by pifithrin-α but insignificantly reduced by wortmannin. Our results indicate that RIBE can be modulated by p53 and PI3K through a NO-dependent and NO-independent pathway, respectively.

  5. Autophagy genes function sequentially to promote apoptotic cell corpse degradation in the engulfing cell

    PubMed Central

    Li, Wei; Zou, Wei; Yang, Yihong; Chai, Yongping; Chen, Baohui; Cheng, Shiya; Tian, Dong

    2012-01-01

    Apoptotic cell degradation is a fundamental process for organism development, and impaired clearance causes inflammatory or autoimmune disease. Although autophagy genes were reported to be essential for exposing the engulfment signal on apoptotic cells, their roles in phagocytes for apoptotic cell removal are not well understood. In this paper, we develop live-cell imaging techniques to study apoptotic cell clearance in the Caenorhabditis elegans Q neuroblast lineage. We show that the autophagy proteins LGG-1/LC3, ATG-18, and EPG-5 were sequentially recruited to internalized apoptotic Q cells in the phagocyte. In atg-18 or epg-5 mutants, apoptotic Q cells were internalized but not properly degraded; this phenotype was fully rescued by the expression of autophagy genes in the phagocyte. Time-lapse analysis of autophagy mutants revealed that recruitment of the small guanosine triphosphatases RAB-5 and RAB-7 to the phagosome and the formation of phagolysosome were all significantly delayed. Thus, autophagy genes act within the phagocyte to promote apoptotic cell degradation. PMID:22451698

  6. Evidence for involvement of cytosolic thioredoxin peroxidase in the excessive resistance of Sf9 Lepidopteran insect cells against radiation-induced apoptosis.

    PubMed

    Hambarde, Shashank; Singh, Vijaypal; Chandna, Sudhir

    2013-01-01

    Lepidopteran insect cells display 50-100 times higher radioresistance compared to human cells, and reportedly have more efficient antioxidant system that can significantly reduce radiation-induced oxidative stress and cell death. However, the antioxidant mechanisms that contribute substantially to this excessive resistance still need to be understood thoroughly. In this study, we investigated the role of thioredoxin peroxidase (TPx) in high-dose γ-radiation response of Sf9 cell line derived from Spodoptera frugiperda, the Fall armyworm. We identified a TPx orthologue (Sf-TPx) in Spodoptera system, with primarily cytosolic localization. Gamma-irradiation at 500 Gy dose significantly up-regulated Sf-TPx, while higher doses (1000 Gy-2000 Gy) had no such effect. G2/M checkpoint induced following 500 Gy was associated with transition of Sf-TPx decamer into enzymatically active dimer. Same effect was observed during G2/M block induced by 5 nM okadaic acid or 10 µM CDK1 (cycline dependent kinase-1) inhibitor roscovitine, thus indicating that radiation-induced Sf-TPx activity is mediated by CDKs. Accumulation of TPx dimer form during G2/M checkpoint might favour higher peroxidase activity facilitating efficient survival at this dose. Confirming this, higher lethal doses (1000 Gy-2000 Gy) caused significantly less accumulation of dimer form and induced dose-dependent apoptosis. A ∼50% knock-down of Sf-TPx by siRNA caused remarkable increase in radiation-induced ROS as well as caspase-3 dependent radiation-induced apoptosis, clearly implying TPx role in the radioresistance of Sf9 cells. Quite importantly, our study demonstrates for the first time that thioredoxin peroxidase contributes significantly in the radioresistance of Lepidopteran Sf9 insect cells, especially in their exemplary resistance against radiation-induced apoptosis. This is an important insight into the antioxidant mechanisms existing in this highly stress-resistant model cell system.

  7. Evidence for Involvement of Cytosolic Thioredoxin Peroxidase in the Excessive Resistance of Sf9 Lepidopteran Insect Cells against Radiation-Induced Apoptosis

    PubMed Central

    Hambarde, Shashank; Singh, Vijaypal; Chandna, Sudhir

    2013-01-01

    Lepidopteran insect cells display 50–100 times higher radioresistance compared to human cells, and reportedly have more efficient antioxidant system that can significantly reduce radiation-induced oxidative stress and cell death. However, the antioxidant mechanisms that contribute substantially to this excessive resistance still need to be understood thoroughly. In this study, we investigated the role of thioredoxin peroxidase (TPx) in high-dose γ-radiation response of Sf9 cell line derived from Spodoptera frugiperda, the Fall armyworm. We identified a TPx orthologue (Sf-TPx) in Spodoptera system, with primarily cytosolic localization. Gamma-irradiation at 500 Gy dose significantly up-regulated Sf-TPx, while higher doses (1000 Gy–2000 Gy) had no such effect. G2/M checkpoint induced following 500 Gy was associated with transition of Sf-TPx decamer into enzymatically active dimer. Same effect was observed during G2/M block induced by 5 nM okadaic acid or 10 µM CDK1 (cycline dependent kinase-1) inhibitor roscovitine, thus indicating that radiation-induced Sf-TPx activity is mediated by CDKs. Accumulation of TPx dimer form during G2/M checkpoint might favour higher peroxidase activity facilitating efficient survival at this dose. Confirming this, higher lethal doses (1000 Gy–2000 Gy) caused significantly less accumulation of dimer form and induced dose-dependent apoptosis. A ∼50% knock-down of Sf-TPx by siRNA caused remarkable increase in radiation-induced ROS as well as caspase-3 dependent radiation-induced apoptosis, clearly implying TPx role in the radioresistance of Sf9 cells. Quite importantly, our study demonstrates for the first time that thioredoxin peroxidase contributes significantly in the radioresistance of Lepidopteran Sf9 insect cells, especially in their exemplary resistance against radiation-induced apoptosis. This is an important insight into the antioxidant mechanisms existing in this highly stress-resistant model cell system. PMID:23505474

  8. Intravenous apoptotic spleen cell infusion induces a TGF-beta-dependent regulatory T-cell expansion

    PubMed Central

    Kleinclauss, François M.; Perruche, Sylvain; Masson, Emeline; De Carvalho Bittencourt, Marcelo; Biichle, Sabeha; Remy-Martin, Jean-Paul; Ferrand, Christophe; Martin, Mael; Bittard, Hugues; Chalopin, Jean-Marc; Seilles, Estelle; Tiberghien, Pierre; Saas, Philippe

    2006-01-01

    Apoptotic leukocytes are endowed with immunomodulatory properties that can be used to enhance hematopoietic engraftment and prevent graft-versus-host disease. This apoptotic cell-induced tolerogenic effect is mediated by host macrophages and not recipient dendritic cells or donor phagocytes present in the bone marrow graft as evidenced by selective cell depletion and trafficking experiments. Furthermore, apoptotic cell infusion is associated with TGF-β-dependent donor CD4+CD25+ T cell expansion. Such cells have a regulatory phenotype (CD62Lhigh and intracellular CTLA-4+), express high levels of Foxp3 mRNA and exert ex vivo suppressive activity through a cell-to-cell contact mechanism. In vivo CD25 depletion after apoptotic cell infusion prevents the apoptotic spleen cell-induced beneficial effects on engraftment and graft-versus-host disease occurrence. This highlights the role of regulatory T cells in the tolerogenic effect of apoptotic spleen cell infusion. This novel association between apoptosis and regulatory T cell expansion may also contribute to preventing deleterious auto-immune responses during normal turnover. PMID:15962005

  9. Radiation-induced cell lethality of samonella typhimurium ATCC 14028: Cooperative effect of hydroxyl radical and oxygen

    SciTech Connect

    Kim, Y.A.; Thayer, D.W.

    1995-10-01

    The lethality of {gamma}-radiation doses of 0.2 to 1.0 kGy for Salmonella typhimurium ATCC 14028 was measured in the presence of air, N{sub 2} and N{sub 2}O and with the hydroxyl radical scavengers formate and polyethylene glycol (PEG), M{sub r} 8,000. Saturation of cell suspensions with either N{sub 2}O or N{sub 2}/N{sub 2}/N{sub 2}O (1:1, v/v) gas was expected to double the number of hydroxyl radicals (OH{center_dot}) and to produce an equivalent increase in lethality, but this did not occur. Adding 10% (v/v) O{sub 2} to either N{sub 2}/N{sub 2}O gas produced approximately the same {gamma}-irradiation lethality for S. typhimurium as did air. Addition of hydroxyl radical scavengers, 40 mM formate and 1.5% (w/v) PEG, significantly reduced the lethality of {gamma} radiation for S. typhimurium in the presence of air but not in the presence of N{sub 2} or N{sub 2}O gases. Membrane-permeable formate provided slightly better protection than nonpermeable PEG. Cells of S. typhimurium grown under anaerobic conditions were more sensitive to radiation, and were less protected by hydroxyl radical scavengers, especially formate, than when cells grown under aerobic conditions were irradiated in the presence of oxygen. Hydroxyl radical scavengers provided no further protection during irradiation in the absence of oxygen. These results indicated that the increased radiation sensitivity of cells grown under anaerobic conditions may be related to superoxide radicals which could increase intercellular damage during irradiation in the presence of oxygen. However, endogenous superoxide dismutase and catalase activities did not protect cells from the radiation-induced lethality of S. typhimurium. Cytoplasmic extracts protected bacterial DNA in vitro in either the presence of absence of oxygen, and no radiation-induced lipid peroxidation of the cellular components was identified by measuring the levels of 2-thiobarbituric acid. 38 refs., 4 figs., 2 tabs.

  10. Effect of intercellular contact on DNA conformation, radiation-induced DNA damage, and mutation in Chinese hamster V79 cells

    SciTech Connect

    Olive, P.L.; Durand, R.E.

    1985-01-01

    Chinese hamster V79 cells, when grown as small spheroids in suspension culture, are more resistant to killing by ionizing radiation than when grown as monolayers. The authors have attempted to determine whether this enhanced survival following irradiation is reflected in DNA damage and repair at the structural level (by measuring alkali-induced DNA unwinding rates from strand breaks) and at the functional level (by measuring resistance to forward mutation at the HGPRT locus). For a given dose of radiation, the unwinding of DNA in high salt/weak alkali was less complete for spheroid DNA than for monolayer DNA, and the rate of repair of radiation damage was faster in spheroid DNA. These differential responses were lost 8 hr after separation of spheroids into single cells, coinciding with loss of radioresistance measured by clonogenicity. In addition, spheroid cells showed fewer numbers of induced mutants per Gray, although, for a given level of survival, the mutation frequency for monolayers and spheroids was identical. These results suggest that conformational changes in DNA resulting from cell growth as spheroids might enhance repair of radiation-induced lesions.

  11. Novel mechanism for the radiation-induced bystander effect: nitric oxide and ethylene determine the response in sponge cells.

    PubMed

    Müller, Werner E G; Ushijima, Hiroshi; Batel, Renato; Krasko, Anatoli; Borejko, Alexandra; Müller, Isabel M; Schröder, Heinz-C

    2006-05-11

    Until now the bystander effect had only been described in vertebrates. In the present study the existence of this effect has been demonstrated for the phylogenetically oldest metazoan phylum, the Porifera. We used the demosponge Suberites domuncula for the experiments in the two-chamber-system. The lower dish contained irradiated "donor" cells (single cells) and the upper dish the primmorphs ("recipient" primmorphs). The "donor" cells were treated with UV-B light (40 mJ/cm2) and 100 microM hydrogen peroxide (H2O2), factors that exist also in the natural marine aquatic environment of sponges; these factors caused a high level of DNA strand breaks followed by a reduced viability of the cells. If these cells were added to the "recipient" primmorphs these 3D-cell cultures started to undergo apoptosis. This effect could be abolished by the NO-specific scavenger PTIO and ethylene. The conclusion that NO is synthesized by the UV-B/H2O2-treated cells was supported analytically. The cDNA encoding the enzyme dimethylarginine dimethylaminohydrolase (DDAH) was isolated from the "donor" cells. High levels of DDAH transcripts were measured in UV-B/H2O2-treated "donor" cells while after ethylene treatment the steady-state level of expression drops drastically. We conclude that in the absence of ethylene the concentration of the physiological inhibitor for the NO synthase ADMA is low, due to the high level of DDAH. In consequence, high amounts of NO are released from "donor" cells which cause apoptosis in "recipient" primmorphs. In contrast, ethylene reduces the DDAH expression with the consequence of higher levels of ADMA which prevent the formation of larger amounts of NO. This study describes the radiation-induced bystander effect also for the most basal metazoans and demonstrates that this effect is controlled by the two gases NO and ethylene.

  12. Use of Human Cadaveric Mesenchymal Stem Cells for Cell Therapy of a Chronic Radiation-Induced Skin Lesion: A Case Report.

    PubMed

    Portas, M; Mansilla, E; Drago, H; Dubner, D; Radl, A; Coppola, A; Di Giorgio, M

    2016-09-01

    Acute and late radiation-induced injury on skin and subcutaneous tissues are associated with substantial morbidity in radiation therapy, interventional procedures and also are of concern in the context of nuclear or radiological accidents. Pathogenesis is initiated by depletion of acutely responding epithelial tissues and damage to vascular endothelial microvessels. Efforts for medical management of severe radiation-induced lesions have been made. Nevertheless, the development of strategies to promote wound healing, including stem cell therapy, is required. From 1997 to 2014, over 248 patients were referred to the Radiopathology Committee of Hospital de Quemados del Gobierno de la Ciudad de Buenos Aires (Burns Hospital) for the diagnosis and therapy of radiation-induced localized lesions. As part of the strategies for the management of severe cases, there is an ongoing research and development protocol on 'Translational Clinical Trial phases I/II to evaluate the safety and efficacy of adult mesenchymal stem cells from bone marrow for the treatment of large burns and radiological lesions'. The object of this work was to describe the actions carried out by the Radiopathology Committee of the Burns Hospital in a chronic case with more than 30 years of evolution without positive response to conventional treatments. The approach involved the evaluation of the tissular compromise of the lesion, the prognosis and the personalized treatment, including regenerative therapy.

  13. Activation-induced CD154 expression abrogates tolerance induced by apoptotic cells*

    PubMed Central

    Gurung, Prajwal; Kucaba, Tamara A.; Ferguson, Thomas A.; Griffith, Thomas S.

    2009-01-01

    The decision to generate a productive immune response or tolerance often depends on the context in which T cells first see Ag. Using a classical system of tolerance induction, we examined the immunological consequence of Ag encountered in the presence of naïve or activated apoptotic cells. Naïve apoptotic cells induced tolerance when injected i.v.; however, previously activated apoptotic cells induced immunity. Further analysis revealed a key role for CD154, as tolerance resulted after i.v. injection of either naïve or activated apoptotic CD154−/− T cells, while co-injection of an agonistic anti-CD40 mAb with naïve apoptotic T cells induced robust immunity. DC fed activated apoptotic T cells in vitro produced IL-12p40 in a CD154-dependent manner, and the use of IL-12p40−/− mice or mAb-mediated neutralization of IL-12 revealed a link between CD154, IL-12, and the ability of activated apoptotic T cells to induce immunity rather than tolerance. Collectively these results show that CD154 expression on apoptotic T cells can determine the outcome of an immune response to Ag recognized within the context of the apoptotic cells, and suggest the balance between naïve and activated apoptotic T cells may dictate whether a productive immune response is encouraged. PMID:19841180

  14. Apoptotic hair cell death after transient cochlear ischemia in gerbils.

    PubMed

    Taniguchi, Masafumi; Hakuba, Nobuhiro; Koga, Kenichiro; Watanabe, Futoshi; Hyodo, Jun; Gyo, Kiyofumi

    2002-12-20

    The mechanisms of cochlear hair cell death following exposure to transient inner ear ischemia were investigated in gerbils histologically. The animals were subjected to ischemic insult by occluding both vertebral arteries for 15 min. Hoechst 33342 nuclear staining showed that inner hair cells (IHCs) underwent sporadic degeneration via nuclear condensation, which peaked 12 hours after the ischemia. Furthermore, nuclear DNA fragmentation was noted by the terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-biotin nick end labeling method. Transmission electron microscopy revealed morphological changes in the IHCs characteristic of apoptosis, including karyopyknosis, chromatin condensation. These findings suggest that apoptotic cell death is the major process in hair cell degeneration in this animal model.

  15. Inhibition of hematopoietic recovery from radiation-induced myelosuppression by natural killer cells

    SciTech Connect

    Pantel, K.; Boertman, J.; Nakeff, A. )

    1990-05-01

    We have examined the role of natural killer (NK) cells in situ in the recovery of marrow hematopoiesis in B6D2F1 mice receiving various doses of total-body irradiation (TBI) as a well-characterized model for treatment-induced myelosuppression. Applying an in situ cytotoxic approach for ablating NK 1.1 cells, we have demonstrated that NK 1.1 cells differentially inhibit the recovery of hematopoietic stem cells (CFU-S) and their progenitor cells committed to granulocyte-macrophage differentiation from a sublethal dose of TBI (9 Gy) while not affecting the recovery of progenitor cells committed to either erythroid or megakaryocyte differentiation from TBI. However, recoveries of CFU-S and progenitor cells were unaffected by the ablation of NK cells prior to a moderate dose of TBI (2 Gy). These findings provide in situ evidence that NK cells are potential inhibitors of hematopoietic recovery from treatment-induced myelosuppression.

  16. CDC42 Gtpase Activation Affects Hela Cell DNA Repair and Proliferation Following UV Radiation-Induced Genotoxic Stress.

    PubMed

    Ascer, Liv G; Magalhaes, Yuli T; Espinha, Gisele; Osaki, Juliana H; Souza, Renan C; Forti, Fabio L

    2015-09-01

    Cell division control protein 42 (CDC42) homolog is a small Rho GTPase enzyme that participates in such processes as cell cycle progression, migration, polarity, adhesion, and transcription. Recent studies suggest that CDC42 is a potent tumor suppressor in different tissues and is related to aging processes. Although DNA damage is crucial in aging, a potential role for CDC42 in genotoxic stress remains to be explored. Migration, survival/proliferation and DNA damage/repair experiments were performed to demonstrate CDC42 involvement in the recovery of HeLa cells exposed to ultraviolet radiation-induced stress. Sub-lines of HeLa cells ectopically expressing the constitutively active CDC42-V12 mutant were generated to examine whether different CDC42-GTP backgrounds might reflect different sensitivities to UV radiation. Our results show that CDC42 constitutive activation does not interfere with HeLa cell migration after UV radiation. However, the minor DNA damage exhibited by the CDC42-V12 mutant exposed to UV radiation most likely results in cell cycle arrest at the G2/M checkpoint and reduced proliferation and survival. HeLa cells and Mock clones, which express endogenous wild-type CDC42 and show normal activity, are more resistant to UV radiation. None of these effects are altered by pharmacological CDC42 inhibition. Finally, the phosphorylation status of the DNA damage response proteins γ-H2AX and p-Chk1 was found to be delayed and attenuated, respectively, in CDC42-V12 clones. In conclusion, the sensitivity of HeLa cells to ultraviolet radiation increases with CDC42 over-activation due to inadequate DNA repair signaling, culminating in G2/M cell accumulation, which is translated into reduced cellular proliferation and survival.

  17. Ultraviolet-A radiation induces changes in cyclin G gene expression in mouse melanoma B16-F1 cells

    PubMed Central

    Pastila, Riikka; Leszczynski, Dariusz

    2007-01-01

    Background We have previously shown that ultraviolet-A (UVA) radiation enhances metastatic lung colonization capacity of B16-F1 melanoma cells. The aim of this study was to examine changes in expression profile of genes in mouse melanoma B16-F1 cells exposed to UVA radiation. Results B16-F1 melanoma cells were exposed to a single UVA radiation dose of 8 J/cm2 and mRNA was isolated 4 h after the end of UVA exposure. Atlas™ Mouse Cancer 1.2 cDNA expression arrays were used for the large-scale screening to identify the genes involved in the regulation of carcinogenesis, tumor progression and metastasis. Physiologically relevant UVA dose induced differential expression in 9 genes in the UVA exposed melanoma cells as compared to the unexposed control cells. The expression of seven genes out of nine was upregulated (HSC70, HSP86, α-B-crystallin, GST mu2, Oxidative stress induced protein OSI, VEGF, cyclin G), whereas the expression of two genes was down-regulated (G-actin, non-muscle cofilin). The gene expression of cyclin G was mostly affected by UVA radiation, increasing by 4.85-folds 4 hour after exposure. The analysis of cyclin G protein expression revealed 1.36-fold increase at the 6 hour time point after UVA exposure. Cell cycle arrest in G2/M phase, which is known to be regulated by cyclin G, occurred at 4-h hour time-point, peaking 8 hours after the end of UVA irradiation, suggesting that cyclin G might play a role in the cell cycle arrest. Conclusion Our results suggest that UVA radiation-induces changes in the expression of several genes. Some of these changes, e.g. in expression of cyclin G, possibly might affect cell physiology (cell cycle arrest). PMID:17474990

  18. 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.

  19. 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.

  20. Radiation Induced Genomic Instability

    SciTech Connect

    Morgan, William F.

    2011-03-01

    Radiation induced genomic instability can be observed in the progeny of irradiated cells multiple generations after irradiation of parental cells. The phenotype is well established both in vivo (Morgan 2003) and in vitro (Morgan 2003), and may be critical in radiation carcinogenesis (Little 2000, Huang et al. 2003). Instability can be induced by both the deposition of energy in irradiated cells as well as by signals transmitted by irradiated (targeted) cells to non-irradiated (non-targeted) cells (Kadhim et al. 1992, Lorimore et al. 1998). Thus both targeted and non-targeted cells can pass on the legacy of radiation to their progeny. However the radiation induced events and cellular processes that respond to both targeted and non-targeted radiation effects that lead to the unstable phenotype remain elusive. The cell system we have used to study radiation induced genomic instability utilizes human hamster GM10115 cells. These cells have a single copy of human chromosome 4 in a background of hamster chromosomes. Instability is evaluated in the clonal progeny of irradiated cells and a clone is considered unstable if it contains three or more metaphase sub-populations involving unique rearrangements of the human chromosome (Marder and Morgan 1993). Many of these unstable clones have been maintained in culture for many years and have been extensively characterized. As initially described by Clutton et al., (Clutton et al. 1996) many of our unstable clones exhibit persistently elevated levels of reactive oxygen species (Limoli et al. 2003), which appear to be due dysfunctional mitochondria (Kim et al. 2006, Kim et al. 2006). Interestingly, but perhaps not surprisingly, our unstable clones do not demonstrate a “mutator phenotype” (Limoli et al. 1997), but they do continue to rearrange their genomes for many years. The limiting factor with this system is the target – the human chromosome. While some clones demonstrate amplification of this chromosome and thus lend

  1. Differential expression of cell adhesion molecules in an ionizing radiation-induced breast cancer model system.

    PubMed

    Calaf, Gloria M; Roy, Debasish; Narayan, Gopeshwar; Balajee, Adayabalam S

    2013-07-01

    Cell-cell adhesion is mediated by members of the cadherin-catenin system and among them E-cadherin and β-catenin are important adhesion molecules for epithelial cell function and preservation of tissue integrity. To investigate the importance of cell adhesion molecules in breast carcinogenesis, we developed an in vitro breast cancer model system wherein immortalized human breast epithelial cell line, MCF-10F, was malignantly transformed by exposure to low doses of high linear energy transfer (LET) α particle radiation (150 keV/µm) and subsequent growth in the presence or absence of 17β-estradiol. This model consisted of human breast epithelial cells in different stages of transformation: i) parental cell line MCF-10F; ii) MCF-l0F continuously grown with estradiol at 10(-8) (Estrogen); iii) a non-malignant cell line (Alpha3); and iv) a malignant and tumorigenic cell line (Alpha5) and the Tumor2 cell line derived from the nude mouse xenograft of the Alpha5 cell line. Expression levels of important cell adhesion molecules such as α-catenin, β-catenin, γ-catenin, E-cadherin and integrin were found to be higher at the protein level in the Alpha5 and Tumor2 cell lines relative to these levels in the non-tumorigenic MCF-10F, Estrogen and Alpha3 cell lines. In corroboration, cDNA expression analysis revealed elevated levels of genes involved in the cell adhesion function [E-cadherin, integrin β6 and desmocollin3 (DSc3)] in the Alpha5 and Tumor2 cell lines relative to the levels in the MCF-10F, Estrogen and Alpha3 cell lines. Collectively, our results suggest that cell adhesion molecules are expressed at higher levels in malignantly transformed breast epithelial cells relative to levels in non-malignant cells. However, reduced levels of adhesion molecules observed in the mouse xenograft-derived Tumor 2 cell line compared to the pre-tumorigenic Alpha5 cell line suggests that the altered expression levels of adhesion molecules depend on the tumor tissue

  2. Cell cycle age dependence for radiation-induced G/sub 2/ arrest: evidence for time-dependent repair

    SciTech Connect

    Rowley, R.

    1985-09-01

    Exponentially growing eucaryotic cells, irradiated in interphase, are delayed in progression to mitosis chiefly by arrest in G/sub 2/. The sensitivity of Chinese hamster ovary cells to G/sub 2/ arrest induction by X rays increases through the cell cycle, up to the X-ray transition point (TP) in G/sub 2/. This age response can be explained by cell cycle age-dependent changes in susceptibility of the target(s) for G/sub 2/ arrest and/or by changes in capability for postirradiation recovery from G/sub 2/ arrest damage. Discrimination between sensitivity changes and repair phenomena is possible only if the level of G/sub 2/ arrest-causing damage sustained by a cell at the time of irradiation and the level ultimately expressed as arrest can be determined. The ability of caffeine to ameliorate radiation-induced G/sub 2/ arrest, while inhibiting repair of G/sub 2/ arrest-causing damage makes such an analysis possible. In the presence of caffeine, progression of irradiated cells was relatively unperturbed, but on caffeine removal, G/sub 2/ arrest was expressed. The duration of G/sub 2/ arrest was independent of the length of the prior caffeine exposure. This finding indicates that the target for G/sub 2/ arrest induction is present throughout the cell cycle and that the level of G/sub 2/ arrest damage incurred is initially constant for all cell cycle phases. The data are consistent with the existence of a time-dependent recovery mechanism to explain the age dependence for radiation induction of G/sub 2/ arrest.

  3. Radiation-Induced RhoGDIβ Cleavage Leads to Perturbation of Cell Polarity: A Possible Link to Cancer Spreading.

    PubMed

    Fujiwara, Mamoru; Okamoto, Mayumi; Hori, Masato; Suga, Hiroshi; Jikihara, Hiroshi; Sugihara, Yuka; Shimamoto, Fumio; Mori, Toshio; Nakaoji, Koichi; Hamada, Kazuhiko; Ota, Takahide; Wiedemuth, Ralf; Temme, Achim; Tatsuka, Masaaki

    2016-11-01

    The equilibrium between proliferation and apoptosis is tightly balanced to maintain tissue homeostasis in normal tissues and even in tumors. Achieving and maintaining such a balance is important for cancer regrowth and spreading after cytotoxic treatments. Caspase-3 activation and tumor cell death following anticancer therapy as well as accompanying cell death pathways are well characterized, but their association to homeostasis of cancerous tissue and tumor progression remains poorly understood. Here we proposed a novel mechanism of cancer spreading induced by caspase-3. RhoGDIβ, known as a direct cleavage substrate of caspase-3, is overexpressed in many epithelial cancers. The N-terminal-truncated RhoGDIβ (ΔN-RhoGDIβ) is accumulated in caspase-3-activated cells. Stable expression of ΔN-RhoGDIβ in HeLa cells did not induce apoptosis, but impaired directional cell migration in a wound-healing assay accompanied by a perturbed direction of cell division at the wound edge. Subcellular protein fractionation experiments revealed that ΔN-RhoGDIβ but not wild-type RhoGDIβ was present in the detergent-soluble cytoplasmic and nuclear fractions and preferentially associated with Cdc42. Furthermore, Cdc42 activity was constitutively inhibited by stable expression of ΔN-RhoGDIβ, resulting in increased radiation-induced compensatory proliferation linking to RhoA activation. Thus, ΔN-RhoGDIβ dominant-negatively regulates Cdc42 activity and contributes to loss of polarity-related functions. The caspase-3-cleaved RhoGDIβ is a possible determinant to promote cancer spreading due to deregulation of directional organization of tumor cell population and inhibition of default equilibrium between proliferation and apoptosis after cytotoxic damage. J. Cell. Physiol. 231: 2493-2505, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  4. Intravenous apoptotic cell infusion as a cell-based therapy toward improving hematopoietic cell transplantation outcome.

    PubMed

    Saas, Philippe; Gaugler, Béatrice; Perruche, Sylvain

    2010-10-01

    Allogeneic hematopoietic cell transplantation (AHCT) is an efficient therapy for different malignant and nonmalignant hematological diseases. However, the use of this therapeutic approach is still limited by some severe toxic side effects, mainly graft-versus-host disease (GvHD). Today, the risk of fatal GvHD restrains the wider application of AHCT to many patients in need of an effective therapy for their high-risk hematologic malignancies. Thus, new strategies, including cell-based therapy approaches, are required. We propose to use intravenous donor apoptotic leukocyte infusion to improve AHCT outcome. In experimental AHCT models, we demonstrated that intravenous apoptotic leukocyte infusion, simultaneously with allogeneic bone marrow grafts, favors hematopoietic engraftment, prevents allo-immunization, and delays acute GvHD onset. Here, we review the different mechanisms and the potential beneficial effects associated with the immunomodulatory properties of apoptotic cells in the AHCT setting. © 2010 New York Academy of Sciences.

  5. Cell culture on polymers prepared by radiation-induced grafting of various monomers.

    PubMed

    Yoshii, F; Kaetsu, I

    1983-12-01

    The adhesion and growth of tissue cells on polymers prepared by radiation grafting was investigated. The apparent rates of initial attachment and growth of Chang liver and C6 cells were promoted on surfaces with increased wettability and with a heterogeneous structure for grafted polyvinyl fluoride film. The degree of cell attachment and growth on surfaces having a dense microblock structure, formed by grafting of methyl methacrylate in acetone solvent, was greater than that caused by other factors, such as wettability.

  6. Radiation-Induced Immunogenic Modulation Enhances T-Cell Killing | Center for Cancer Research

    Cancer.gov

    For many types of cancer, including breast, lung, and prostate carcinomas, radiation therapy is the standard of care. However, limits placed on the tolerable levels of radiation exposure coupled with heterogeneity of biological tissue result in cases where not all tumor cells receive a lethal dose of radiation. Preclinical studies have shown that exposing tumor cells to lethal doses of radiation can elicit cell death while inducing some antitumor immunity, described as immunogenic cell death (ICD). However, in a clinical setting, immune responses elicited by radiation alone rarely result in protective immunity, as tumor relapse often occurs.

  7. IGFBP3 and BAG1 enhance radiation-induced apoptosis in squamous esophageal cancer cells

    SciTech Connect

    Yoshino, Kei; Motoyama, Satoru; Koyota, Souichi; Shibuya, Kaori; Usami, Shuetsu; Maruyama, Kiyotomi; Saito, Hajime; Minamiya, Yoshihiro; Sugiyama, Toshihiro; Ogawa, Jun-ichi

    2011-01-28

    Research highlights: {yields} TE-12 cell had greater radiosensitivity and higher levels of caspase 3/7 activity for radiotherapy than TE-5 or TE-9 cells. {yields} The expression of IGFBP3 and BAG1 was five or more times higher in TE-12 cell in DNA microarrays analysis. {yields} Knocking down IGFBP3 and/or BAG1 expression using targeted siRNA diminished their susceptibility to radiation. -- Abstract: Identification of reliable markers of radiosensitivity and the key molecules that enhance the susceptibility of esophageal cancer cells to anticancer treatments would be highly desirable. To identify molecules that confer radiosensitivity to esophageal squamous carcinoma cells, we assessed the radiosensitivities of the TE-5, TE-9 and TE-12 cloneA1 cell lines. TE-12 cloneA1 cells showed significantly greater susceptibility to radiotherapy at 5 and 10 Gy than either TE-5 or TE-9 cells. Consistent with that finding, 24 h after irradiation (5 Gy), TE-12 cloneA1 cells showed higher levels of caspase 3/7 activity than TE-5 or TE-9 cells. When we used DNA microarrays to compare the gene expression profiles of TE-5 and TE-12 cloneA1 cells, we found that the mRNA and protein expression of insulin-like growth factor binding protein 3 (IGFBP3) and Bcl-2-associated athanogene 1 (BAG1) was five or more times higher in TE-12 cloneA1 cells than TE-5 cells. Conversely, knocking down expression of IGFBP3 and BAG1 mRNA in TE-12 cloneA1 cells using small interfering RNA (siRNA) significantly reduced radiosensitivity. These data suggest that IGFBP3 and BAG1 may be key markers of radiosensitivity that enhance the susceptibility of squamous cell esophageal cancer to radiotherapy. IGFBP3 and BAG1 may thus be useful targets for improved and more individualized treatments for patients with esophageal squamous cell carcinoma.

  8. Intestinal and peri-tumoral lymphatic endothelial cells are resistant to radiation-induced apoptosis

    SciTech Connect

    Sung, Hoon Ki; Morisada, Tohru; Cho, Chung-Hyun; Oike, Yuichi; Lee, Jayhun; Sung, Eon Ki; Chung, Jae Hoon; Suda, Toshio; Koh, Gou Young . E-mail: gykoh@kaist.ac.kr

    2006-06-30

    Radiation therapy is a widely used cancer treatment, but it is unable to completely block cancer metastasis. The lymphatic vasculature serves as the primary route for metastatic spread, but little is known about how lymphatic endothelial cells respond to radiation. Here, we show that lymphatic endothelial cells in the small intestine and peri-tumor areas are highly resistant to radiation injury, while blood vessel endothelial cells in the small intestine are relatively sensitive. Our results suggest the need for alternative therapeutic modalities that can block lymphatic endothelial cell survival, and thus disrupt the integrity of lymphatic vessels in peri-tumor areas.

  9. Comparative studies on radiation-induced micronuclei and chromosomal aberrations in V79 cells

    SciTech Connect

    Keshava, C.; Ong., T. |; Nath, J.

    1994-12-31

    Induction of micronuclei (MN) and structural chromosomal aberrations (SCA) by physical agents extensively in a variety of cell lines for genotoxicity assessment. However, comparative data on the relationship between these two cytogenetic endpoints are limited. This study compares MN and SCA formation in V79 Chinese hamster lung cells treated with X-rays and UV radiation. Four replicate cultures of exponentially growing cells were exposed to four doses of X-rays (100 to 800 rad). For two replicate cultures, cytochalasin B (3 {mu}g/ml) was added and cells harvested 16 h later for MN and cell cycle kinetics assessments. For the remaining two replicate cultures, colcemid (0.025{mu}g/ml) was added 16 h post-treatment and harvested 2 h later for SCA and mitotic index (MI) analysis. This experiment was duplicated using four doses of UV radiation (100 to 800 {mu}joules x 10{sup 2}/cm{sup 2}). In the x-ray experiment, a dose-related decrease in the % of binucleated (BN) cells and MI was noted. Also, there was a clear dose-related increase in micronucleated binucleate (MNBN) and aberrant cells. Similar dose-response, but with lower frequencies, was observed in the UV radiation treatment. These data suggest that there is a good relationship between chromosome damage as measured by the % of MNBN and aberrant cells and cytotoxicity as measured by the % of BN cells and MI in these assays.

  10. Cell-Assisted Lipotransfer Improves Volume Retention in Irradiated Recipient Sites and Rescues Radiation-Induced Skin Changes

    PubMed Central

    Luan, Anna; Duscher, Dominik; Whittam, Alexander J.; Paik, Kevin J.; Zielins, Elizabeth R.; Brett, Elizabeth A.; Atashroo, David A.; Hu, Michael S.; Lee, Gordon K.; Gurtner, Geoffrey C.; Longaker, Michael T.; Wan, Derrick C.

    2016-01-01

    Radiation therapy is not only a mainstay in the treatment of many malignancies but also results in collateral obliteration of microvasculature and dermal/subcutaneous fibrosis. Soft tissue reconstruction of hypovascular, irradiated recipient sites through fat grafting remains challenging; however, a coincident improvement in surrounding skin quality has been noted. Cell-assisted lipotransfer (CAL), the enrichment of fat with additional adipose-derived stem cells (ASCs) from the stromal vascular fraction, has been shown to improve fat volume retention, and enhanced outcomes may also be achieved with CAL at irradiated sites. Supplementing fat grafts with additional ASCs may also augment the regenerative effect on radiation-damaged skin. In this study, we demonstrate the ability for CAL to enhance fat graft volume retention when placed beneath the irradiated scalps of immunocompromised mice. Histologic metrics of fat graft survival were also appreciated, with improved structural qualities and vascularity. Finally, rehabilitation of radiation-induced soft tissue changes were also noted, as enhanced amelioration of dermal thickness, collagen content, skin vascularity, and biomechanical measures were all observed with CAL compared to unsupplemented fat grafts. Supplementation of fat grafts with ASCs therefore shows promise for reconstruction of complex soft tissue defects following adjuvant radiotherapy. PMID:26661694

  11. Progressive behavioral changes during the maturation of rats with early radiation-induced hypoplasia of fascia dentata granule cells

    SciTech Connect

    Mickley, G.A.; Ferguson, J.L.; Mulvihill, M.A.; Nemeth, T.J. )

    1989-07-01

    Localized exposure of the neonatal rat brain to X-rays produces neuronal hypoplasia specific to the granule cell layer of the hippocampal dentate gyrus. This brain damage causes locomotor hyperactivity, slowed acquisition of passive avoidance tasks and long bouts of spontaneous turning (without reversals) in a bowl apparatus. Here we report how these behavioral deficits change as a function of subject aging and behavioral test replications. Portions of the neonatal rat cerebral hemispheres were X-irradiated in order to selectively damage the granule cells of the dentate gyrus. The brains of experimental animals received a fractionated dose of X rays (13 Gy total) over postnatal days 1 to 16 and control animals were sham-irradiated. Rats between the ages of 71-462 days were tested 3 separate times on each of the following 3 behavioral tests: (1) spontaneous locomotion, (2) passive avoidance acquisition, and (3) spontaneous circling in a large plastic hemisphere. Rats with radiation-induced damage to the fascia dentata exhibited long bouts of slow turns without reversals. Once they began, irradiated subjects perseverated in turning to an extent significantly greater than sham-irradiated control subjects. This irradiation effect was significant during all test series. Moreover, in time, spontaneous perseverative turning was significantly potentiated in rats with hippocampal damage but increased only slightly in controls. Early radiation exposure produced locomotor hyperactivity in young rats. While activity levels of controls remained fairly stable throughout the course of the experiment, the hyperactivity of the irradiated animals decreased significantly as they matured.

  12. Nano-silymarin provides protection against γ-radiation-induced oxidative stress in cultured human embryonic kidney cells.

    PubMed

    Adhikari, Manish; Arora, Rajesh

    2015-10-01

    Radiation can produce biological damage, mainly oxidative stress, via production of free radicals, including reactive oxygen species (ROS). Nanoparticles are of interest as radioprotective agents, particularly due to their high solubility and bioavailability. Silymarin is a hepatoprotective agent but has poor oral bioavailability. Silymarin was formulated as a nanoemulsion with the aim of improving its bioavailability and therapeutic efficacy. In the present study, we evaluated self-nanoemulsifying drug delivery systems (SNEDDS) formulated with surfactants and co-surfactants. Nano-silymarin was characterized by estimating % transmittance, globule size, and polydispersity index, and by transmission electron microscopy (TEM). The nano-silymarin obtained was in the range of 3-8nm diameter. With regard to DNA damage, measured by a plasmid relaxation assay, maximum protection was obtained at 10μg/mL. Cytotoxicity of nano-silymarin to human embryonic kidney (HEK) cells was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. Protective efficacy against γ-radiation was assessed by reduction in micronucleus frequency and ROS generation, using the 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) assay. Radiation-induced apoptosis was estimated by microscopic analysis and cell-cycle estimation. Nano-silymarin was radioprotective, supporting the possibility of developing new approaches to radiation protection via nanotechnology. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. High-LET Radiation Induced Chromosome Aberrations in Normal and Ataxia Telangiectasia Fibroblast Cells

    NASA Astrophysics Data System (ADS)

    Kawata, Tetsuya; George, Ms Kerry; Cucinotta, Francis A.; Shigematsu, Naoyuki; Ito, Hisao; Furusawa, Yoshiya; Uno, Takashi

    We investigated the effects of heavy ions beams on chromosomal aberrations in normal and AT cells. Normal and AT fibroblast cells arrested at G0/G1 phase were irradiated with 2 Gy of X-rays, 490 MeV/u Silicon (LET 55 keV/micron), 500 MeV/u Iron (LET 185 keV/micron) and 200 MeV/u Iron (LET 440 keV/micron) particles, and then cells were allowed to repair for 24 hours at 37 degrees before subculture. Calyculin-A induced PCC method was employed to collect G2/M chromosomes and whole DNA probes 1 and 3 were used to analyze chromosomal aberrations such as color-junctions, deletions, simple exchanges (incomplete and reciprocal exchanges) and complex-type exchanges. The percentages of aberrant cells were higher when normal and AT cells were exposed to heavy ions compared to X-rays, and had a tendency to increase with increasing LET up to 185 keV/micron and then decreased at 440 keV/micron. When the frequency of color-junctions per cell was compared after X-ray exposure, AT cells had around three times higher frequency of color-junctions (mis-rejoining) than normal cells. However, at 185 keV/micron there was no difference in the frequency of color-junctions between two cell lines. It was also found that the frequency of simple exchanges per cell was almost constant in AT cells regardless LET levels, but it was LET dependent for normal cells. Interestingly, the frequency of simple exchanges was higher for normal fibroblast cells when it was compared at 185 keV/micron, but AT cells had more complex-type exchanges at the same LET levels. Heavy ions are more efficient in inducing chromosome aberrations in normal and AT cells compared to X-rays, and the aberration types between normal and AT fibroblast appeared different probably due to difference in the ATM gene function.

  14. Radiation-Induced Bystander Effects in A549 Cells Exposed to 6 MV X-rays.

    PubMed

    Yang, Shuning; Xu, Jing; Shao, Weixian; Geng, Chong; Li, Jia; Guo, Feng; Miao, Hui; Shen, Wenbin; Ye, Tao; Liu, Yazhou; Xu, Haiting; Zhang, Xuguang

    2015-07-01

    The aim of the study is to explore the bystander effects in A549 cells that have been exposed to 6MV X-ray. Control group, irradiated group, irradiated conditioned medium (ICM)-received group, and fresh medium group were designed in this study. A549 cells in the logarithmic growth phase were irradiated with 6MV X-ray at 0, 0.5, 1, 1.5, and 2. In ICM-received group, post-irradiation A549 cells were cultured for 3 h and were transferred into non-irradiated A549 cells for further cultivation. Clone forming test was applied to detect the survival fraction of cells. Annexin V-FITC/PI double-staining assay was used to detect the apoptosis of A549 cells 24, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation, and the curves of apoptosis were drawn. The changes in the cell cycles 4, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation were detected using PI staining flow cytometry. With the increase of irradiation dose, the survival fraction of A549 cells after the application of 0.5 Gy irradiation was decreasing continuously. In comparison to the control group, the apoptosis rate of the ICM-received group was increased in a time-dependent pattern, with the highest apoptosis rate observed at 72 h (p < 0.05). Cell count in G2/M stages was obviously increased compared with that of the control group (p < 0.05), with the highest count observed at 72 h, after which G2/M stage arrest was diminished. ICM can cause apparent A549 cell damage, indicating that 6MV X-ray irradiation can induce bystander effect on A549 cells, which reaches a peak at 72 h.

  15. Mitochondrial Staining Allows Robust Elimination of Apoptotic and Damaged Cells during Cell Sorting

    PubMed Central

    Ponomarev, Eugeny D.; Tsytsykova, Alla; Armant, Myriam; Vorobjev, Ivan A.

    2014-01-01

    High-speed fluorescence-activated cell sorting is relevant for a plethora of applications, such as PCR-based techniques, microarrays, cloning, and propagation of selected cell populations. We suggest a simple cell-sorting technique to eliminate early and late apoptotic and necrotic cells, with good signal-to-noise ratio and a high-purity yield. The mitochondrial potential dye, TMRE (tetramethylrhodamine ethyl ester perchlorate), was used to separate viable and non-apoptotic cells from the cell sorting samples. TMRE staining is reversible and does not affect cell proliferation and viability. Sorted TMRE+ cells contained a negligible percentage of apoptotic and damaged cells and had a higher proliferative potential as compared with their counterpart cells, sorted on the basis of staining with DNA viability dye. This novel sorting technique using TMRE does not interfere with subsequent functional assays and is a method of choice for the enrichment of functionally active, unbiased cell populations. PMID:24394470

  16. Mitochondrial staining allows robust elimination of apoptotic and damaged cells during cell sorting.

    PubMed

    Barteneva, Natasha S; Ponomarev, Eugeny D; Tsytsykova, Alla; Armant, Myriam; Vorobjev, Ivan A

    2014-04-01

    High-speed fluorescence-activated cell sorting is relevant for a plethora of applications, such as PCR-based techniques, microarrays, cloning, and propagation of selected cell populations. We suggest a simple cell-sorting technique to eliminate early and late apoptotic and necrotic cells, with good signal-to-noise ratio and a high-purity yield. The mitochondrial potential dye, TMRE (tetramethylrhodamine ethyl ester perchlorate), was used to separate viable and non-apoptotic cells from the cell sorting samples. TMRE staining is reversible and does not affect cell proliferation and viability. Sorted TMRE(+) cells contained a negligible percentage of apoptotic and damaged cells and had a higher proliferative potential as compared with their counterpart cells, sorted on the basis of staining with DNA viability dye. This novel sorting technique using TMRE does not interfere with subsequent functional assays and is a method of choice for the enrichment of functionally active, unbiased cell populations.

  17. Derivation of hepatocytes from bone marrow cells in mice after radiation-induced myeloablation.

    PubMed

    Theise, N D; Badve, S; Saxena, R; Henegariu, O; Sell, S; Crawford, J M; Krause, D S

    2000-01-01

    Following a report of skeletal muscle regeneration from bone marrow cells, we investigated whether hepatocytes could also derive in vivo from bone marrow cells. A cohort of lethally irradiated B6D2F1 female mice received whole bone marrow transplants from age-matched male donors and were sacrificed at days 1, 3, 5, and 7 and months 2, 4, and 6 posttransplantation (n = 3 for each time point). Additionally, 2 archival female mice of the same strain who had previously been recipients of 200 male fluorescence-activated cell sorter (FACS)-sorted CD34(+)lin(-) cells were sacrificed 8 months posttransplantation under the same protocol. Fluorescence in situ hybridization (FISH) for the Y-chromosome was performed on liver tissue. Y-positive hepatocytes, up to 2.2% of total hepatocytes, were identified in 1 animal at 7 days posttransplantation and in all animals sacrificed 2 months or longer posttransplantation. Simultaneous FISH for the Y-chromosome and albumin messenger RNA (mRNA) confirmed male-derived cells were mature hepatocytes. These animals had received lethal doses of irradiation at the time of bone marrow transplantation, but this induced no overt, histologically demonstrable, acute hepatic injury, including inflammation, necrosis, oval cell proliferation, or scarring. We conclude that hepatocytes can derive from bone marrow cells after irradiation in the absence of severe acute injury. Also, the small subpopulation of CD34(+)lin(-) bone marrow cells is capable of such hepatic engraftment.

  18. Susceptibility of BRCA2 Heterozygous Normal Mammary Epithelial Cells to Radiation-Induced Transformation

    DTIC Science & Technology

    2005-10-01

    mouse embryonic fibroblasts (MEF) with less severe BRCA2 truncations (22-24). Particularly, spontaneous accumulation of chromosomal abnormalities ...Scully, R. Stable interaction between the products of the BRCA1 and BRCA2 tumor suppressor genes in mitotic and meiotic cells, Mol Cell. 2: 317-28

  19. Mast Cell Growth Factor Enhances Multilineage Hematopoietic Recovery in Vivo Following Radiation-Induced Aplasia

    DTIC Science & Technology

    1994-01-01

    GM-CFC), and peripheral white blood cells macrocytic anemia that is resistant to erythropoietin treat- (WBC), red 1 lood cells (RBC), and platelets...MacVittie TJ (1993) A GM-CSF/IL-3 fusion protein tary mouse anemias (SI/Sid and W/WV) deficient in promotes neutrophil and platelet recovery in

  20. Effect of caffeine on radiation-induced apoptosis in TK6 cells

    SciTech Connect

    Zhen, W.; Vaughan, A.T.M.

    1995-02-01

    Apoptosis has been measured in cells of the human TK6 lymphoblastoid cell line by recording the release of endonuclease-digested DNA from affected cells using flow cytometry. In asynchronously dividing cells, DNA degradation characteristic of apoptosis was first seen 12 h after irradiation as a defined DNA fluorescent peak of sub-G{sub 1}-phase content, reaching a maximum of 30-50% of the population by 24-72 h. Treating cells with 2 mM caffeine either before or up to 3 h after irradiation eliminated the degradation of DNA entirely. In addition, the percentage of cells in which apoptosis could be detected microscopically decreased from 62.4 {+-} 0.95% to 16.7 {+-} 1.5% 72 h after caffeine treatment. Delaying caffeine treatment for 12 h after irradiation reduced DNA degradation by approximately 50% compared to cells receiving radiation alone. DNA degradation induced by serum deprivation was unaffected by caffeine treatment. These data support the contention that irradiation of TK6 cells produces a long-lived cellular signal which triggers apoptosis. Apoptosis produced by serum deprivation does not operate through the same pathway. 36 refs., 5 figs.

  1. Nonimmunogenic radiation-induced lymphoma: immunity induction by a somatic cell hybrid

    SciTech Connect

    Yefenof, E.; Goldapfel, M.; Ber, R.

    1982-05-01

    The cell line designated PIR-2 is a nonimmunogenic X-ray-induced thymoma of C57BL/6 origin that is unable to induce antitumor immunity in syngeneic lymphocytes in vitro and in mice in vivo. Fusion of PIR-2 with an allogeneic universal fuser A9HT (clone 3c) resulted in the establishment of a somatic cell hybrid designated A9/PIR. C57BL/6 lymphocytes sensitized in vitro with A9/PIR could lyse parental PIR-2 cells, as well as other syngeneic tumors. However, immunization of mice with the hybrid significantly enhanced PIR-2 tumor takes while it partially protected the animals against a challenge with unrelated syngeneic tumors. The results imply that somatic cell hybridization can increase the immunogenicity of an otherwise nonimmunogenic tumor. However, in view of the enhancing effects of hybrid preimmunization on parental tumor cell growth, the possible application of this approach for immunotherapy is questionable.

  2. Radiation-Induced Glycogen Accumulation Detected by Single Cell Raman Spectroscopy Is Associated with Radioresistance that Can Be Reversed by Metformin

    PubMed Central

    Matthews, Quinn; Isabelle, Martin; Harder, Samantha J.; Smazynski, Julian; Beckham, Wayne; Brolo, Alexandre G.; Jirasek, Andrew; Lum, Julian J.

    2015-01-01

    Altered cellular metabolism is a hallmark of tumor cells and contributes to a host of properties associated with resistance to radiotherapy. Detection of radiation-induced biochemical changes can reveal unique metabolic pathways affecting radiosensitivity that may serve as attractive therapeutic targets. Using clinically relevant doses of radiation, we performed label-free single cell Raman spectroscopy on a series of human cancer cell lines and detected radiation-induced accumulation of intracellular glycogen. The increase in glycogen post-irradiation was highest in lung (H460) and breast (MCF7) tumor cells compared to prostate (LNCaP) tumor cells. In response to radiation, the appearance of this glycogen signature correlated with radiation resistance. Moreover, the buildup of glycogen was linked to the phosphorylation of GSK-3β, a canonical modulator of cell survival following radiation exposure and a key regulator of glycogen metabolism. When MCF7 cells were irradiated in the presence of the anti-diabetic drug metformin, there was a significant decrease in the amount of radiation-induced glycogen. The suppression of glycogen by metformin following radiation was associated with increased radiosensitivity. In contrast to MCF7 cells, metformin had minimal effects on both the level of glycogen in H460 cells following radiation and radiosensitivity. Our data demonstrate a novel approach of spectral monitoring by Raman spectroscopy to assess changes in the levels of intracellular glycogen as a potential marker and resistance mechanism to radiation therapy. PMID:26280348

  3. Ionizing Radiation-Induced DNA Damage and Its Repair in Human Cells

    SciTech Connect

    Dizdaroglu, Miral

    1999-05-12

    DNA damage in mammalian chromatin in vitro and in cultured mammalian cells including human cells was studied. In the first phase of these studies, a cell culture laboratory was established. Necessary equipment including an incubator, a sterile laminar flow hood and several centrifuges was purchased. We have successfully grown several cell lines such as murine hybridoma cells, V79 cells and human K562 leukemia cells. This was followed by the establishment of a methodology for the isolation of chromatin from cells. This was a very important step, because a routine and successful isolation of chromatin was a prerequisite for the success of the further studies in this project, the aim of which was the measurement of DNA darnage in mammalian chromatin in vitro and in cultured cells. Chromatin isolation was accomplished using a slightly modified procedure of the one described by Mee & Adelstein (1981). For identification and quantitation of DNA damage in cells, analysis of chromatin was preferred over the analysis of "naked DNA" for the following reasons: i. DNA may not be extracted efficiently from nucleoprotein in exposed cells, due to formation of DNA-protein cross-links, ii. the extractability of DNA is well known to decrease with increasing doses of radiation, iii. portions of DNA may not be extracted due to fragmentation, iv. unextracted DNA may contain a significant portion of damaged DNA bases and DNA-protein cross-links. The technique of gas chromatography/mass spectrometry (GC/MS), which was used in the present project, permits the identification and quantitation of modified DNA bases in chromatin in the presence of proteins without the necessity of first isolating DNA from chromatin. This has been demonstrated previously by the results from our laboratory and by the results obtained during the course of the present project. The quality of isolated chromatin was tested by measurement of its content of DNA, proteins, and RNA, by analysis of its protein

  4. Clinical significance of radiation-induced CD133 expression in residual rectal cancer cells after chemoradiotherapy.

    PubMed

    Kawamoto, Aya; Tanaka, Koji; Saigusa, Susumu; Toiyama, Yuji; Morimoto, Yuhki; Fujikawa, Hiroyuki; Iwata, Takashi; Matsushita, Kohei; Yokoe, Takeshi; Yasuda, Hiromi; Inoue, Yasuhiro; Miki, Chikao; Kusunoki, Masato

    2012-03-01

    CD133 and CD44 have been considered as markers for colorectal cancer stem cells (CSCs). The association of CD133 and CD44 expression with radiation has not been fully examined in rectal cancer. Both CD133 (PROM) and CD44 mRNA levels were measured in post-chemoradiotherapy (CRT) specimens of 52 rectal cancer patients using real-time RT-PCR and compared to clinicopathological variables and clinical outcome. Their protein levels were examined in the radiation-treated HT29 human colon cancer cell line. Post-CRT CD133 in residual cancer cells was significantly higher than matched pre-CRT CD133 in biopsy specimens (n=30). By contrast, CD44 was significantly lower in post-CRT specimens (P<0.01). CD133 was associated with distant recurrence after CRT followed by surgery (P<0.05). Patients with elevated CD133 in residual cancer cells showed poor disease-free survival (P<0.05). No significant association between post-CRT CD44 and clinical outcome was found. The in vitro study showed that CD133 protein was increased in a radiation dose-dependent manner, despite of the decreased number of clonogenic radiation-surviving cells. CD44 protein was decreased after irradiation. CD133, but not CD44, was increased in radiation-resistant surviving colon cancer cells. Post-CRT CD133 in residual cancer cells may predict metachronous distant recurrence and poor survival of rectal cancer patients after CRT.

  5. Optical imaging of radiation-induced metabolic changes in radiation-sensitive and resistant cancer cells

    NASA Astrophysics Data System (ADS)

    Alhallak, Kinan; Jenkins, Samir V.; Lee, David E.; Greene, Nicholas P.; Quinn, Kyle P.; Griffin, Robert J.; Dings, Ruud P. M.; Rajaram, Narasimhan

    2017-06-01

    Radiation resistance remains a significant problem for cancer patients, especially due to the time required to definitively determine treatment outcome. For fractionated radiation therapy, nearly 7 to 8 weeks can elapse before a tumor is deemed to be radiation-resistant. We used the optical redox ratio of FAD/(FAD+NADH) to identify early metabolic changes in radiation-resistant lung cancer cells. These radiation-resistant human A549 lung cancer cells were developed by exposing the parental A549 cells to repeated doses of radiation (2 Gy). Although there were no significant differences in the optical redox ratio between the parental and resistant cell lines prior to radiation, there was a significant decrease in the optical redox ratio of the radiation-resistant cells 24 h after a single radiation exposure (p=0.01). This change in the redox ratio was indicative of increased catabolism of glucose in the resistant cells after radiation and was associated with significantly greater protein content of hypoxia-inducible factor 1 (HIF-1α), a key promoter of glycolytic metabolism. Our results demonstrate that the optical redox ratio could provide a rapid method of determining radiation resistance status based on early metabolic changes in cancer cells.

  6. [Radiation-induces increased tumor cell aggressiveness of tumors of the glioblastomas?].

    PubMed

    Falk, Alexander T; Moncharmont, Coralie; Guilbert, Matthieu; Guy, Jean-Baptiste; Alphonse, Gersende; Trone, Jane-Chloé; Rivoirard, Romain; Gilormini, Marion; Toillon, Robert-Alain; Rodriguez-Lafrasse, Claire; Magné, Nicolas

    2014-09-01

    Glioblastoma multiform is the most common and aggressive brain tumor with a worse prognostic. Ionizing radiation is a cornerstone in the treatment of glioblastome with chemo-radiation association being the actual standard. As a paradoxal effect, it has been suggested that radiotherapy could have a deleterious effect on local recurrence of cancer. In vivo studies have studied the effect of radiotherapy on biological modification and pathogenous effect of cancer cells. It seems that ionizing radiations with photon could activate oncogenic pathways in glioblastoma cell lines. We realized a review of the literature of photon-enhanced effect on invasion and migration of glioblastoma cells by radiotherapy.

  7. Apoptotic effect of noscapine in breast cancer cell lines.

    PubMed

    Quisbert-Valenzuela, Edwin O; Calaf, Gloria M

    2016-06-01

    Cancer is a public health problem in the world and breast cancer is the most frequently cancer in women. Approximately 15% of the breast cancers are triple-negative. Apoptosis regulates normal growth, homeostasis, development, embryogenesis and appropriate strategy to treat cancer. Bax is a protein pro-apoptotic enhancer of apoptosis in contrast to Bcl-2 with antiapoptotic properties. Initiator caspase-9 and caspase-8 are features of intrinsic and extrinsic apoptosis pathway, respectively. NF-κB is a transcription factor known to be involved in the initiation and progression of breast cancer. Noscapine, an alkaloid derived from opium is used as antitussive and showed antitumor properties that induced apoptosis in cancer cell lines. The aim of the present study was to determine the apoptotic effect of noscapine in breast cancer cell lines compared to breast normal cell line. Three cell lines were used: i) a control breast cell line MCF-10F; ii) a luminal-like adenocarcinoma triple-positive breast cell line MCF-7; iii) breast cancer triple-negative cell line MDA-MB-231. Our results showed that noscapine had lower toxicity in normal cells and was an effective anticancer agent that induced apoptosis in breast cancer cells because it increases Bax gene and protein expression in three cell lines, while decreases Bcl-xL gene expression, and Bcl-2 protein expression decreased in breast cancer cell lines. Therefore, Bax/Bcl-2 ratio increased in the three cell lines. This drug increased caspase-9 gene expression in breast cancer cell lines and caspase-8 gene expression increased in MCF-10F and MDA-MB-231. Furthermore, it increased cleavage of caspase-8, suggesting that noscapine-induced apoptosis is probably due to the involvement of extrinsic and intrinsic apoptosis pathways. Antiapoptotic gene and protein expression diminished and proapoptotic gene and protein expression increased noscapine-induced expression, probably due to decrease in NF-κB gene and protein expression

  8. Radiation-induced bystander effect and adaptive response in mammalian cells

    NASA Technical Reports Server (NTRS)

    Zhou, H.; Randers-Pehrson, G.; Waldren, C. A.; Hei, T. K.

    2004-01-01

    Two conflicting phenomena, bystander effect and adaptive response, are important in determining the biological responses at low doses of radiation and have the potential to impact the shape of the dose-response relationship. Using the Columbia University charged-particle microbeam and the highly sensitive AL cell mutagenic assay, we show here that non-irradiated cells acquire mutagenesis through direct contact with cells whose nuclei have been traversed with a single alpha particle each. Pretreatment of cells with a low dose of X-rays four hours before alpha particle irradiation significantly decreased this bystander mutagenic response. Results from the present study address some of the fundamental issues regarding both the actual target and radiation dose effect and can contribute to our current understanding in radiation risk assessment. c2004 COSPAR. Published by Elsevier Ltd. All rights reserved.

  9. Radiation-induced bystander effect and adaptive response in mammalian cells

    NASA Technical Reports Server (NTRS)

    Zhou, H.; Randers-Pehrson, G.; Waldren, C. A.; Hei, T. K.

    2004-01-01

    Two conflicting phenomena, bystander effect and adaptive response, are important in determining the biological responses at low doses of radiation and have the potential to impact the shape of the dose-response relationship. Using the Columbia University charged-particle microbeam and the highly sensitive AL cell mutagenic assay, we show here that non-irradiated cells acquire mutagenesis through direct contact with cells whose nuclei have been traversed with a single alpha particle each. Pretreatment of cells with a low dose of X-rays four hours before alpha particle irradiation significantly decreased this bystander mutagenic response. Results from the present study address some of the fundamental issues regarding both the actual target and radiation dose effect and can contribute to our current understanding in radiation risk assessment. c2004 COSPAR. Published by Elsevier Ltd. All rights reserved.

  10. Pharmacologic profiling of phosphoinositide 3-kinase inhibitors as mitigators of ionizing radiation-induced cell death.

    PubMed

    Lazo, John S; Sharlow, Elizabeth R; Epperly, Michael W; Lira, Ana; Leimgruber, Stephanie; Skoda, Erin M; Wipf, Peter; Greenberger, Joel S

    2013-12-01

    Ionizing radiation (IR) induces genotoxic stress that triggers adaptive cellular responses, such as activation of the phosphoinositide 3-kinase (PI3K)/Akt signaling cascade. Pluripotent cells are the most important population affected by IR because they are required for cellular replenishment. Despite the clear danger to large population centers, we still lack safe and effective therapies to abrogate the life-threatening effects of any accidental or intentional IR exposure. Therefore, we computationally analyzed the chemical structural similarity of previously published small molecules that, when given after IR, mitigate cell death and found a chemical cluster that was populated with PI3K inhibitors. Subsequently, we evaluated structurally diverse PI3K inhibitors. It is remarkable that 9 of 14 PI3K inhibitors mitigated γIR-induced death in pluripotent NCCIT cells as measured by caspase 3/7 activation. A single intraperitoneal dose of LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one], administered to mice at 4 or 24 hours, or PX-867 [(4S,4aR,5R,6aS,9aR,Z)-11-hydroxy-4-(methoxymethyl)-4a,6a-dimethyl-2,7,10-trioxo-1-(pyrrolidin-1-ylmethylene)-1,2,4,4a,5,6,6a,7,8,9,9a,10-dodecahydroindeno[4,5-H]isochromen-5-yl acetate (CID24798773)], administered 4 hours after a lethal dose of γIR, statistically significantly (P < 0.02) enhanced in vivo survival. Because cell cycle checkpoints are important regulators of cell survival after IR, we examined cell cycle distribution in NCCIT cells after γIR and PI3K inhibitor treatment. LY294002 and PX-867 treatment of nonirradiated cells produced a marked decrease in S phase cells with a concomitant increase in the G1 population. In irradiated cells, LY294002 and PX-867 treatment also decreased S phase and increased the G1 and G2 populations. Treatment with LY294002 or PX-867 decreased γIR-induced DNA damage as measured by γH2AX, suggesting reduced DNA damage. These results indicate pharmacologic inhibition of PI3K after

  11. Radiation-Induced Germ Cell Mutations-Their Detection and Modification.

    DTIC Science & Technology

    1987-06-30

    representative of cells irradiated as differentiating spermatogonia . No reduction in the fertilization * rate was detected in animals mated 100 or more days... spermatogonia . This damage is manifest only as a failure to differentiate and proliferate an inner cell mass atop the trophectoderm outgrowth. The nature of...Ford, C.E., Searle, A.G., Evans, E.P. and B.J. West. Differential transmission of translocation induced in spermatogonia of mice by irradiation

  12. Ionizing Radiation-Induced Responses in Human Cells with Differing TP53 Status

    PubMed Central

    Mirzayans, Razmik; Andrais, Bonnie; Scott, April; Wang, Ying W.; Murray, David

    2013-01-01

    Ionizing radiation triggers diverse responses in human cells encompassing apoptosis, necrosis, stress-induced premature senescence (SIPS), autophagy, and endopolyploidy (e.g., multinucleation). Most of these responses result in loss of colony-forming ability in the clonogenic survival assay. However, not all modes of so-called clonogenic cell “death” are necessarily advantageous for therapeutic outcome in cancer radiotherapy. For example, the crosstalk between SIPS and autophagy is considered to influence the capacity of the tumor cells to maintain a prolonged state of growth inhibition that unfortunately can be succeeded by tumor regrowth and disease recurrence. Likewise, endopolyploid giant cells are able to segregate into near diploid descendants that continue mitotic activities. Herein we review the current knowledge on the roles that the p53 and p21WAF1 tumor suppressors play in determining the fate of human fibroblasts (normal and Li-Fraumeni syndrome) and solid tumor-derived cells after exposure to ionizing radiation. In addition, we discuss the important role of WIP1, a p53-regulated oncogene, in the temporal regulation of the DNA damage response and its contribution to p53 dynamics post-irradiation. This article highlights the complexity of the DNA damage response and provides an impetus for rethinking the nature of cancer cell resistance to therapeutic agents. PMID:24232458

  13. X-ray radiation-induced effects in human mammary epithelial cells investigated by Raman microspectroscopy

    NASA Astrophysics Data System (ADS)

    Risi, R.; Manti, L.; Perna, G.; Lasalvia, M.; Capozzi, V.; Delfino, I.; Lepore, M.

    2012-06-01

    Micro-Raman technique can be particularly useful to investigate the chemical changes induced in structure, protein, nucleic acid, lipid, and carbohydrate contents of cells. The aim of this work is to inspect the possibility to employ Raman microspectroscopy to detect biochemical modifications in human mammary epithelial cells after exposure to different Xray doses. The samples consisted of cells cultured on polylysine-coated glass coverslips. After the exposition, control and treated cells were washed in phosphate-buffered saline (PBS) and then fixed in paraformaldehyde 3.7%. They were examined using a confocal micro-Raman system equipped with a He-Ne laser (λ = 632.8 nm; power on the sample= 3.5mW). Differences in the intensity ratio of specific Raman vibrational markers commonly assigned to phenylalanine and tyrosine amino acids (at 1000, 1030, 1618 cm-1), DNA bases (787, 1090, 1305 cm-1), and amide III (1237, and 1265 cm-1) with respect a reference peak (the one of lipids at 1450 cm-1) were evidenced between control and exposed cells. These differences may be indicative of damage in exposed cells as the fragmentation of individual amino acids and DNA bases, crosslink effects in molecular structure of DNA and protein conformational change that especially tend to "unwind" the protein due to the breaking of hydrogen bonds between peptide chains.

  14. Radiation induces premature chromatid separation via the miR-142-3p/Bod1 pathway in carcinoma cells

    PubMed Central

    Pan, Dong; Du, Yarong; Ren, Zhenxin; Chen, Yaxiong; Li, Xiaoman; Wang, Jufang; Hu, Burong

    2016-01-01

    Radiation-induced genomic instability plays a vital role in carcinogenesis. Bod1 is required for proper chromosome biorientation, and Bod1 depletion increases premature chromatid separation. MiR-142-3p influences cell cycle progression and inhibits proliferation and invasion in cervical carcinoma cells. We found that radiation induced premature chromatid separation and altered miR-142-3p and Bod1 expression in 786-O and A549 cells. Overexpression of miR-142-3p increased premature chromatid separation and G2/M cell cycle arrest in 786-O cells by suppressing Bod1 expression. We also found that either overexpression of miR-142-3p or knockdown of Bod1 sensitized 786-O and A549 cells to X-ray radiation. Overexpression of Bod1 inhibited radiation- and miR-142-3p-induced premature chromatid separation and increased resistance to radiation in 786-O and A549 cells. Taken together, these results suggest that radiation alters miR-142-3p and Bod1 expression in carcinoma cells, and thus contributes to early stages of radiation-induced genomic instability. Combining ionizing radiation with epigenetic regulation may help improve cancer therapies. PMID:27527863

  15. Ionizing radiation-induced cell death is partly caused by increase of mitochondrial reactive oxygen species in normal human fibroblast cells.

    PubMed

    Kobashigawa, Shinko; Kashino, Genro; Suzuki, Keiji; Yamashita, Shunichi; Mori, Hiromu

    2015-04-01

    Radiation-induced cell death is thought to be caused by nuclear DNA damage that cannot be repaired. However, in this study we found that a delayed increase of mitochondrial reactive oxygen species (ROS) is responsible for some of the radiation-induced cell death in normal human fibroblast cells. We have previously reported that there is a delayed increase of mitochondrial (·)O2(-), measured using MitoSOX™ Red reagent, due to gamma irradiation. This is dependent on Drp1 localization to mitochondria. Here, we show that knockdown of Drp1 expression reduces the level of DNA double-strand breaks (DSBs) remaining 3 days after 6 Gy irradiation. Furthermore, cells with knockdown of Drp1 expression are more resistant to gamma radiation. We then tested whether the delayed increase of ROS causes DNA damage. The antioxidant, 2-glucopyranoside ascorbic acid (AA-2G), was applied before or after irradiation to inhibit ROS production during irradiation or to inhibit delayed ROS production from mitochondria. Interestingly, 1 h after exposure, the AA-2G treatment reduced the level of DSBs remaining 3 days after 6 Gy irradiation. In addition, irradiated AA-2G-treated cells were more resistant to radiation than the untreated cells. These results indicate that delayed mitochondrial ROS production may cause some of the cell death after irradiation.

  16. Radiation-induced senescence-like terminal growth arrest in thyroid cells.

    PubMed

    Podtcheko, Alexei; Namba, Hiroyuki; Saenko, Vladimir; Ohtsuru, Akira; Starenki, Dmitriy; Meirmanov, Serik; Polona, Iryna; Rogounovitch, Tatiana; Yamashita, Shunichi

    2005-04-01

    Premature senescence may play an important role as an acute, drug-, or ionizing radiation (IR)-inducible growth arrest program along with interphase apoptosis and mitotic catastrophe. The aim of the study was to evaluate whether IR can induce senescence-like phenotype (SLP) associated with terminal growth arrest in the thyroid cells, and if so, to evaluate impact of terminal growth arrest associated with SLP in intrinsic radiosensitivity of various thyroid carcinomas. The induction of SLP in thyroid cells were identified by: (1) senescence associated beta-galactosidase (SA-beta-Gal) staining method, (2) dual-flow cytometric analysis of cell proliferation and side light scatter using vital staining with PKH-2 fluorescent dye, (3) double labeling for 5-bromodeoxyuridine and SA- beta-Gal, (4) Staining for SA-beta-Gal with consequent antithyroglobulin immunohistochemistry. IR induced SLP associated with terminal growth arrest in four thyroid cancer cells lines and in primary thyrocytes in time- and dose-dependent manner. Analysis of relationship between induction of SLP and radiosensitivity revealed a trend in which more radioresistant cell lines strongly tended to show lower specific SLP yields (r = -0.93, p = 0.068). We find out that SA-beta-Gal staining is detectable in irradiated ARO xenotransplants, but not in control tumors. We, therefore, conclude that induction of SLP with terminal growth arrest contribute to the elimination of clonogenic populations after IR.

  17. Radiation-induced Akt activation modulates radioresistance in human glioblastoma cells

    PubMed Central

    Li, Hui-Fang; Kim, Jung-Sik; Waldman, Todd

    2009-01-01

    Background Ionizing radiation (IR) therapy is a primary treatment for glioblastoma multiforme (GBM), a common and devastating brain tumor in humans. IR has been shown to induce PI3K-Akt activation in many cell types, and activation of the PI3K-Akt signaling pathway has been correlated with radioresistance. Methods Initially, the effects of IR on Akt activation were assessed in multiple human GBM cell lines. Next, to evaluate a potential causative role of IR-induced Akt activation on radiosensitivity, Akt activation was inhibited during IR with several complementary genetic and pharmacological approaches, and radiosensitivity measured using clonogenic survival assays. Results Three of the eight cell lines tested demonstrated IR-induced Akt activation. Further studies revealed that IR-induced Akt activation was dependent upon the presence of a serum factor, and could be inhibited by the EGFR inhibitor AG1478. Inhibition of PI3K activation with LY294002, or with inducible wild-type PTEN, inhibition of EGFR, as well as direct inhibition of Akt with two Akt inhibitors during irradiation increased the radiosensitivity of U87MG cells. Conclusion These results suggest that Akt may be a central player in a feedback loop whereby activation of Akt induced by IR increases radioresistance of GBM cells. Targeting the Akt signaling pathway may have important therapeutic implications when used in combination with IR in the treatment of a subset of brain tumor patients. PMID:19828040

  18. Molecular Mechanisms of Radiation-Induced Genomic Instability in Human Cells

    SciTech Connect

    Howard L. Liber; Jeffrey L. Schwartz

    2005-10-31

    There are many different model systems that have been used to study chromosome instability. What is clear from all these studies is that conclusions concerning chromosome instability depend greatly on the model system and instability endpoint that is studied. The model system for our studies was the human B-lymphoblastoid cell line TK6. TK6 was isolated from a spontaneously immortalized lymphoblast culture. Thus there was no outside genetic manipulation used to immortalize them. TK6 is a relatively stable p53-normal immortal cell line (37). It shows low gene and chromosome mutation frequencies (19;28;31). Our general approach to studying instability in TK6 cells has been to isolate individual clones and analyze gene and chromosome mutation frequencies in each. This approach maximizes the possibility of detecting low frequency events that might be selected against in mass cultures.

  19. Radiation-induced cell inactivation as a cause for cancer promotion

    NASA Astrophysics Data System (ADS)

    Heidenreich, W. F.; Paretzke, H. G.

    In space research, estimates of health risks from high-LET radiation, as well as from mixed fields are needed. Features of several applications of the biologically based two step clonal expansion (TSCE) model on data with high-LET radiation, normally α-particles from radon and from Thorotrast, are reviewed. One conclusion is that radiation might not only influence the initiating event in carcinogenesis, but may also act as a promoter. A possible mechanism which gives a promoting action from cell inactivation is presented for the organs "lung", with a two-dimensional arrangement of the cells at risk, and for "liver" where the sensitive cells are distributed in all three dimensions. Inferences for dose-response curves at low doses and dose rates are drawn. For liver, the number and size of premalignant clones is estimated from the cancer data.

  20. Hypoxia- and radiation-inducible, breast cell-specific targeting of retroviral vectors

    SciTech Connect

    Lipnik, Karoline; Greco, Olga; Scott, Simon; Knapp, Elzbieta; Mayrhofer, Elisabeth; Rosenfellner, Doris; Guenzburg, Walter H.; Salmons, Brian; Hohenadl, Christine . E-mail: christine.hohenadl@vu-wien.ac.at

    2006-05-25

    To facilitate a more efficient radiation and chemotherapy of mammary tumours, synthetic enhancer elements responsive to hypoxia and ionizing radiation were coupled to the mammary-specific minimal promoter of the murine whey acidic protein (WAP) encoding gene. The modified WAP promoter was introduced into a retroviral promoter conversion (ProCon) vector. Expression of a transduced reporter gene in response to hypoxia and radiation was analysed in stably infected mammary cancer cell lines and an up to 9-fold increase in gene expression demonstrated in comparison to the respective basic vector. Expression analyses in vitro, moreover, demonstrated a widely preserved mammary cell-specific promoter activity. For in vivo analyses, xenograft tumours consisting of infected human mammary adenocarcinoma cells were established in SCID/beige mice. Immunohistochemical analyses demonstrated a hypoxia-specific, markedly increased WAP promoter-driven expression in these tumours. Thus, this retroviral vector will facilitate a targeted gene therapeutic approach exploiting the unique environmental condition in solid tumours.

  1. Catalase inhibits ionizing radiation-induced apoptosis in hematopoietic stem and progenitor cells.

    PubMed

    Xiao, Xia; Luo, Hongmei; Vanek, Kenneth N; LaRue, Amanda C; Schulte, Bradley A; Wang, Gavin Y

    2015-06-01

    Hematologic toxicity is a major cause of mortality in radiation emergency scenarios and a primary side effect concern in patients undergoing chemo-radiotherapy. Therefore, there is a critical need for the development of novel and more effective approaches to manage this side effect. Catalase is a potent antioxidant enzyme that coverts hydrogen peroxide into hydrogen and water. In this study, we evaluated the efficacy of catalase as a protectant against ionizing radiation (IR)-induced toxicity in hematopoietic stem and progenitor cells (HSPCs). The results revealed that catalase treatment markedly inhibits IR-induced apoptosis in murine hematopoietic stem cells and hematopoietic progenitor cells. Subsequent colony-forming cell and cobble-stone area-forming cell assays showed that catalase-treated HSPCs can not only survive irradiation-induced apoptosis but also have higher clonogenic capacity, compared with vehicle-treated cells. Moreover, transplantation of catalase-treated irradiated HSPCs results in high levels of multi-lineage and long-term engraftments, whereas vehicle-treated irradiated HSPCs exhibit very limited hematopoiesis reconstituting capacity. Mechanistically, catalase treatment attenuates IR-induced DNA double-strand breaks and inhibits reactive oxygen species. Unexpectedly, we found that the radioprotective effect of catalase is associated with activation of the signal transducer and activator of transcription 3 (STAT3) signaling pathway and pharmacological inhibition of STAT3 abolishes the protective activity of catalase, suggesting that catalase may protect HSPCs against IR-induced toxicity via promoting STAT3 activation. Collectively, these results demonstrate a previously unrecognized mechanism by which catalase inhibits IR-induced DNA damage and apoptosis in HSPCs.

  2. Radiation-induced DNA double-strand break rejoining in human tumour cells.

    PubMed Central

    Núñez, M. I.; Villalobos, M.; Olea, N.; Valenzuela, M. T.; Pedraza, V.; McMillan, T. J.; Ruiz de Almodóvar, J. M.

    1995-01-01

    Five established human breast cancer cell lines and one established human bladder cancer cell line of varying radiosensitivity have been used to determine whether the rejoining of DNA double-strand breaks (dsbs) shows a correlation with radiosensitivity. The kinetics of dsb rejoining was biphasic and both components proceeded exponentially with time. The half-time (t1/2) of rejoining ranged from 18.0 +/- 1.4 to 36.4 +/- 3.2 min (fast rejoining process) and from 1.5 +/- 0.2 to 5.1 +/- 0.2 h (slow rejoining process). We found a statistically significant relationship between the survival fraction at 2 Gy (SF2) and the t1/2 of the fast rejoining component (r = 0.949, P = 0.0039). Our results suggest that cell lines which show rapid rejoining are more radioresistant. These results support the view that, as well as the level of damage induction that we have reported previously, the repair process is a major determinant of cellular radiosensitivity. It is possible that the differences found in DNA dsb rejoining and the differences in DNA dsb induction are related by a common mechanism, e.g. conformation of chromatin in the cell. PMID:7841046

  3. Ionizing radiation-induced mutation of human cells with different DNA repair capacities

    NASA Astrophysics Data System (ADS)

    Amundson, S. A.; Chen, D. J.

    We have observed significant differences in the response to ionizing radiation of two closely related human cell lines, and now compare the effects on these lines of both low and intermediate LET radiation. Compared to TK6, WTK1 has an enhanced X-ray survival, and is also more resistant to cell killing by alpha-particles. The hprt locus is more mutable in WTK1 than in TK6 by both X-rays and alpha-particles. WTK1 is also more mutable by alpha-particles than by X-rays at the hprt locus. X-ray-induced mutation at the heterozygous tk locus in WTK1 is about 25 fold higher than in TK6, while alpha-particle-induced mutation is nearly 50 fold higher at this locus. Also, the slowly growing tk- mutants, which comprise the majority of spontaneous and X-ray-inducedtk - mutants of TK6, were not induced significantly by alpha-particles. Previously, we showed that TK6 has a reduced capacity for recombination compared with WTK1, and therefore, these results indicate that recombinational repair may contribute to both cell survival and mutation-induction following exposure to ionizing radiation. Such a mechanism may aid cell survival, but could also result in increased deleterious effects such as the unmasking of recessive mutations in cancer suppresser genes.

  4. Radiation-induced PGE2 sustains human glioma cells growth and survival through EGF signaling.

    PubMed

    Brocard, Emeline; Oizel, Kristell; Lalier, Lisenn; Pecqueur, Claire; Paris, François; Vallette, François M; Oliver, Lisa

    2015-03-30

    Glioblastoma Multiforme (GBM) is the most common brain cancer in adults. Radiotherapy (RT) is the most effective post-operative treatment for the patients even though GBM is one of the most radio-resistant tumors. Dead or dying cells within the tumor are thought to promote resistance to treatment through mechanisms that are very poorly understood. We have evaluated the role of Prostaglandin E2 (PGE2), a versatile bioactive lipid, in GBM radio-resistance. We used an in vitro approach using 3D primary cultures derived from representative GBM patients. We show that irradiated glioma cells produced and released PGE2 in important quantities independently of the induction of cell death. We demonstrate that the addition of PGE2 enhances cell survival and proliferation though its ability to trans-activate the Epithelial Growth Factor receptor (EGFR) and to activate β-catenin. Indeed, PGE2 can substitute for EGF to promote primary cultures survival and growth in vitro and the effect is likely to occur though the Prostaglandin E2 receptor EP2.

  5. Radiation-induced PGE2 sustains human glioma cell growth and survival through EGF signaling

    PubMed Central

    Lalier, Lisenn; Pecqueur, Claire; Paris, François; Vallette, François M.; Oliver, Lisa

    2015-01-01

    Glioblastoma Multiforme (GBM) is the most common brain cancer in adults. Radiotherapy (RT) is the most effective post-operative treatment for the patients even though GBM is one of the most radio-resistant tumors. Dead or dying cells within the tumor are thought to promote resistance to treatment through mechanisms that are very poorly understood. We have evaluated the role of Prostaglandin E2 (PGE2), a versatile bioactive lipid, in GBM radio-resistance. We used an in vitro approach using 3D primary cultures derived from representative GBM patients. We show that irradiated glioma cells produced and released PGE2 in important quantities independently of the induction of cell death. We demonstrate that the addition of PGE2 enhances cell survival and proliferation though its ability to trans-activate the Epithelial Growth Factor receptor (EGFR) and to activate β-catenin. Indeed, PGE2 can substitute for EGF to promote primary cultures survival and growth in vitro and the effect is likely to occur though the Prostaglandin E2 receptor EP2. PMID:25749386

  6. Role of ion channels in ionizing radiation-induced cell death.

    PubMed

    Huber, Stephan M; Butz, Lena; Stegen, Benjamin; Klumpp, Lukas; Klumpp, Dominik; Eckert, Franziska

    2015-10-01

    Neoadjuvant, adjuvant or definitive fractionated radiation therapy are implemented in first line anti-cancer treatment regimens of many tumor entities. Ionizing radiation kills the tumor cells mainly by causing double strand breaks of their DNA through formation of intermediate radicals. Survival of the tumor cells depends on both, their capacity of oxidative defense and their efficacy of DNA repair. By damaging the targeted cells, ionizing radiation triggers a plethora of stress responses. Among those is the modulation of ion channels such as Ca2+-activated K+ channels or Ca2+-permeable nonselective cation channels belonging to the super-family of transient receptor potential channels. Radiogenic activation of these channels may contribute to radiogenic cell death as well as to DNA repair, glucose fueling, radiogenic hypermigration or lowering of the oxidative stress burden. The present review article introduces these channels and summarizes our current knowledge on the mechanisms underlying radiogenic ion channel modulation. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. A novel multitarget model of radiation-induced cell killing based on the Gaussian distribution.

    PubMed

    Zhao, Lei; Mi, Dong; Sun, Yeqing

    2017-03-08

    The multitarget version of the traditional target theory based on the Poisson distribution is still used to describe the dose-survival curves of cells after ionizing radiation in radiobiology and radiotherapy. However, noting that the usual ionizing radiation damage is the result of two sequential stochastic processes, the probability distribution of the damage number per cell should follow a compound Poisson distribution, like e.g. Neyman's distribution of type A (N. A.). In consideration of that the Gaussian distribution can be considered as the approximation of the N. A. in the case of high flux, a multitarget model based on the Gaussian distribution is proposed to describe the cell inactivation effects in low linear energy transfer (LET) radiation with high dose-rate. Theoretical analysis and experimental data fitting indicate that the present theory is superior to the traditional multitarget model and similar to the Linear - Quadratic (LQ) model in describing the biological effects of low-LET radiation with high dose-rate, and the parameter ratio in the present model can be used as an alternative indicator to reflect the radiation damage and radiosensitivity of the cells.

  8. Ionizing radiation-induced mutation of human cells with different DNA repair capacities

    SciTech Connect

    Amundson, S.A.; Chen, D.J.

    1994-12-31

    We have observed significant differences in the response to ionizing radiation of two closely related human cell lines, and now compare the effects on these lines of both low and intermediate LET radiation. Compared to TK6, WTK1 has an enhanced X-ray survival, and is also more resistant to cell killing by {alpha}-particles. The hprt locus is more mutable in WTK1 than in TK6 by both X-rays and {alpha}-particles. WTK1 is also more mutable by {alpha}-particles than by X-rays at the hprt locus. X-ray-induced mutation at the heterozygous tk locus in WTK1 is about 25 fold higher than in TK6, while {alpha}-particle-induced mutation is nearly 50 fold higher at this locus. Also, the slowly growing tk- mutants, which comprise the majority of spontaneous and X-ray-induced tk- mutants of TK6, were not induced significantly by {alpha}-particles. Previously, we showed that TK6 has a reduced capacity for recombination compared with WTK1, and therefore, these results indicate that recombinational repair may contribute to both cell survival and mutation-induction following exposure to ionizing radiation. Such a mechanism may aid cell survival, but could also result in increased deleterious effects such as the unmasking of recessive mutations in cancer suppresser genes.

  9. Apoptotic pathway induced by noscapine in human myelogenous leukemic cells.

    PubMed

    Heidari, Nastaran; Goliaei, Bahram; Moghaddam, Parvaneh Rahimi; Rahbar-Roshandel, Nahid; Mahmoudian, Massoud

    2007-11-01

    It has been shown that noscapine, an opium-derived phthalideisoquinoline alkaloid that is currently being used as an oral antitussive drug, induces apoptosis in myeloid leukemia cells. The molecular mechanism responsible for the anticancer effects of noscapine is poorly understood. In the current study, the apoptotic effects of noscapine on two myeloid cell lines, apoptosis-proficient HL60 cells and apoptosis-resistant K562 cells, were analyzed. An increase in the activity of caspase-2, -3, -6, -8 and -9, poly(ADP ribose) polymerase cleavage, detection of phosphatidylserine on the outer layer of the cell membrane, nucleation of chromatin, and DNA fragmentation suggested the induction of apoptosis. Noscapine increased the Bax/Bcl-2 ratio with a significant decrease of Bcl-2 expression accompanied with Bcl-2 phosphorylation. Using an inhibitory approach, the activation of the caspase cascade involved in the noscapine-induced apoptosis was analyzed. We observed no inhibitory effect of the caspase-8 inhibitor on caspase-9 activity. In view of these results and taking into consideration that K562 cells are Fas-null, we suggested that caspase-8 is activated in a Fas-independent manner downstream of caspase-9. In conclusion, noscapine can induce apoptosis in both apoptosis-proficient and apoptosis-resistant leukemic cells, and it can be a novel candidate in the treatment of hematological malignancies.

  10. T cell targeting and phagocytosis of apoptotic biliary epithelial cells in primary biliary cirrhosis.

    PubMed

    Allina, Jorge; Hu, Bin; Sullivan, Daniel M; Fiel, Maria Isabel; Thung, Swan N; Bronk, Steven F; Huebert, Robert C; van de Water, Judy; LaRusso, Nicholas F; Gershwin, M E; Gores, Gregory J; Odin, Joseph A

    2006-12-01

    Primary biliary cirrhosis (PBC) is characterized by loss of tolerance against ubiquitously expressed mitochondrial autoantigens followed by biliary and salivary gland epithelial cell (BEC and SGEC) destruction by autoreactive T cells. It is unclear why BECs and SGECs are targeted. Previous work demonstrated that the reduced form of the major PBC autoantigen predominated in apoptotic BECs and SGECs as opposed to an oxidized form in other apoptotic cells. This led to the hypothesis that presentation of novel self-peptides from phagocytosed apoptotic BECs might contribute to BEC targeting by autoreactive T cells. The effect of autoantigen redox status on self-peptide formation was examined along with the phagocytic ability of BECs. Oxidation of PBC autoantigens first was shown to be due to protein S-glutathionylation of lipoyllysine residues. Absence of protein S-glutathionylation generated novel self-peptides and affected T cell recognition of a lipoyllysine containing peptide. Liver biopsy staining revealed BEC phagocytosis of apoptotic BECs (3.74+/-2.90% of BEC) was present in PBC (7 of 7 cases) but not in normal livers (0 of 3). BECs have the ability to present novel mitochondrial self-peptides derived from phagocytosed apoptotic BECs. Apoptotic cell phagocytosis by non-professional phagocytes may influence the tissue specificity of autoimmune diseases.

  11. HMGB1/RAGE axis promotes autophagy and protects keratinocytes from ultraviolet radiation-induced cell death.

    PubMed

    Mou, Kuanhou; Liu, Wei; Han, Dan; Li, Pan

    2017-03-01

    The primary cause of skin cancer is ultraviolet (UV) light from the sun. Keratinocytes are the predominant cell type in the epidermis and form a barrier against environmental damage, especially from UV light irradiation. Autophagy is a self-digestion mechanism for energy homeostasis at critical times during development and as a response to stress. High-mobility group protein 1 (HMGB1) is a highly conserved nuclear protein that is associated with cell autophagy. We investigated the role of HMGB1 in keratinocytes exposed to UV irradiation and its regulation of keratinocyte autophagy. Specimens of UV-exposed human skin were assayed immunohistochemically for HMGB1. HaCaT immortalized human keratinocytes were used to investigate the mechanism of HMGB1 translocation induced by UV irradiation. Levels of cytosolic reactive oxygen species (ROS) were determined by H2DCF assay, apoptosis was assayed by flow cytometry and western-blot after lentivirus-mediated shRNA targeting of HMGB1 in keratinocytes by UV irradiation. Phosphorylated-Erk1/2 expression was assayed by western blotting. HMGB1 and its receptor (receptor for advanced glycation end products, RAGE) were both expressed by HaCaT cells, and HMGB1 was transferred from the nucleus to the cytoplasm after UV irradiation in both HaCaT and human skin keratinocytes. Knockdown of HMGB1 expression by lentivirus-mediated shRNA limited UV-induced autophagy and led to increased apoptosis of HaCaT cells. Pharmacological inhibition of HMGB1 cytoplasmic translocation by agents such as ethyl pyruvate limits starvation-induced autophagy. UV irradiation led to phosphorylation of Erk1/2 in HaCaT cells. Inhibition of RAGE and Erk1/2 limited HaCaT cell autophagy. Autocrine HMGB1 modulated HaCaT autophagy via a RAGE/HMGB1/extracellular signal-regulated Erk1/2-dependent pathway to protect keratinocytes from apoptosis during UV irradiation. Copyright © 2016 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All

  12. Pyrrolidine dithiocarbamate (PDTC) blocks apoptosis and promotes ionizing radiation-induced necrosis of freshly-isolated normal mouse spleen cells.

    PubMed

    Thompson, John S; Asmis, Reto; Tapp, Andrea A; Nelson, Brandy; Chu, Yanxia; Glass, Judith; Moneyhon, Micheal; Brown, Stephen A

    2010-06-01

    Ionizing radiation (IR) is a pro-oxidant that kills cells by both apoptotic and necrotic mechanisms. Pyrrolidine dithiocarbamate (PDTC) is a thiol-containing compound that may act either as a pro- or anti-oxidant depending on the experimental conditions. This study was designed to determine whether PDTC would reduce or enhance IR-induced cell death of freshly-isolated normal mouse B6/129 spleen cells (NMSC). We determined the effect of increasing doses of IR, PDTC alone and PDTC followed by IR on the viability of NMSC. Annexin V and propidium iodide (Annexin V/PI) staining demonstrated a dose and time-dependent relationship in which PDTC enhanced the percentage of IR-induced apoptotic/necrotic NMSC. Trypan blue dye inclusion confirmed that a loss of membrane integrity was occurring 1 h after incubation with PDTC plus IR. Reduction in the glutathione (GSH)/glutathione disulfide (GSSG) ratio and GSH demonstrated that both IR (8.5 Gy) and PDTC acted as pro-oxidants, but their mechanisms of action differed: In contrast to IR, which promoted p53 activation and caspase 3/7-mediated apoptosis, PDTC inhibited IR-induced p53 and caspase 3/7 activity. However, PDTC increased H(2)O(2) formation and necrosis, resulting in an overall increase in IR-induced cell death. Catalase prevented the PDTC-induced increase in IR cytotoxicity implicating the generation of H(2)O(2) as a major factor in this mechanism. These results demonstrate that in NMSC PDTC acts as pro-oxidant and enhances IR-induced cell cytotoxicity by increasing H(2)O(2)formation and thiol oxidation. As such, they strongly suggest that the use of PDTC as an adjunct to reduce radiation toxicity should be avoided.

  13. Influence of the circadian rhythm in cell division on radiation-induced mitotic delay in vivo

    SciTech Connect

    Rubin, N.H.

    1982-01-01

    Mitotic delay is described as a classical response to radiation; however, circadian rhythmicity in cell division in vivo has not been considered by many authors. The present study investigated the relation between fluctuations reported as mitotic delay and recovery in vivo and circadian oscillations in mitotic index in mouse corneal epithelium. One aspect involved single doses (approximately 600 rad) given to mice at different circadian stages. The normal circadian rhythm in cell division was never obliterated. Inhibition of mitosis was evident but unpredictable, ranging from 6 to 15 hr after irradiation. Recovery was evident only during the daily increase in mitotic index of controls. The classical interpretation of recovery from mitotic delay may be in an in vitro phenomenon not reflecting in vivo responses, which are apparently strongly circadian stage dependent. The second portion of the study demonstrated a dose-response effect on length of mitotic delay and, to a lesser extent, degree of recovery.

  14. Particle Radiation-Induced Nontargeted Effects in Bone-Marrow-Derived Endothelial Progenitor Cells

    PubMed Central

    Sasi, Sharath P.; Park, Daniel; Muralidharan, Sujatha; Wage, Justin; Kiladjian, Albert; Onufrak, Jillian; Enderling, Heiko; Yan, Xinhua; Goukassian, David A.

    2015-01-01

    Bone-marrow- (BM-) derived endothelial progenitor cells (EPCs) are critical for endothelial cell maintenance and repair. During future space exploration missions astronauts will be exposed to space irradiation (IR) composed of a spectrum of low-fluence protons (1H) and high charge and energy (HZE) nuclei (e.g., iron-56Fe) for extended time. How the space-type IR affects BM-EPCs is limited. In media transfer experiments in vitro we studied nontargeted effects induced by 1H- and 56Fe-IR conditioned medium (CM), which showed significant increase in the number of p-H2AX foci in nonirradiated EPCs between 2 and 24 h. A 2–15-fold increase in the levels of various cytokines and chemokines was observed in both types of IR-CM at 24 h. Ex vivo analysis of BM-EPCs from single, low-dose, full-body 1H- and 56Fe-IR mice demonstrated a cyclical (early 5–24 h and delayed 28 days) increase in apoptosis. This early increase in BM-EPC apoptosis may be the effect of direct IR exposure, whereas late increase in apoptosis could be a result of nontargeted effects (NTE) in the cells that were not traversed by IR directly. Identifying the role of specific cytokines responsible for IR-induced NTE and inhibiting such NTE may prevent long-term and cyclical loss of stem and progenitors cells in the BM milieu. PMID:26074973

  15. Radiation-induced senescence-like phenotype in proliferating and plateau-phase vascular endothelial cells

    SciTech Connect

    Igarashi, Kaori; Sakimoto, Ippei; Kataoka, Keiko; Ohta, Keisuke; Miura, Masahiko

    2007-09-10

    The effects of ionizing radiation (IR) on tumor angiogenesis still remain largely unknown. In this study, we found that IR (8 Gy) induces a high-frequency (80-90%) senescence-like phenotype in vascular endothelial cells (ECs) undergoing exponential growth. This finding allowed us to characterize the IR-induced senescence-like (IRSL) phenotype by examining the gene expression profiles and in vitro angiogenic activities of these ECs. The expression levels of genes associated with cell cycle progression and DNA replication were remarkably reduced in the IRSL ECs. Additionally, the in vitro invasion and migration activities of these cells through Matrigel were significantly suppressed. We also found that confluent ECs exhibited a high-frequency IRSL phenotype when they were replated immediately after irradiation, whereas incubation in plateau-phase conditions reduced the induction of this phenotype and enhanced colony formation. The kinetics of DNA double-strand break repair, which showed a faster time course in confluent ECs than in growing ECs, may contribute to the protective mechanism associated with the IRSL phenotype. These results imply that the IRSL phenotype may be important for determining the angiogenic activity of ECs following irradiation. The present study should contribute to the understanding of the effects of IR on tumor angiogenesis.

  16. Galleria mellonella lysozyme induces apoptotic changes in Candida albicans cells.

    PubMed

    Sowa-Jasiłek, Aneta; Zdybicka-Barabas, Agnieszka; Stączek, Sylwia; Wydrych, Jerzy; Skrzypiec, Krzysztof; Mak, Paweł; Deryło, Kamil; Tchórzewski, Marek; Cytryńska, Małgorzata

    2016-12-01

    The greater wax moth Galleria mellonella has been increasingly used as a model host to determine Candida albicans virulence and efficacy of antifungal treatment. The G. mellonella lysozyme, similarly to its human counterpart, is a member of the c-type family of lysozymes that exhibits antibacterial and antifungal activity. However, in contrast to the relatively well explained bactericidal action, the mechanism of fungistatic and/or fungicidal activity of lysozymes is still not clear. In the present study we provide the direct evidences that the G. mellonella lysozyme binds to the protoplasts as well as to the intact C. albicans cells and decreases the survival rate of both these forms in a time-dependent manner. No enzymatic activity of the lysozyme towards typical chitinase and β-glucanase substrates was detected, indicating that hydrolysis of main fungal cell wall components is not responsible for anti-Candida activity of the lysozyme. On the other hand, pre-treatment of cells with tetraethylammonium, a potassium channel blocker, prevented them from the lysozyme action, suggesting that lysozyme acts by induction of programmed cell death. In fact, the C. albicans cells treated with the lysozyme exhibited typical apoptotic features, i.e. loss of mitochondrial membrane potential, phosphatidylserine exposure in the outer leaflet of the cell membrane, as well as chromatin condensation and DNA fragmentation.

  17. The scavenger receptor SCARF1 mediates apoptotic cell clearance and prevents autoimmunity

    PubMed Central

    Ramirez-Ortiz, Zaida G.; Pendergraft, William F.; Prasad, Amit; Byrne, Michael H.; Iram, Tal; Blanchette, Christopher J.; Luster, Andrew D.; Hacohen, Nir; Khoury, Joseph El; Means, Terry K.

    2013-01-01

    Clearance of apoptotic cells is critical for control of tissue homeostasis however the full range of receptor(s) on phagocytes responsible for recognition of apoptotic cells remains to be identified. Here we show that dendritic cells (DCs), macrophages and endothelial cells use scavenger receptor type F family member 1 (SCARF1) to recognize and engulf apoptotic cells via C1q. Loss of SCARF1 impairs uptake of apoptotic cells. Consequently, in SCARF1-deficient mice, dying cells accumulate in tissues leading to a lupus-like disease with the spontaneous generation of autoantibodies to DNA-containing antigens, immune cell activation, dermatitis and nephritis. The discovery of SCARF1 interactions with C1q and apoptotic cells provides insights into molecular mechanisms involved in maintenance of tolerance and prevention of autoimmune disease. PMID:23892722

  18. Cooperative binding of Annexin A5 to phosphatidylserine on apoptotic cell membranes

    NASA Astrophysics Data System (ADS)

    Janko, Christina; Jeremic, Ivica; Biermann, Mona; Chaurio, Ricardo; Schorn, Christine; Muñoz, Luis E.; Herrmann, Martin

    2013-12-01

    Healthy cells exhibit an asymmetric plasma membrane with phosphatidylserine (PS) located on the cytoplasmic leaflet of the plasma membrane bilayer. Annexin A5-FITC, a PS binding protein, is commonly used to evaluate apoptosis in flow cytometry. PS exposed by apoptotic cells serves as a major ‘eat-me’ signal for phagocytes. Although exposition of PS has been observed after alternative stimuli, no clearance of viable, PS exposing cells has been detected. Thus, besides PS exposure, membranes of viable and apoptotic cells might exhibit specific characteristics. Here, we show that Annexin A5 binds in a cooperative manner to different types of dead cells. Shrunken apoptotic cells thereby showed the highest Hill coefficient values. Contrarily, parafomaldehyde fixation of apoptotic cells completely abrogates the cooperativity effect seen with dead and dying cells. We tend to speculate that the cooperative binding of Annexin A5 to the membranes of apoptotic cells reflects higher fluidity of the exposed membranes facilitating PS clustering.

  19. Radiation-Induced Micro-RNA Expression Changes in Peripheral Blood Cells of Radiotherapy Patients

    SciTech Connect

    Templin, Thomas; Paul, Sunirmal; Amundson, Sally A.; Young, Erik F.; Barker, Christopher A.; Wolden, Suzanne L.; Smilenov, Lubomir B.

    2011-06-01

    Purpose: MicroRNAs (miRNAs), a class of noncoding small RNAs that regulate gene expression, are involved in numerous physiologic processes in normal and malignant cells. Our in vivo study measured miRNA and gene expression changes in human blood cells in response to ionizing radiation, to develop miRNA signatures that can be used as biomarkers for radiation exposure. Methods and Materials: Blood from 8 radiotherapy patients in complete remission 1 or 2 was collected immediately before and 4 hours after total body irradiation with 1.25 Gy x-rays. Both miRNA and gene expression changes were measured by means of quantitative polymerase chain reaction and microarray hybridization, respectively. Hierarchic clustering, multidimensional scaling, class prediction, and gene ontology analysis were performed to investigate the potential of miRNAs to serve as radiation biomarkers and to elucidate their likely physiologic roles in the radiation response. Results: The expression levels of 45 miRNAs were statistically significantly upregulated 4 hours after irradiation with 1.25 Gy x-rays, 27 of them in every patient. Nonirradiated and irradiated samples form separate clusters in hierarchic clustering and multidimensional scaling. Out of 223 differentially expressed genes, 37 were both downregulated and predicted targets of the upregulated miRNAs. Paired and unpaired miRNA-based classifiers that we developed can predict the class membership of a sample with unknown irradiation status, with accuracies of 100% when all 45 upregulated miRNAs are included. Both miRNA control of and gene involvement in biologic processes such as hemopoiesis and the immune response are increased after irradiation, whereas metabolic processes are underrepresented among all differentially expressed genes and the genes controlled by miRNAs. Conclusions: Exposure to ionizing radiation leads to the upregulation of the expression of a considerable proportion of the human miRNAome of peripheral blood cells

  20. Ionizing radiation-induced metabolic oxidative stress and prolonged cell injury

    PubMed Central

    Azzam, Edouard I.; Jay-Gerin, Jean-Paul; Pain, Debkumar

    2013-01-01

    Cellular exposure to ionizing radiation leads to oxidizing events that alter atomic structure through direct interactions of radiation with target macromolecules or via products of water radiolysis. Further, the oxidative damage may spread from the targeted to neighboring, non-targeted bystander cells through redox-modulated intercellular communication mechanisms. To cope with the induced stress and the changes in the redox environment, organisms elicit transient responses at the molecular, cellular and tissue levels to counteract toxic effects of radiation. Metabolic pathways are induced during and shortly after the exposure. Depending on radiation dose, dose-rate and quality, these protective mechanisms may or may not be sufficient to cope with the stress. When the harmful effects exceed those of homeostatic biochemical processes, induced biological changes persist and may be propagated to progeny cells. Physiological levels of reactive oxygen and nitrogen species play critical roles in many cellular functions. In irradiated cells, levels of these reactive species may be increased due to perturbations in oxidative metabolism and chronic inflammatory responses, thereby contributing to the long-term effects of exposure to ionizing radiation on genomic stability. Here, in addition to immediate biological effects of water radiolysis on DNA damage, we also discuss the role of mitochondria in the delayed outcomes of ionization radiation. Defects in mitochondrial functions lead to accelerated aging and numerous pathological conditions. Different types of radiation vary in their linear energy transfer (LET) properties, and we discuss their effects on various aspects of mitochondrial physiology. These include short and long-term in vitro and in vivo effects on mitochondrial DNA, mitochondrial protein import and metabolic and antioxidant enzymes. PMID:22182453

  1. Enhanced acoustic startle responding in rats with radiation-induced hippocampal granule cell hypoplasia

    SciTech Connect

    Mickley, G.A.; Ferguson, J.L.

    1989-01-01

    Irradiation of the neonatal rat hippocampus reduces the proliferation of granule cells in the dentate gyrus and results in locomotor hyperactivity, behavioral preservation, and deficits on some learned tasks. In order to address the role of changes in stimulus salience and behavioral inhibition in animals with this type of brain damage, irradiated and normal rats were compared in their startle reactions to an acoustic stimulus. Irradiated rats startled with a consistently higher amplitude than control and were more likely to exhibit startle responses. These animals with hippocampal damage also failed to habituate to the startle stimulus and, under certain circumstances, showed potentiated startle responses after many tone presentations.

  2. Mechanisms of Low Dose Radiation-induced T helper Cell Function

    SciTech Connect

    Gridley, Daila S.

    2008-10-31

    Exposure to radiation above levels normally encountered on Earth can occur during wartime, accidents such as those at Three Mile Island and Chernobyl, and detonation of “dirty bombs” by terrorists. Relatively high levels of radiation exposure can also occur in certain occupations (low-level waste sites, nuclear power plants, nuclear medicine facilities, airline industry, and space agencies). Depression or dysfunction of the highly radiosensitive cells of the immune system can lead to serious consequences, including increased risk for infections, cancer, hypersensitivity reactions, poor wound healing, and other pathologies. The focus of this research was on the T helper (Th) subset of lymphocytes that secrete cytokines (proteins), and thus control many actions and interactions of other cell types that make up what is collectively known as the immune system. The Department of Energy (DOE) Low Dose Radiation Program is concerned with mechanisms altered by exposure to high energy photons (x- and gamma-rays), protons and electrons. This study compared, for the first time, the low-dose effects of two of these radiation forms, photons and protons, on the response of Th cells, as well as other cell types with which they communicate. The research provided insights regarding gene expression patterns and capacity to secrete potent immunostimulatory and immunosuppressive cytokines, some of which are implicated in pathophysiological processes. Furthermore, the photon versus proton comparison was important not only to healthy individuals who may be exposed, but also to patients undergoing radiotherapy, since many medical centers in the United States, as well as worldwide, are now building proton accelerators. The overall hypothesis of this study was that whole-body exposure to low-dose photons (gamma-rays) will alter CD4+ Th cell function. We further proposed that exposure to low-dose proton radiation will induce a different pattern of gene and functional changes compared to

  3. Does radio frequency radiation induce micronuclei frequency in exfoliated bladder cells of diabetic rats?

    PubMed

    Gurbuz, N; Sirav, B; Kuzay, D; Ozer, C; Seyhan, N

    2015-07-01

    For many years there has been a discussion among both experts and the general public regarding the effects of radio frequency (RF) radiation on the human organism. The purpose of the present study was to evaluate the relationship of micronucleui (MN) frequency and RF radiation in exfoliated bladder cells of non-diabetic and diabetic rats. Three groups were used in the experiment: Group I (n=6): diabetic group without RF exposure; Group II (n=6): diabetic group exposed 2100 MHz RF radiation and Group III (n=6): control animals (non-diabetic group, no RF exposure). RF exposure in the experiment resulted in a whole body average SAR of 0.24 W/kg with an ERMS field of 17.5 V/m in non-thermal levels. Results showed that there was no statistically important differences between non-RF exposed diabetes group and control group; Group I and Group III (p>0.05). There was no statistically important differences between diabetes group and diabetes+RF exposed group (Group I and Group II) (p>0.05). RF exposure did not result in increased MN frequencies in exfoliated bladder cells of diabetic rats with respect to control animals (Group II and Group III), either and this result found no statistically important (p>0.05). This study suggested no possible genotoxic effects of RF radiation among human beings especially with chronic disorders, such as diabetes.

  4. Detection of space radiation-induced double strand breaks as a track in cell nucleus

    SciTech Connect

    Ohnishi, Takeo; Takahashi, Akihisa; Nagamatsu, Aiko; Omori, Katsunori; Suzuki, Hiromi; Shimazu, Toru; Ishioka, Noriaki

    2009-12-18

    To identify DNA damage induced by space radiations such as the high linear energy transfer (LET) particles, phospho-H2AX ({gamma}H2AX) foci formation was analyzed in human cells frozen in an International Space Station freezer for 133 days. After recovering the frozen sample to the earth, the cells were cultured for 30 min, and then fixed. Here we show a track of {gamma}H2AX positive foci in them by immuno-cytochemical methods. It is suggested that space radiations, especially high LET particles, induced DSBs as a track. From the formation of the tracks in nuclei, exposure dose rate was calculated to be 0.7 mSv per day as relatively high-energy space radiations of Fe-ions (500 MeV/u, 200 keV/{mu}m). From the physical dosimetry with CR-39 plastic nuclear track detectors and thermo-luminescent dosimeters, dose rate was 0.5 mSv per day. These values the exposed dose rate were similar between biological and physical dosimetries.

  5. Role of ion transport in control of apoptotic cell death.

    PubMed

    Lang, Florian; Hoffmann, Else K

    2012-07-01

    Cell shrinkage is a hallmark and contributes to signaling of apoptosis. Apoptotic cell shrinkage requires ion transport across the cell membrane involving K(+) channels, Cl(-) or anion channels, Na(+)/H(+) exchange, Na(+),K(+),Cl(-) cotransport, and Na(+)/K(+)ATPase. Activation of K(+) channels fosters K(+) exit with decrease of cytosolic K(+) concentration, activation of anion channels triggers exit of Cl(-), organic osmolytes, and HCO3(-). Cellular loss of K(+) and organic osmolytes as well as cytosolic acidification favor apoptosis. Ca(2+) entry through Ca(2+)-permeable cation channels may result in apoptosis by affecting mitochondrial integrity, stimulating proteinases, inducing cell shrinkage due to activation of Ca(2+)-sensitive K(+) channels, and triggering cell-membrane scrambling. Signaling involved in the modification of cell-volume regulatory ion transport during apoptosis include mitogen-activated kinases p38, JNK, ERK1/2, MEKK1, MKK4, the small G proteins Cdc42, and/or Rac and the transcription factor p53. Osmosensing involves integrin receptors, focal adhesion kinases, and tyrosine kinase receptors. Hyperosmotic shock leads to vesicular acidification followed by activation of acid sphingomyelinase, ceramide formation, release of reactive oxygen species, activation of the tyrosine kinase Yes with subsequent stimulation of CD95 trafficking to the cell membrane. Apoptosis is counteracted by mechanisms involved in regulatory volume increase (RVI), by organic osmolytes, by focal adhesion kinase, and by heat-shock proteins. Clearly, our knowledge on the interplay between cell-volume regulatory mechanisms and suicidal cell death is still far from complete and substantial additional experimental effort is needed to elucidate the role of cell-volume regulatory mechanisms in suicidal cell death. 2012 American Physiological Society. Compr Physiol 2:2037-2061, 2012.

  6. Radiation induced nuclear factor kappa-B signaling cascade study in mammalian cells by improved detection systems

    NASA Astrophysics Data System (ADS)

    Chishti, Arif Ali; Baumstark-Khan, Christa; Hellweg, Christine; Reitz, Guenther

    To enable long-term human space flight cellular radiation response to densely ionizing radiation needs to be better understood for developing appropriate countermeasures to mitigate acute effects and late radiation risks for the astronaut. The biological effectiveness of accelerated heavy ions with high linear energy transfer (LET) for effecting DNA damage response pathways as a gateway to cell death or survival is of major concern, not only for tumor radiotherapy but also for new regimes of space missions. Ionizing radiation modulates several signaling pathways resulting in transcription factor activation. NF-kappaB is one of the important transcription factors that respond to changes in the environment of a mammalian cell and plays a key role in many biological processes relevant to radiation response, such as apoptosis, inflammation and carcinogenesis. From medical and biological point of view it is important to understand radiation induced NF-kappaB signaling cascade. For studying NF-kappaB signaling, green fluorescent proteins EGFP and d2EGFP were used previously (Advances in Space Research, 36: 1673-1679, 2005). The current study aims to improve reporter assays by the use of a destabilized variant of red fluorescent protein tdTomato (DD-tdTomato) which gives high fluorescence signals and a better signal/noise ratio for NF-kappaB activation. The reporter system HEK-pNFkappaB-DD-tdTomato-C8 is a dual reporter system which can provide both discrete and cumulative signals after exposure to ionizing radiation (X-rays, heavy ions). In the presence of Shield-1, the fluorescent protein DD-tdTomato is not degraded but accumulated inside the cell which helps to quantify the fold induction of NF-kappaB-dependent gene expression. The minimum dose required to activate NF-kappaB is 6 Gy but accumulated signals data shows that NF-kappaB is activated after 3 Gy in the presence of Shield-1. Average dose and number of heavy ions’ hits per nucleus necessary to double the NF

  7. RADIATION-INDUCED MICRO-RNA EXPRESSION CHANGES IN PERIPHERAL BLOOD CELLS OF RADIOTHERAPY PATIENTS

    PubMed Central

    Templin, Thomas; Paul, Sunirmal; Amundson, Sally A.; Young, Erik F.; Barker, Christopher A.; Wolden, Suzanne L.; Smilenov, Lubomir B.

    2013-01-01

    Purpose MicroRNAs (miRNAs), a class of noncoding small RNAs that regulate gene expression, are involved in numerous physiologic processes in normal and malignant cells. Our in vivo study measured miRNA and gene expression changes in human blood cells in response to ionizing radiation, to develop miRNA signatures that can be used as biomarkers for radiation exposure. Methods and Materials Blood from 8 radiotherapy patients in complete remission 1 or 2 was collected immediately before and 4 hours after total body irradiation with 1.25 Gy x-rays. Both miRNA and gene expression changes were measured by means of quantitative polymerase chain reaction and microarray hybridization, respectively. Hierarchic clustering, multidimensional scaling, class prediction, and gene ontology analysis were performed to investigate the potential of miRNAs to serve as radiation biomarkers and to elucidate their likely physiologic roles in the radiation response. Results The expression levels of 45 miRNAs were statistically significantly upregulated 4 hours after irradiation with 1.25 Gy x-rays, 27 of them in every patient. Nonirradiated and irradiated samples form separate clusters in hierarchic clustering and multidimensional scaling. Out of 223 differentially expressed genes, 37 were both down-regulated and predicted targets of the upregulated miRNAs. Paired and unpaired miRNA-based classifiers that we developed can predict the class membership of a sample with unknown irradiation status, with accuracies of 100% when all 45 upregulated miRNAs are included. Both miRNA control of and gene involvement in biologic processes such as hemopoiesis and the immune response are increased after irradiation, whereas metabolic processes are underrepresented among all differentially expressed genes and the genes controlled by miRNAs. Conclusions Exposure to ionizing radiation leads to the upregulation of the expression of a considerable proportion of the human miRNAome of peripheral blood cells

  8. Plasmacytoid dendritic cells play a major role in apoptotic leukocyte-induced immune modulation.

    PubMed

    Bonnefoy, Francis; Perruche, Sylvain; Couturier, Mélanie; Sedrati, Abdeslem; Sun, Yunwei; Tiberghien, Pierre; Gaugler, Béatrice; Saas, Philippe

    2011-05-15

    Several APCs participate in apoptotic cell-induced immune modulation. Whether plasmacytoid dendritic cells (PDCs) are involved in this process has not yet been characterized. Using a mouse model of allogeneic bone marrow engraftment, we demonstrated that donor bone marrow PDCs are required for both donor apoptotic cell-induced engraftment and regulatory T cell (Treg) increase. We confirmed in naive mice receiving i.v. syngeneic apoptotic cell infusion that PDCs from the spleen induce ex vivo Treg commitment. We showed that PDCs did not interact directly with apoptotic cells. In contrast, in vivo macrophage depletion experiments using clodronate-loaded liposome infusion and coculture experiments with supernatant from macrophages incubated with apoptotic cells showed that PDCs required macrophage-derived soluble factors--including TGF-β--to exert their immunomodulatory functions. Overall, PDCs may be considered as the major APC involved in Treg stimulation/generation in the setting of an immunosuppressive environment obtained by apoptotic cell infusion. These findings show that like other APCs, PDC functions are influenced, at least indirectly, by exposure to blood-borne apoptotic cells. This might correspond with an additional mechanism preventing unwanted immune responses against self-antigens clustered at the cell surface of apoptotic cells occurring during normal cell turnover.

  9. Regulatory T Cells Contribute to the Inhibition of Radiation-Induced Acute Lung Inflammation via Bee Venom Phospholipase A2 in Mice

    PubMed Central

    Shin, Dasom; Lee, Gihyun; Sohn, Sung-Hwa; Park, Soojin; Jung, Kyung-Hwa; Lee, Ji Min; Yang, Jieun; Cho, Jaeho; Bae, Hyunsu

    2016-01-01

    Bee venom has long been used to treat various inflammatory diseases, such as rheumatoid arthritis and multiple sclerosis. Previously, we reported that bee venom phospholipase A2 (bvPLA2) has an anti-inflammatory effect through the induction of regulatory T cells. Radiotherapy is a common anti-cancer method, but often causes adverse effects, such as inflammation. This study was conducted to evaluate the protective effects of bvPLA2 in radiation-induced acute lung inflammation. Mice were focally irradiated with 75 Gy of X-rays in the lung and administered bvPLA2 six times after radiation. To evaluate the level of inflammation, the number of immune cells, mRNA level of inflammatory cytokine, and histological changes in the lung were measured. BvPLA2 treatment reduced the accumulation of immune cells, such as macrophages, neutrophils, lymphocytes, and eosinophils. In addition, bvPLA2 treatment decreased inflammasome-, chemokine-, cytokine- and fibrosis-related genes’ mRNA expression. The histological results also demonstrated the attenuating effect of bvPLA2 on radiation-induced lung inflammation. Furthermore, regulatory T cell depletion abolished the therapeutic effects of bvPLA2 in radiation-induced pneumonitis, implicating the anti-inflammatory effects of bvPLA2 are dependent upon regulatory T cells. These results support the therapeutic potential of bvPLA2 in radiation pneumonitis and fibrosis treatments. PMID:27144583

  10. Apoptotic cell death by the novel natural compound, cinobufotalin.

    PubMed

    Emam, Heba; Zhao, Qing-Li; Furusawa, Yukihiro; Refaat, Alaa; Ahmed, Kanwal; Kadowaki, Makoto; Kondo, Takashi

    2012-09-30

    Cinobufotalin (CB), one of the bufadienolides prepared from toad venom, was investigated for its cytotoxicity, and the underneath mechanism involved. We primarily utilized DNA fragmentation assay and microscopic observation to assess the effect of various doses of CB in human lymphoma U937 cells. Following that, we investigated other parameters involved in cell death mechanism such as reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and apoptotic proteins activation. HeLa cells were concomitantly used to generalize the data observed. Our results show that CB caused significant DNA fragmentation, decrease of MMP, and an increase in the intracellular Ca(2+) ion and ROS production. In addition, CB induced upregulation of Fas protein, proteolytic activation of cytochrome c, caspase-2, -3, -8 and -9 together with the activation of Bid and Bax. Our findings were further validated using either Fas/FasL antagonist or pan-caspase inhibitor to significantly inhibit CB-induced DNA fragmentation. In our study, we suggest that CB induces caspase dependent cell death in U937 cells, and that Fas plays a role in CB-induced apoptosis. Altogether, our data provides novel insights of the mechanism of action of CB and its potential as a future chemotherapeutic agent. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  11. Insights into the apoptotic death of immune cells in sepsis.

    PubMed

    Luan, Ying-yi; Yao, Yong-ming; Xiao, Xian-zhong; Sheng, Zhi-yong

    2015-01-01

    Sepsis with subsequent multiple-organ dysfunction is a distinct systemic inflammatory response to concealed or obvious infection, and it is a leading cause of death in intensive care units. Thus, one of the key goals in critical care medicine is to develop novel therapeutic strategies that will affect favorably on outcome of septic patients. In addition to systemic response to infection, apoptosis is implicated to be an important mechanism of the death of immune cells, including neutrophils, macrophages, T lymphocytes, and dendritic cells, and it is usually followed by the development of multiple-organ failure in sepsis. The implication of apoptosis of immune cells is now highlighted by multiple studies that demonstrate that prevention of cell apoptosis can improve survival in relevant animal models of severe sepsis. In this review, we focus on major apoptotic death pathways and molecular mechanisms that regulate apoptosis of different immune cells, and advances in these areas that may be translated into more promising therapies for the prevention and treatment of severe sepsis.

  12. Lysosomal photodamage induces cell death via mitochondrial apoptotic pathway

    NASA Astrophysics Data System (ADS)

    Liu, Lei; Wang, Xian-wang; Li, Hui

    2009-11-01

    Lysosomal photosensitizers have been used in photodynamic therapy (PDT). Combination of such photosensitizers and light causes lysosomal photodamage, inducing cell death. The lysosomal disruption can lead to apoptosis but its signaling pathways remain to be elucidated. In this study, we selected N-aspartyl chlorin e6 (NPe6), an effective photosensitizer which preferentially accumulates in lysosomes, to study the mechanism of apoptosis caused by lysosomal photodamage. Apoptosis in living human lung adenocarcinoma cells treated by NPe6-PDT was studied using real-time single-cell analysis. In this study, the fluorescence probes Cyto c-GFP and DsRed-Mit were used to detect the spatial and temporal changes of cytochrome c in real-time in sub-cell level; the Rhodamine 123 dyes were used to monitor the changes of mitochondrial membrane potential. The results showed that, after PDT treatment,the mitochondrial membrane potential decreased, and cytochrome c released from mitochondria; The caspase-3 was activated obviously. These results suggested that lysosomal photodamage activates mitochondrial apoptotic pathway to induce cell death.

  13. High- and low-LET Radiation-induced Chromosome Aberrations in Human Epithelial Cells Cultured in 3-dimensional Matrices

    NASA Technical Reports Server (NTRS)

    Hada, M.; George K.; Cucinotta, F. A.; Wu, H.

    2008-01-01

    Energetic heavy ions pose a great health risk to astronauts who participate in extended ISS missions and will be an even greater concern for future manned lunar and Mars missions. High-LET heavy ions are particularly effective in causing various biological effects, including cell inactivation, genetic mutations, cataracts and cancer induction. Most of these biological endpoints are closely related to chromosomal damage, which can be utilized as a biomarker for radiation insults. Previously, we had studied low- and high-LET radiation-induced chromosome aberrations in human epithelial cells cultured in 2-dimension (2D) using the multicolor banding fluorescence in situ hybridization (mBAND) technique. However, it has been realized that the biological response to radiation insult in a 2D in vitro cellular environment can differ significantly from the response in 3-dimension (3D) or at the actual tissue level. In this study, we cultured human epithelial cells in 3D to provide a more suitable model for human tissue. Human mammary epithelial cells (CH184B5F5/M10) were grown in Matrigel to form 3D structures, and exposed to Fe-ions at NASA Space Radiation Laboratory (NSRL) at the Brookhaven National Laboratory or 137Cs-gamma radiation source at the University of Texas MD Anderson Cancer Center. After exposure, cells were allowed to repair for 16hr before dissociation and subcultured at low density in 2D. G2 and metaphase chromosomes in the first cell cycle were collected in the first cell cycle after irradiation using a chemical-induced premature chromosome condensation (PCC) technique, and chromosome aberrations were analyzed using mBAND technique. With this technique, individually painted chromosomal bands on one chromosome allowed the identification of interchromosomal aberrations (translocation to unpainted chromosomes) and intrachromosomal aberrations (inversions and deletions within a single painted chromosome). Our data indicate a significant difference in the

  14. High- and low-LET Radiation-induced Chromosome Aberrations in Human Epithelial Cells Cultured in 3-dimensional Matrices

    NASA Technical Reports Server (NTRS)

    Hada, M.; George K.; Cucinotta, F. A.; Wu, H.

    2008-01-01

    Energetic heavy ions pose a great health risk to astronauts who participate in extended ISS missions and will be an even greater concern for future manned lunar and Mars missions. High-LET heavy ions are particularly effective in causing various biological effects, including cell inactivation, genetic mutations, cataracts and cancer induction. Most of these biological endpoints are closely related to chromosomal damage, which can be utilized as a biomarker for radiation insults. Previously, we had studied low- and high-LET radiation-induced chromosome aberrations in human epithelial cells cultured in 2-dimension (2D) using the multicolor banding fluorescence in situ hybridization (mBAND) technique. However, it has been realized that the biological response to radiation insult in a 2D in vitro cellular environment can differ significantly from the response in 3-dimension (3D) or at the actual tissue level. In this study, we cultured human epithelial cells in 3D to provide a more suitable model for human tissue. Human mammary epithelial cells (CH184B5F5/M10) were grown in Matrigel to form 3D structures, and exposed to Fe-ions at NASA Space Radiation Laboratory (NSRL) at the Brookhaven National Laboratory or 137Cs-gamma radiation source at the University of Texas MD Anderson Cancer Center. After exposure, cells were allowed to repair for 16hr before dissociation and subcultured at low density in 2D. G2 and metaphase chromosomes in the first cell cycle were collected in the first cell cycle after irradiation using a chemical-induced premature chromosome condensation (PCC) technique, and chromosome aberrations were analyzed using mBAND technique. With this technique, individually painted chromosomal bands on one chromosome allowed the identification of interchromosomal aberrations (translocation to unpainted chromosomes) and intrachromosomal aberrations (inversions and deletions within a single painted chromosome). Our data indicate a significant difference in the

  15. Dinitrophenol Inhibits the Rejoining of Radiation-Induced DNA Breaks by L-Cells

    PubMed Central

    Moss, A. J.; Dalrymple, Glenn V.; Sanders, J. L.; Wilkinson, K. P.; Nash, John C.

    1971-01-01

    The production and rejoining of X-ray-induced single-stranded DNA breaks was studied using the alkaline sucrose density gradient technique and by measuring the disappearance of both 5′ termini and 3′-OH termini using polynucleotide kinase and DNA polymerase, respectively. All studies were conducted using L-cell suspensions irradiated both in the presence and absence of 2,4-dinitrophenol (DNP), an uncoupler of oxidative phosphorylation. Results show that the induction of single-stranded DNA breaks probably includes a nucleolytic component in addition to indirect free radical effects. A greater number of breaks were produced in the absence of DNP, suggesting that depressed adenosine triphosphate (ATP) levels reduce endogenous nucleolytic activity. The rejoining mechanism is enzymatic and requires an available ATP supply for operation. In the presence of DNP no DNA rejoining was observed following 30 min incubation after 10,000 rad. These results suggest that DNA breaks produced may be characterized by 5′-PO4-3′-OH termini and are rejoined by DNA ligase. PMID:5542611

  16. Abdominal {gamma}-Radiation Induces an Accumulation of Function-Impaired Regulatory T Cells in the Small Intestine

    SciTech Connect

    Billiard, Fabienne; Buard, Valerie; Benderitter, Marc; Linard, Christine

    2011-07-01

    Purpose: To assess the frequency and the functional characteristics of one major component of immune tolerance, the CD4{sup +}FoxP3{sup +} regulatory T cells (Tregs) in a mouse model of abdominal irradiation. Methods and Materials: Mice were exposed to a single abdominal dose of {gamma}-radiation (10 Gy). We evaluated small intestine Treg infiltration by Foxp3 immunostaining and the functional suppressive activity of Tregs isolated from mesenteric lymph nodes. Results: Foxp3 immunostaining showed that radiation induced a long-term infiltration of the intestine by Tregs (levels 5.5 times greater than in controls). Co-culture of Tregs from mesenteric lymph nodes with CD4{sup +} effector cells showed that the Tregs had lost their suppressive function. This loss was associated with a significant decrease in the levels of Foxp3, TGF-{beta}, and CTLA-4 mRNA, all required for optimal Treg function. At Day 90 after irradiation, Tregs regained their suppressive activity as forkhead box P3 (Foxp3), transforming growth factor beta (TGF-{beta}), and cytotoxic T-lymphocyte antigen 4 (CTLA-4) expression returned to normal. Analysis of the secretory function of mesenteric lymph node Tregs, activated in vitro with anti-CD3/anti-CD28 Abs, showed that this dysfunction was independent of a defect in interleukin-10 secretion. Conclusion: Radiation caused a long-term accumulation of function-impaired Foxp3{sup +}CD4{sup +} Tregs in the intestine. Our study provides new insights into how radiation affects the immune tolerance in peripheral tissues.

  17. Barium inhibits arsenic-mediated apoptotic cell death in human squamous cell carcinoma cells.

    PubMed

    Yajima, Ichiro; Uemura, Noriyuki; Nizam, Saika; Khalequzzaman, Md; Thang, Nguyen D; Kumasaka, Mayuko Y; Akhand, Anwarul A; Shekhar, Hossain U; Nakajima, Tamie; Kato, Masashi

    2012-06-01

    Our fieldwork showed more than 1 μM (145.1 μg/L) barium in about 3 μM (210.7 μg/L) arsenic-polluted drinking well water (n = 72) in cancer-prone areas in Bangladesh, while the mean concentrations of nine other elements in the water were less than 3 μg/L. The types of cancer include squamous cell carcinomas (SCC). We hypothesized that barium modulates arsenic-mediated biological effects, and we examined the effect of barium (1 μM) on arsenic (3 μM)-mediated apoptotic cell death of human HSC-5 and A431 SCC cells in vitro. Arsenic promoted SCC apoptosis with increased reactive oxygen species (ROS) production and JNK1/2 and caspase-3 activation (apoptotic pathway). In contrast, arsenic also inhibited SCC apoptosis with increased NF-κB activity and X-linked inhibitor of apoptosis protein (XIAP) expression level and decreased JNK activity (antiapoptotic pathway). These results suggest that arsenic bidirectionally promotes apoptotic and antiapoptotic pathways in SCC cells. Interestingly, barium in the presence of arsenic increased NF-κB activity and XIAP expression and decreased JNK activity without affecting ROS production, resulting in the inhibition of the arsenic-mediated apoptotic pathway. Since the anticancer effect of arsenic is mainly dependent on cancer apoptosis, barium-mediated inhibition of arsenic-induced apoptosis may promote progression of SCC in patients in Bangladesh who keep drinking barium and arsenic-polluted water after the development of cancer. Thus, we newly showed that barium in the presence of arsenic might inhibit arsenic-mediated cancer apoptosis with the modulation of the balance between arsenic-mediated promotive and suppressive apoptotic pathways.

  18. Upregulation of NRF2 through autophagy/ERK 1/2 ameliorates ionizing radiation induced cell death of human osteosarcoma U-2 OS.

    PubMed

    Chen, Ni; Zhang, Rui; Konishi, Teruaki; Wang, Jun

    2017-01-01

    The antioxidative response mediated by transcription factor NRF2 is thought to be a pivotal cellular defense system against various extrinsic stresses. It has been reported that activation of the NRF2 pathway confers cells with resistance to ionizing radiation-induced damage. However, the underlying mechanism remains largely unknown. In the current research, it was found that α-particle radiation has the ability to stimulate NRF2 expression in human osteosarcoma U-2 OS cells. Knockdown of cellular NRF2 level by shRNA-mediated gene silencing decreased the survival rate, increased the micronucleus formation rate and apoptosis rate in irradiated cells. Consistently, knockdown of NRF2 resulted in decreased expression of p65 and Bcl-2, and increased expression of p53 and Bax. Besides, it was observed that increased expression of NRF2 was partially dependent on radiation induced phosphorylation of ERK 1/2. Further results showed that radiation promoted autophagy flux which leads to the enhanced phosphorylation of ERK 1/2, as evidenced by the resultls that knockdown of ATG5 (Autophagy protein 5) gene by shRNA suppressed both radiation induced ERK 1/2 phosphorylation and NRF2 upregulation. Based on these results, it is proposed that attenuation of NRF2 antioxidative pathway can sensitize U-2 OS cells to radiation, where NRF2 antioxidative response is regulated by autophagy mediated activation of ERK 1/2 kinases.

  19. Distinct roles of Ape1 protein, an enzyme involved in DNA repair, in high or low linear energy transfer ionizing radiation-induced cell killing.

    PubMed

    Wang, Hongyan; Wang, Xiang; Chen, Guangnan; Zhang, Xiangming; Tang, Xiaobing; Park, Dongkyoo; Cucinotta, Francis A; Yu, David S; Deng, Xingming; Dynan, William S; Doetsch, Paul W; Wang, Ya

    2014-10-31

    High linear energy transfer (LET) radiation from space heavy charged particles or a heavier ion radiotherapy machine kills more cells than low LET radiation, mainly because high LET radiation-induced DNA damage is more difficult to repair. Relative biological effectiveness (RBE) is the ratio of the effects generated by high LET radiation to low LET radiation. Previously, our group and others demonstrated that the cell-killing RBE is involved in the interference of high LET radiation with non-homologous end joining but not homologous recombination repair. This effect is attributable, in part, to the small DNA fragments (≤40 bp) directly produced by high LET radiation, the size of which prevents Ku protein from efficiently binding to the two ends of one fragment at the same time, thereby reducing non-homologous end joining efficiency. Here we demonstrate that Ape1, an enzyme required for processing apurinic/apyrimidinic (known as abasic) sites, is also involved in the generation of small DNA fragments during the repair of high LET radiation-induced base damage, which contributes to the higher RBE of high LET radiation-induced cell killing. This discovery opens a new direction to develop approaches for either protecting astronauts from exposure to space radiation or benefiting cancer patients by sensitizing tumor cells to high LET radiotherapy. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Prospects of apoptotic cell-based therapies for transplantation and inflammatory diseases.

    PubMed

    Saas, Philippe; Kaminski, Sandra; Perruche, Sylvain

    2013-10-01

    Apoptotic cell removal or interactions of early-stage apoptotic cells with immune cells are associated with an immunomodulatory microenvironment that can be harnessed to exert therapeutic effects. While the involved immune mechanisms are still being deciphered, apoptotic cell infusion has been tested in different experimental models where inflammation is deregulated. This includes chronic and acute inflammatory disorders such as arthritis, contact hypersensitivity and acute myocardial infarction. Apoptotic cell infusion has also been used in transplantation settings to prevent or treat acute and chronic rejection, as well as to limit acute graft-versus-host disease associated with allogeneic hematopoietic cell transplantation. Here, we review the mechanisms involved in apoptotic cell-induced immunomodulation and data obtained in preclinical models of transplantation and inflammatory diseases.

  1. Concise Review: Apoptotic Cell-Based Therapies-Rationale, Preclinical Results and Future Clinical Developments.

    PubMed

    Saas, Philippe; Daguindau, Etienne; Perruche, Sylvain

    2016-06-01

    The objectives of this review are to summarize the experimental data obtained using apoptotic cell-based therapies, and then to discuss future clinical developments. Indeed, apoptotic cells exhibit immunomodulatory properties that are reviewed here by focusing on more recent mechanisms. These immunomodulatory mechanisms are in particular linked to the clearance of apoptotic cells (called also efferocytosis) by phagocytes, such as macrophages, and the induction of regulatory T cells. Thus, apoptotic cell-based therapies have been used to prevent or treat experimental inflammatory diseases. Based on these studies, we have identified critical steps to design future clinical trials. This includes: the administration route, the number and schedule of administration, the appropriate apoptotic cell type to be used, as well as the apoptotic signal. We also have analyzed the clinical relevancy of apoptotic-cell-based therapies in experimental models. Additional experimental data are required concerning the treatment of inflammatory diseases (excepted for sepsis) before considering future clinical trials. In contrast, apoptotic cells have been shown to favor engraftment and to reduce acute graft-versus-host disease (GvHD) in different relevant models of transplantation. This has led to the conduct of a phase 1/2a clinical trial to alleviate GvHD. The absence of toxic effects obtained in this trial may support the development of other clinical studies based on this new cell therapy. Stem Cells 2016;34:1464-1473. © 2016 AlphaMed Press.

  2. Elastase-mediated phosphatidylserine receptor cleavage impairs apoptotic cell clearance in cystic fibrosis and bronchiectasis

    PubMed Central

    Vandivier, R. William; Fadok, Valerie A.; Hoffmann, Peter R.; Bratton, Donna L.; Penvari, Churee; Brown, Kevin K.; Brain, Joseph D.; Accurso, Frank J.; Henson, Peter M.

    2002-01-01

    Cystic fibrosis is characterized by an early and sustained influx of inflammatory cells into the airways and by release of proteases. Resolution of inflammation is normally associated with the orderly removal of dying apoptotic inflammatory cells through cell recognition receptors, such as the phosphatidylserine receptor, CD36, and αv integrins. Accordingly, removal of apoptotic inflammatory cells may be impaired in persistent inflammatory responses such as that seen in cystic fibrosis airways. Examination of sputa from cystic fibrosis and non–cystic fibrosis bronchiectasis patients demonstrated an abundance of apoptotic cells, in excess of that seen in patients with chronic bronchitis. In vitro, cystic fibrosis and bronchiectasis airway fluid directly inhibited apoptotic cell removal by alveolar macrophages in a neutrophil elastase-dependent manner, suggesting that elastase may impair apoptotic cell clearance in vivo. Flow cytometry demonstrated that neutrophil elastase cleaved the phosphatidylserine receptor, but not CD36 or CD32 (FcγRII). Cleavage of the phosphatidylserine receptor by neutrophil elastase specifically disrupted phagocytosis of apoptotic cells, implying a potential mechanism for delayed apoptotic cell clearance in vivo. Therefore, defective airway clearance of apoptotic cells in cystic fibrosis and bronchiectasis may be due to elastase-mediated cleavage of phosphatidylserine receptor on phagocytes and may contribute to ongoing airway inflammation. PMID:11877474

  3. Pro-apoptotic versus anti-apoptotic properties of dietary resveratrol on tumoral and normal cardiac cells.

    PubMed

    Baarine, Mauhamad; Thandapilly, Sijo Joseph; Louis, Xavier Lieben; Mazué, Frédéric; Yu, Liping; Delmas, Dominique; Netticadan, Thomas; Lizard, Gérard; Latruffe, Norbert

    2011-05-01

    Resveratrol is a natural dietary polyphenol found in grape skin, red wine, and various other food products. Resveratrol has proved to be an effective chemopreventive agent for different malignant tumors. It has also been shown to prevent vascular alterations such as atherosclerosis and inflammatory-associated events. In view of these observations, we investigated the anti-proliferative and pro-apoptotic activities of resveratrol on a tumoral cardiac cell line (HL-1 NB) derived from mouse tumoral atrial cardiac myocytes. These effects were compared with those found on normal neonatal mouse cardiomyocytes. HL-1 NB cells and neonatal cardiomyocytes were treated with resveratrol (5, 30, and/or 100 μM) for different times of culture (24, 48, and/or 72 h). Resveratrol effects were determined by various microscopical and flow cytometric methods. After resveratrol treatment, a strong inhibition of tumoral cardiac HL1-NB cell growth associated with a loss of cell adhesion was observed. This cell proliferation arrest was associated with an apoptotic process revealed by an increased percentage of cells with fragmented and/or condensed nuclei (characteristic of apoptotic cells) identified after staining with Hoechst 33342 and by the presence of cells in subG1. At the opposite, on normal cardiomyocytes, no cytotoxic effects of resveratrol were observed, and a protective effect of resveratrol against norepinephrine-induced apoptosis was found on normal cardiomyocytes. Altogether, the present data demonstrate that resveratrol (1) induces apoptosis of tumoral cardiac HL1-NB cells, (2) does not induce cell death on normal cardiomyocytes, and (3) prevents norepinephrine-induced apoptosis on normal cardiomyocytes.

  4. Gamma-radiation-induced ATM-dependent signalling in human T-lymphocyte leukemic cells, MOLT-4.

    PubMed

    Tichý, Ales; Záskodová, Darina; Rezácová, Martina; Vávrová, Jirina; Vokurková, Doris; Pejchal, Jaroslav; Vilasová, Zdena; Cerman, Jaroslav; Osterreicher, Jan

    2007-01-01

    ATM kinase (ATM) is essential for activation of cell cycle check points and DNA repair in response to ionizing radiation (IR). In this work we studied the molecular mechanisms regulating DNA repair and cell death in human T-lymphocyte leukemic cells, MOLT-4. Apoptosis was evaluated by flow-cytometric detection of annexin V. Early apoptotic cells were determined as sub-G1 cells and late apoptotic cells were determined as APO2.7-positive ones. Proteins involved in ATM signalling pathway were analysed by Western-blotting. We observed a rapid (0.5 h) phosphorylation of ATM declining after 6 h after irradiation by all the doses studied (1.5, 3.0, and 7.5 Gy). Checkpoint kinase-2 (Chk-2) was also phosphorylated after 0.5 h but its phosphorylated form persisted 4, 2, and 1 h after the doses of 1.5, 3.0, and 7.5 Gy, respectively. The amount of p53 protein and its form phosphorylated on Ser-392 increased 1 h after irradiation (1-10 Gy). The lethal dose of 7.5 Gy caused an immediate induction and phosphorylation of p53 after 0.5 h post-irradiation. At the time of phosphorylation of p53, we found simultaneous phosphorylation of the oncoprotein Mdm2 on Ser-166. Neither ATM nor its downstream targets showed a dose-dependent response after 1 h when irradiated by the doses of 1-10 Gy. MOLT-4 cells were very sensitive to the effect of IR. Even low doses, such as 1.5 Gy, induced apoptosis 16 h after irradiation (evaluated according to the cleavage of nuclear lamin B to a 48-kDa fragment). IR-induced molecular signalling after exposure to all the tested doses was triggered by rapid phosphorylation of ATM and Chk-2. Subsequent induction of p53 protein and its phosphorylation was accompanied by concomitant phosphorylation of its negative regulator, oncoprotein Mdm2, and followed by induction of apoptosis.

  5. Arbutin, an intracellular hydroxyl radical scavenger, protects radiation-induced apoptosis in human lymphoma U937 cells.

    PubMed

    Wu, Li-Hua; Li, Peng; Zhao, Qing-Li; Piao, Jin-Lan; Jiao, Yu-Fei; Kadowaki, Makoto; Kondo, Takashi

    2014-11-01

    Ionizing radiation (IR) can generate reactive oxygen species (ROS). Excessive ROS have the potential to damage cellular macromolecules including DNA, proteins, and lipids and eventually lead to cell death. In this study, we evaluated the potential of arbutin, a drug chosen from a series of traditional herbal medicine by measuring intracellular hydroxyl radical scavenging ability in X-irradiated U937 cells. Arbutin (hydroquinone-β-D-glucopyranoside), a naturally occurring glucoside of hydroquinone, has been traditionally used to treat pigmentary disorders. However, there are no reports describing the effect of arbutin on IR-induced apoptosis. We confirmed that arbutin can protect cells from apoptosis induced by X-irradiation. The combination of arbutin and X-irradiation could reduce intracellular hydroxyl radical production and prevent mitochondrial membrane potential loss. It also could down-regulate the expression of phospho-JNK, phospho-p38 in whole cell lysate and activate Bax in mitochondria. Arbutin also inhibits cytochrome C release from mitochondria to cytosol. To verify the role of JNK in X-irradiation-induced apoptosis, the cells were pretreated with a JNK inhibitor, and found that JNK inhibitor could reduce apoptosis induced by X-irradiation. Taken together, our data indicate that arbutin plays an anti-apoptotic role via decreasing intracellular hydroxyl radical production, inhibition of Bax-mitochondria pathway and activation of the JNK/p38 MAPK pathway.

  6. Mitochondrial D310 D-Loop instability and histological subtypes in radiation-induced cutaneous basal cell carcinomas.

    PubMed

    Boaventura, Paula; Pereira, Dina; Mendes, Adélia; Batista, Rui; da Silva, André Ferreira; Guimarães, Isabel; Honavar, Mrinalini; Teixeira-Gomes, José; Lopes, José Manuel; Máximo, Valdemar; Soares, Paula

    2014-01-01

    Basal cell carcinoma (BCC) is the most frequent skin cancer. An elevated prevalence of BCC has been associated with radiation, namely after the Tinea capitis epilation treatment, being these tumors described as more aggressive. Mitochondrial DNA (mtDNA) mutations have been reported in many human tumors, but their occurrence in BCC is poorly documented. The purpose of this work was to evaluate BCC histological subtypes in individuals subjected to X-ray epilation for Tinea capitis treatment when compared to non-irradiated patients. Moreover we also wanted to evaluate mitochondrial D-Loop instability in both groups of BCCs in order to compare the frequency of D-Loop mutations in post-irradiation BCC versus sporadic BCC. 228 histological specimens corresponding to BCCs from 75 irradiated patients and 60 non-irradiated patients were re-evaluated for histological subtype. Subsequently, we sequenced the D-Loop 310 repeat in blood, oral mucosa, tumor lesions and, whenever available, non-tumoral adjacent tissue from these patients. The infiltrative subtype of BCC, considered to be more aggressive, was significantly more frequent in irradiated patients. BCC D-Loop D310 mutation rate was significantly higher in irradiated BCCs than in the non-irradiated ones. Moreover, it was associated with a higher irradiation dose. The presence of mtDNA heteroplasmy in patients' blood was associated with a higher mutation rate in the BCCs suggesting that a more unstable genotype could predispose to mtDNA somatic mutation. Our results suggest that radiation-induced BCCs may be considered to be more aggressive tumors. Further studies are needed to clarify the role of mtDNA D-Loop mutations in tumors from irradiated patients. Copyright © 2013 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  7. A fluorescent toolkit for spatiotemporal tracking of apoptotic cells in living Drosophila tissues.

    PubMed

    Schott, Sonia; Ambrosini, Arnaud; Barbaste, Audrey; Benassayag, Corinne; Gracia, Mélanie; Proag, Amsha; Rayer, Mégane; Monier, Bruno; Suzanne, Magali

    2017-09-04

    Far from being passive, apoptotic cells influence their environment. For instance, they promote tissue folding, myoblast fusion and modulate tumor growth. Understanding the role of apoptotic cells necessitates their efficient tracking within living tissues, a task which is currently challenging. In order to easily spot apoptotic cells in developing Drosophila tissues, we generated a series of fly lines expressing different fluorescent sensors of caspase activity. We show that three of these reporters (GFP, Cerulean and Venus derived molecules) are detected specifically in apoptotic cells and throughout the whole process of programmed cell death. These reporters allow the specific visualization of apoptotic cells directly within living tissues, without any post-acquisition processing. They overcome the limitations of other apoptosis detection methods developed so far and notably, they can be combined with any kind of fluorophore. © 2017. Published by The Company of Biologists Ltd.

  8. Defining the optimal window for cranial transplantation of human induced pluripotent stem cell-derived cells to ameliorate radiation-induced cognitive impairment.

    PubMed

    Acharya, Munjal M; Martirosian, Vahan; Christie, Lori-Ann; Riparip, Lara; Strnadel, Jan; Parihar, Vipan K; Limoli, Charles L

    2015-01-01

    Past preclinical studies have demonstrated the capability of using human stem cell transplantation in the irradiated brain to ameliorate radiation-induced cognitive dysfunction. Intrahippocampal transplantation of human embryonic stem cells and human neural stem cells (hNSCs) was found to functionally restore cognition in rats 1 and 4 months after cranial irradiation. To optimize the potential therapeutic benefits of human stem cell transplantation, we have further defined optimal transplantation windows for maximizing cognitive benefits after irradiation and used induced pluripotent stem cell-derived hNSCs (iPSC-hNSCs) that may eventually help minimize graft rejection in the host brain. For these studies, animals given an acute head-only dose of 10 Gy were grafted with iPSC-hNSCs at 2 days, 2 weeks, or 4 weeks following irradiation. Animals receiving stem cell grafts showed improved hippocampal spatial memory and contextual fear-conditioning performance compared with irradiated sham-surgery controls when analyzed 1 month after transplantation surgery. Importantly, superior performance was evident when stem cell grafting was delayed by 4 weeks following irradiation compared with animals grafted at earlier times. Analysis of the 4-week cohort showed that the surviving grafted cells migrated throughout the CA1 and CA3 subfields of the host hippocampus and differentiated into neuronal (∼39%) and astroglial (∼14%) subtypes. Furthermore, radiation-induced inflammation was significantly attenuated across multiple hippocampal subfields in animals receiving iPSC-hNSCs at 4 weeks after irradiation. These studies expand our prior findings to demonstrate that protracted stem cell grafting provides improved cognitive benefits following irradiation that are associated with reduced neuroinflammation.

  9. Increased sensitivity of early apoptotic cells to complement-mediated lysis.

    PubMed

    Attali, Gitit; Gancz, Dana; Fishelson, Zvi

    2004-11-01

    Opsonization of apoptotic cells with complement proteins contributes to their clearance by phagocytes. Little is known about the lytic effects of complement on apoptotic cells. Sensitivity of cells treated with anti-Fas antibody (Jurkat cells), staurosporine or etoposide (Raji cells) to lysis by complement was examined. As shown here, early apoptotic cells are more sensitive to lysis by antibody and complement than control cells. More complement C3 and C9 bound to apoptotic than to control cells, even though antibody binding was similar. Enhanced killing and C3/C9 deposition were blocked by benzyloxy-Val-Ala-Asp-fluoromethylketone, a pan-caspase inhibitor. Complement-mediated lysis of early apoptotic cells was also prevented by inhibitors of caspases 6, 8, 9 or 10. In contrast, caspase inhibitors had no effect on the lysis of non-apoptotic Jurkat and Raji cells. Early apoptotic Jurkat cells were also more sensitive to lysis by the pore formers streptolysin O and melittin. Sensitivity of Jurkat Bcl-2 transfectants to lysis by complement was analyzed. Enhanced Bcl-2 expression was associated with reduced C3 deposition and lower sensitivity to complement-mediated lysis. These results demonstrate that at an early stage in apoptosis, following caspase activation, cells become sensitive to necrotic-type death by complement and other pore formers. Furthermore, they suggest that Bcl-2 is actively protecting Jurkat cells from complement-mediated lysis.

  10. Amelioration of radiation-induced skin injury by adenovirus-mediated heme oxygenase-1 (HO-1) overexpression in rats

    PubMed Central

    2012-01-01

    Objective Radiation-induced skin injury remains a serious concern for radiation therapy. Heme oxygenase-1 (HO-1), the rate-limiting enzyme in heme catabolism, has been reported to have potential antioxidant and anti-apoptotic properties. However, the role of HO-1 in radiation-induced skin damage remains unclear. This study aims to elucidate the effects of HO-1 on radiation-induced skin injury in rats. Methods A control adenovirus (Ad-EGFP) and a recombinant adenovirus (Ad-HO1-EGFP) were constructed. Rats were irradiated to the buttock skin with a single dose of 45 Gy followed by a subcutaneous injection of PBS, 5 × 109 genomic copies of Ad-EGFP or Ad-HO1-EGFP (n = 8). After treatment, the skin MDA concentration, SOD activity and apoptosis were measured. The expression of antioxidant and pro-apoptotic genes was determined by RT-PCR and real-time PCR. Skin reactions were measured at regular intervals using the semi-quantitative skin injury score. Results Subcutaneous injection of Ad-HO1-EGFP infected both epidermal and dermal cells and could spread to the surrounding regions. Radiation exposure upregulated the transcription of the antioxidant enzyme genes, including SOD-1, GPx2 and endogenous HO-1. HO-1 overexpression decreased lipid peroxidation and inhibited the induction of ROS scavenging proteins. Moreover, HO-1 exerted an anti-apoptotic effect by suppressing FAS and FASL expression. Subcutaneous injection of Ad-HO1-EGFP demonstrated significant improvement in radiation-induced skin injury. Conclusions The present study provides evidences for the protective role of HO-1 in alleviating radiation-induced skin damage in rats, which is helpful for the development of therapy for radiation-induced skin injury. PMID:22247972

  11. [Direct assay of radiation-induced DNA base lesions to mammalian cells]. Final progress report, September 1, 1991--November 1, 1993

    SciTech Connect

    Not Available

    1993-12-31

    We have successfully developed the GC/MS technique so that an assessment of base damage in mammalian cells can be accomplished. The technique now has a sensitivity that will allow one to perform research in the low dose region suitable for hazards evaluation. The research on the hydrated DNA molecule has been seminal in generating a better understanding of the mechanisms by which low LET radiation induces DNA damage in mammalian cells. Also reported here are (1) the methodology for hydrating and irradiating DNA has been developed, (2) the procedures for identifying and quantitating radiation-induced DNA damage by HPLC and GC/MS have been mastered, (3) an hypotheses that radiation-induced damage in closely associated water molecules can result in DNA damage which is indistinguishable from that caused by direct ionization of the DNA has been generated and supported by experimental data, and (4) mathematical expressions that relate DNA lesion formation to the important parameters in the above hypotheses have been constructed so that the predictions of the hypotheses can now be tested.

  12. The apoptotic effect of simvastatin via the upregulation of BIM in nonsmall cell lung cancer cells.

    PubMed

    Lee, Hwa Young; Kim, In Kyoung; Lee, Hye In; Mo, Jin Young; Yeo, Chang Dong; Kang, Hyeon Hui; Moon, Hwa Sik; Lee, Sang Haak

    2016-01-01

    Statins are known to have pleiotropic effects that induce cell death in certain cancer cells. BIM is a member of the bcl-2 gene family, which promotes apoptotic cell death. This study investigated the hypothesis that simvastatin has pro-apoptotic effects in epidermal growth factor receptor (EGFR)-mutated lung cancer cell lines via the upregulation of the expression of the BIM protein. The cytotoxic effects of simvastatin on gefitinib-sensitive (HCC827, E716-A750del) and -resistant (H1975, T790M + L858R) nonsmall cell lung cancer (NSCLC) cells were compared. Cell proliferation and expression of apoptosis-related and EGFR downstream signaling proteins were evaluated. Expression of BIM was compared in H1975 cells after treatment with simvastatin or gefitinib. SiRNA-mediated BIM depletion was performed to confirm whether the cytotoxicity of simvastatin was mediated by the expression of BIM. H1975 cells showed significantly reduced viability compared with HCC827 cells after treatment with simvastatin (2 μM) for 48 hours. In simvastatin-treated H1975 cells, expression of pro-apoptotic proteins was increased and the phosphorylation of ERK 1/2 (p-ERK 1/2) was reduced. Expression of BIM was suppressed by gefitinib (1 μM) treatment in H1975 cells, but it was significantly increased by treatment with simvastatin. BIM depletion by siRNA transfection enhanced the viability of H1975 cells that received simvastatin treatment and increased their expression of anti-apoptotic proteins. Simvastatin restored the expression of BIM to induce apoptotic cell death in NSCLC cells harboring an EGFR-resistant mutation. Our study suggests the potential utility of simvastatin as a BIM-targeted treatment for NSCLC.

  13. Radiation-induced gliomas

    PubMed Central

    Prasad, Gautam; Haas-Kogan, Daphne A.

    2013-01-01

    Radiation-induced gliomas represent a relatively rare but well-characterized entity in the neuro-oncologic literature. Extensive retrospective cohort data in pediatric populations after therapeutic intracranial radiation show a clearly increased risk in glioma incidence that is both patient age- and radiation dose/volume-dependent. Data in adults are more limited but show heightened risk in certain groups exposed to radiation. In both populations, there is no evidence linking increased risk associated with routine exposure to diagnostic radiation. At the molecular level, recent studies have found distinct genetic differences between radiation-induced gliomas and their spontaneously-occurring counterparts. Clinically, there is understandable reluctance on the part of clinicians to re-treat patients due to concern for cumulative neurotoxicity. However, available data suggest that aggressive intervention can lead to improved outcomes in patients with radiation-induced gliomas. PMID:19831840

  14. Radiation-induced mitotic cell death and glioblastoma radioresistance: a new regulating pathway controlled by integrin-linked kinase, hypoxia-inducible factor 1 alpha and survivin in U87 cells.

    PubMed

    Lanvin, Olivia; Monferran, Sylvie; Delmas, Caroline; Couderc, Bettina; Toulas, Christine; Cohen-Jonathan-Moyal, Elizabeth

    2013-09-01

    We have previously shown that integrin-linked kinase (ILK) regulates U87 glioblastoma cell radioresistance by modulating the main radiation-induced cell death mechanism in solid tumours, the mitotic cell death. To decipher the biological pathways involved in these mechanisms, we constructed a U87 glioblastoma cell model expressing an inducible shRNA directed against ILK (U87shILK). We then demonstrated that silencing ILK enhanced radiation-induced centrosome overduplication, leading to radiation-induced mitotic cell death. In this model, ionising radiations induce hypoxia-inducible factor 1 alpha (HIF-1α) stabilisation which is inhibited by silencing ILK. Moreover, silencing HIF-1α in U87 cells reduced the surviving fraction after 2 Gy irradiation by increasing cell sensitivity to radiation-induced mitotic cell death and centrosome amplification. Because it is known that HIF-1α controls survivin expression, we then looked at the ILK silencing effect on survivin expression. We show that survivin expression is decreased in U87shILK cells. Furthermore, treating U87 cells with the specific survivin suppressor YM155 significantly increased the percentage of giant multinucleated cells, centrosomal overduplication and thus U87 cell radiosensitivity. In consequence, we decipher here a new pathway of glioma radioresistance via the regulation of radiation-induced centrosome duplication and therefore mitotic cell death by ILK, HIF-1α and survivin. This work identifies new targets in glioblastoma with the intention of radiosensitising these highly radioresistant tumours. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. Radiation-induced genomic instability

    NASA Technical Reports Server (NTRS)

    Kronenberg, A.

    1994-01-01

    Quantitative assessment of the heritable somatic effects of ionizing radiation exposures has relied upon the assumption that radiation-induced lesions were 'fixed' in the DNA prior to the first postirradiation mitosis. Lesion conversion was thought to occur during the initial round of DNA replication or as a consequence of error-prone enzymatic processing of lesions. The standard experimental protocols for the assessment of a variety of radiation-induced endpoints (cell death, specific locus mutations, neoplastic transformation and chromosome aberrations) evaluate these various endpoints at a single snapshot in time. In contrast with the aforementioned approaches, some studies have specifically assessed radiation effects as a function of time following exposure. Evidence has accumulated in support of the hypothesis that radiation exposure induces a persistent destabilization of the genome. This instability has been observed as a delayed expression of lethal mutations, as an enhanced rate of accumulation of non-lethal heritable alterations, and as a progressive intraclonal chromosomal heterogeneity. The genetic controls and biochemical mechanisms underlying radiation-induced genomic instability have not yet been delineated. The aim is to integrate the accumulated evidence that suggests that radiation exposure has a persistent effect on the stability of the mammalian genome.

  16. Radiation-induced genomic instability

    NASA Technical Reports Server (NTRS)

    Kronenberg, A.

    1994-01-01

    Quantitative assessment of the heritable somatic effects of ionizing radiation exposures has relied upon the assumption that radiation-induced lesions were 'fixed' in the DNA prior to the first postirradiation mitosis. Lesion conversion was thought to occur during the initial round of DNA replication or as a consequence of error-prone enzymatic processing of lesions. The standard experimental protocols for the assessment of a variety of radiation-induced endpoints (cell death, specific locus mutations, neoplastic transformation and chromosome aberrations) evaluate these various endpoints at a single snapshot in time. In contrast with the aforementioned approaches, some studies have specifically assessed radiation effects as a function of time following exposure. Evidence has accumulated in support of the hypothesis that radiation exposure induces a persistent destabilization of the genome. This instability has been observed as a delayed expression of lethal mutations, as an enhanced rate of accumulation of non-lethal heritable alterations, and as a progressive intraclonal chromosomal heterogeneity. The genetic controls and biochemical mechanisms underlying radiation-induced genomic instability have not yet been delineated. The aim is to integrate the accumulated evidence that suggests that radiation exposure has a persistent effect on the stability of the mammalian genome.

  17. Cell-permeable intrinsic cellular inhibitors of apoptosis protect and rescue intestinal epithelial cells from radiation-induced cell death.

    PubMed

    Matsuzaki-Horibuchi, Shiori; Yasuda, Takeshi; Sakaguchi, Nagako; Yamaguchi, Yoshihiro; Akashi, Makoto

    2015-01-01

    One of the important mechanisms for gastrointestinal (GI) injury following high-dose radiation exposure is apoptosis of epithelial cells. X-linked inhibitor of apoptosis (XIAP) and cellular IAP2 (cIAP2) are intrinsic cellular inhibitors of apoptosis. In order to study the effects of exogenously added IAPs on apoptosis in intestinal epithelial cells, we constructed bacterial expression plasmids containing genes of XIAP (full-length, BIR2 domain and BIR3-RING domain with and without mutations of auto-ubiquitylation sites) and cIAP2 proteins fused to a protein-transduction domain (PTD) derived from HIV-1 Tat protein (TAT) and purified these cell-permeable recombinant proteins. When the TAT-conjugated IAPs were added to rat intestinal epithelial cells IEC6, these proteins were effectively delivered into the cells and inhibited apoptosis, even when added after irradiation. Our results suggest that PTD-mediated delivery of IAPs may have clinical potential, not only for radioprotection but also for rescuing the GI system from radiation injuries.

  18. Cell-permeable intrinsic cellular inhibitors of apoptosis protect and rescue intestinal epithelial cells from radiation-induced cell death

    PubMed Central

    Matsuzaki-Horibuchi, Shiori; Yasuda, Takeshi; Sakaguchi, Nagako; Yamaguchi, Yoshihiro; Akashi, Makoto

    2015-01-01

    One of the important mechanisms for gastrointestinal (GI) injury following high-dose radiation exposure is apoptosis of epithelial cells. X-linked inhibitor of apoptosis (XIAP) and cellular IAP2 (cIAP2) are intrinsic cellular inhibitors of apoptosis. In order to study the effects of exogenously added IAPs on apoptosis in intestinal epithelial cells, we constructed bacterial expression plasmids containing genes of XIAP (full-length, BIR2 domain and BIR3-RING domain with and without mutations of auto-ubiquitylation sites) and cIAP2 proteins fused to a protein-transduction domain (PTD) derived from HIV-1 Tat protein (TAT) and purified these cell-permeable recombinant proteins. When the TAT-conjugated IAPs were added to rat intestinal epithelial cells IEC6, these proteins were effectively delivered into the cells and inhibited apoptosis, even when added after irradiation. Our results suggest that PTD-mediated delivery of IAPs may have clinical potential, not only for radioprotection but also for rescuing the GI system from radiation injuries. PMID:25359904

  19. Apoptotic cell death is initiated during normothermic ischemia in human kidneys.

    PubMed

    Wolfs, Tim G A M; de Vries, Bart; Walter, Sarah J; Peutz-Kootstra, Carine J; van Heurn, L W Ernest; Oosterhof, Gosse O N; Buurman, Wim A

    2005-01-01

    Ischemic damage plays an important role in post-transplant organ failure. Activation of the apoptotic cascade is crucially involved in post-ischemic inflammation resulting in tissue damage and organ dysfunction. Here we investigate the initiation of the apoptotic cascade during normothermic ischemia in human kidneys using a model for normothermic ischemia with kidneys nephrectomized because of renal cell carcinoma. Ex vivo, kidneys were stored at 37 degrees C, and consecutive biopsies were taken from disease-free tissue. Pro- and anti-apoptotic proteins were assessed by Western blotting and immunofluorescence. During normothermic ischemia the pro-apoptotic proteins Bax and activated caspase-9 increased with ischemia time, whereas caspase-8 was not activated. The anti-apoptotic proteins Bcl-2 and cFLIP decreased in time. Data on Bcl-2 and Bax were supported by immunofluorescence for Bcl-2 and activated Bax. However, activation of the central effector caspase-3, essential for execution of the apoptotic process, was not detected. In conclusion, during normothermic ischemia the apoptotic cascade in the human kidney is initiated, but not fulfilled. Our data show that the duration of ischemia significantly correlates with activation of the apoptotic cascade. These findings provide insight in the initiation of apoptotic cell-death during warm ischemia and may be useful in the assessment of ischemic injury.

  20. [Ionizing radiation-induced DNA damage and its repair in human cells]. Progress report, [April 1, 1993--February 28, 1994

    SciTech Connect

    Not Available

    1994-07-01

    The excision of radiation-induced lesions in DNA by a DNA repair enzyme complex, namely the UvrABC nuclease complex, has been investigated. Irradiated DNA was treated with the enzyme complex. DNA fractions were analyzed by gas chromatography/isotope-dilution mass spectrometry. The results showed that a number pyrimidine- and purine-derived lesions in DNA were excised by the UvrABC nuclease complex and that the enzyme complex does not act on radiation-induced DNA lesions as a glycosylase. This means that it does not excise individual base products, but it excises oligomers containing these lesions. A number of pyrimidine-derived lesions that were no substrates for other DNA repair enzymes investigated in our laboratory were substrates for the UvrABC nuclease complex.

  1. Radiation-induced pneumothorax

    SciTech Connect

    Epstein, D.M.; Littman, P.; Gefter, W.B.; Miller, W.T.; Raney, R.B. Jr.

    1983-01-01

    Pneumothorax is an uncommon complication of radiation therapy to the chest. The proposed pathogenesis is radiation-induced fibrosis promoting subpleural bleb formation that ruptures resulting in pneumothorax. We report on two young patients with primary sarcomas without pulmonary metastases who developed spontaneous pneumothorax after irradiation. Neither patient had antecedent radiographic evidence of pulmonary fibrosis.

  2. Gingerol sensitizes TRAIL-induced apoptotic cell death of glioblastoma cells

    SciTech Connect

    Lee, Dae-Hee; Kim, Dong-Wook; Jung, Chang-Hwa; Lee, Yong J.; Park, Daeho

    2014-09-15

    Glioblastoma multiforme (GBM) is the most lethal and aggressive astrocytoma of primary brain tumors in adults. Although there are many clinical trials to induce the cell death of glioblastoma cells, most glioblastoma cells have been reported to be resistant to TRAIL-induced apoptosis. Here, we showed that gingerol as a major component of ginger can induce TRAIL-mediated apoptosis of glioblastoma. Gingerol increased death receptor (DR) 5 levels in a p53-dependent manner. Furthermore, gingerol decreased the expression level of anti-apoptotic proteins (survivin, c-FLIP, Bcl-2, and XIAP) and increased pro-apoptotic protein, Bax and truncate Bid, by generating reactive oxygen species (ROS). We also found that the sensitizing effects of gingerol in TRAIL-induced cell death were blocked by scavenging ROS or overexpressing anti-apoptotic protein (Bcl-2). Therefore, we showed the functions of gingerol as a sensitizing agent to induce cell death of TRAIL-resistant glioblastoma cells. This study gives rise to the possibility of applying gingerol as an anti-tumor agent that can be used for the purpose of combination treatment with TRAIL in TRAIL-resistant glioblastoma tumor therapy. - Highlights: • Most GBM cells have been reported to be resistant to TRAIL-induced apoptosis. • Gingerol enhances the expression level of anti-apoptotic proteins by ROS. • Gingerol enhances TRAIL-induced apoptosis through actions on the ROS–Bcl2 pathway.

  3. The Use of Lectin Histochemistry for Detecting Apoptotic Cells in the Seminiferous Epithelium.

    PubMed

    Seco-Rovira, Vicente; Beltrán-Frutos, Ester; Hernández-Martínez, Jesús; Ferrer, Concepción; Pastor, Luis Miguel

    2017-01-01

    Lectin histochemistry is commonly used to characterize the pattern of glycoconjugates in cells and tissues. Recent studies show that alterations in these glycoconjugates are associated with the entry of cells into apoptosis. A widely used technique for the detection of apoptotic cell death is TUNEL. In this chapter, we study the sensitivity of both techniques to identify apoptotic cells in the testis of photo-inhibited Syrian hamster.

  4. Ceramide synthases 2, 5, and 6 confer distinct roles in radiation-induced apoptosis in HeLa cells.

    PubMed

    Mesicek, Judith; Lee, Hyunmi; Feldman, Taya; Jiang, Xuejun; Skobeleva, Anastasia; Berdyshev, Evgeny V; Haimovitz-Friedman, Adriana; Fuks, Zvi; Kolesnick, Richard

    2010-09-01

    The role of ceramide neo-genesis in cellular stress response signaling is gaining increasing attention with recent progress in elucidating the novel roles and biochemical properties of the ceramide synthase (CerS) enzymes. Selective tissue and subcellular distribution of the six mammalian CerS isoforms, combined with distinct fatty acyl chain length substrate preferences, implicate differential functions of specific ceramide species in cellular signaling. We report here that ionizing radiation (IR) induces de novo synthesis of ceramide to influence HeLa cell apoptosis by specifically activating CerS isoforms 2, 5, and 6 that generate opposing anti- and pro-apoptotic ceramides in mitochondrial membranes. Overexpression of CerS2 resulted in partial protection from IR-induced apoptosis whereas overexpression of CerS5 increased apoptosis in HeLa cells. Knockdown studies determined that CerS2 is responsible for all observable IR-induced C(24:0) CerS activity, and while CerS5 and CerS6 each confer approximately 50% of the C(16:0) CerS baseline synthetic activity, both are required for IR-induced activity. Additionally, co-immunoprecipitation studies suggest that CerS2, 5, and 6 might exist as heterocomplexes in HeLa cells, providing further insight into the regulation of CerS proteins. These data add to the growing body of evidence demonstrating interplay among the CerS proteins in a stress stimulus-, cell type- and subcellular compartment-specific manner.

  5. Measurement of phagocytic engulfment of apoptotic cells by macrophages using pHrodo succinimidyl ester.

    PubMed

    Aziz, Monowar; Yang, Weng-Lang; Wang, Ping

    2013-01-01

    Considerable interest has emerged towards phagocytosis of apoptotic cells, due to its intricate molecular mechanisms and important regulatory functions in development, homoeostasis, and immune tolerance. Impaired clearance of apoptotic cells leads to immune-mediated disorders. Current quantification methods of the engulfment of apoptotic cells by macrophages are potentially flawed by several limitations. Adherent macrophage populations are overlaid with apoptotic targets in suspension and then co-cultured for a definite period, which may give rise to two different features: (1) engulfed and (2) non-engulfed macrophages that are surface-bound cell populations. Rigorous washing to dislodge surface-bound apoptotic cells before assessment of phagocytosis may lead to loss of phagocytes, thereby skewing the apparent magnitude of the overall phagocytic response. There is a need for simple and reliable methods to clearly determine the internalization of apoptotic cells. In this unit, we demonstrate the use of pHrodo-succinimidyl ester (SE), a pH-sensitive fluorescent dye, to label the apoptotic cells for monitoring the phagocytosis. After engulfment, the intensity of pHrodo light emission will be elevated due to the pH change inside of macrophages. The shift of pHrodo light emission can be detected by a flow cytometer or using a fluorescence microscope.

  6. Apoptotic cells trigger a membrane-initiated pathway to increase ABCA1

    PubMed Central

    Fond, Aaron M.; Lee, Chang Sup; Schulman, Ira G.; Kiss, Robert S.; Ravichandran, Kodi S.

    2015-01-01

    Macrophages clear millions of apoptotic cells daily and, during this process, take up large quantities of cholesterol. The membrane transporter ABCA1 is a key player in cholesterol efflux from macrophages and has been shown via human genetic studies to provide protection against cardiovascular disease. How the apoptotic cell clearance process is linked to macrophage ABCA1 expression is not known. Here, we identified a plasma membrane–initiated signaling pathway that drives a rapid upregulation of ABCA1 mRNA and protein. This pathway involves the phagocytic receptor brain-specific angiogenesis inhibitor 1 (BAI1), which recognizes phosphatidylserine on apoptotic cells, and the intracellular signaling intermediates engulfment cell motility 1 (ELMO1) and Rac1, as ABCA1 induction was attenuated in primary macrophages from mice lacking these molecules. Moreover, this apoptotic cell–initiated pathway functioned independently of the liver X receptor (LXR) sterol–sensing machinery that is known to regulate ABCA1 expression and cholesterol efflux. When placed on a high-fat diet, mice lacking BAI1 had increased numbers of apoptotic cells in their aortic roots, which correlated with altered lipid profiles. In contrast, macrophages from engineered mice with transgenic BAI1 overexpression showed greater ABCA1 induction in response to apoptotic cells compared with those from control animals. Collectively, these data identify a membrane-initiated pathway that is triggered by apoptotic cells to enhance ABCA1 within engulfing phagocytes and with functional consequences in vivo. PMID:26075824

  7. The Role of Target and Bystander Cells in Dose-Response Relationship of Radiation-Induced Bystander Effects in Two Cell Lines

    PubMed Central

    Soleymanifard, Shokouhozaman; Bahreyni Toossi, Mohammad Taghi; Sazgarnia, Ameneh; Mohebbi, Shokoufe

    2013-01-01

    Objective(s): Radiation effect induced in nonirradiated cells which are adjacent or far from irradiated cells is termed radiation-induced bystander effect (RIBE). Published data on dose-response relationship of RIBE is controversial. In the present study the role of targeted and bystander cells in RIBE dose-response relationship of two cell lines have been investigated. Materials and Methods: Two cell lines (QU-DB and MRC5) which had previously exhibited different dose-response relationship were selected. In the previous study the two cell lines received medium from autologous irradiated cells and the results showed that the magnitude of damages induced in QU-DB cells was dependent on dose unlike MRC5 cells. In the present study, the same cells irradiated with 0.5, 2 and 4 Gy gamma rays and their conditioned media were transferred to nonautologous bystander cells; such that the bystander effects due to cross-interaction between them were studied. Micronucleus assay was performed to measure the magnitude of damages induced in bystander cells (RIBE level). Results: QU-DB cells exhibited a dose-dependent response. RIBE level in MRC5 cells which received medium from 0.5 and 2 Gy QU-DB irradiated cells was not statistically different, but surprisingly when they received medium from 4Gy irradiated QU-DB cells, RIBE was abrogated. Conclusion: Results pertaining to QU-DB and MRC5 cells indicated that both target and bystander cells determined the outcome. Triggering the bystander effect depended on the radiation dose and the target cell-type, but when RIBE was triggered, dose-response relationship was predominantly determined by the bystander cell type. PMID:24298387

  8. Adipose Mesenchymal Stem Cell Secretome Modulated in Hypoxia for Remodeling of Radiation-Induced Salivary Gland Damage

    PubMed Central

    An, Hye-Young; Shin, Hyun-Soo; Choi, Jeong-Seok; Kim, Hun Jung

    2015-01-01

    Background and Purpose This study was conducted to determine whether a secretome from mesenchymal stem cells (MSC) modulated by hypoxic conditions to contain therapeutic factors contributes to salivary gland (SG) tissue remodeling and has the potential to improve irradiation (IR)-induced salivary hypofunction in a mouse model. Materials and Methods Human adipose mesenchymal stem cells (hAdMSC) were isolated, expanded, and exposed to hypoxic conditions (O2 < 5%). The hypoxia-conditioned medium was then filtered to a high molecular weight fraction and prepared as a hAdMSC secretome. The hAdMSC secretome was subsequently infused into the tail vein of C3H mice immediately after local IR once a day for seven consecutive days. The control group received equal volume (500 μL) of vehicle (PBS) only. SG function and structural tissue remodeling by the hAdMSC secretome were investigated. Human parotid epithelial cells (HPEC) were obtained, expanded in vitro, and then irradiated and treated with either the hypoxia-conditioned medium or a normoxic control medium. Cell proliferation and IR-induced cell death were examined to determine the mechanism by which the hAdMSC secretome exerted its effects. Results The conditioned hAdMSC secretome contained high levels of GM-CSF, VEGF, IL-6, and IGF-1. Repeated systemic infusion with the hAdMSC secretome resulted in improved salivation capacity and increased levels of salivary proteins, including amylase and EGF, relative to the PBS group. The microscopic structural integrity of SG was maintained and salivary epithelial (AQP-5), endothelial (CD31), myoepithelial (α-SMA) and SG progenitor cells (c-Kit) were successfully protected from radiation damage and remodeled. The hAdMSC secretome strongly induced proliferation of HPEC and led to a significant decrease in cell death in vivo and in vitro. Moreover, the anti-apoptotic effects of the hAdMSC secretome were found to be promoted after hypoxia-preconditioning relative to normoxia

  9. Bioactive compounds from crocodile (Crocodylus siamensis) white blood cells induced apoptotic cell death in hela cells.

    PubMed

    Patathananone, Supawadee; Thammasirirak, Sompong; Daduang, Jureerut; Chung, Jing Gung; Temsiripong, Yosapong; Daduang, Sakda

    2016-08-01

    Crocodile (Crocodylus siamensis) white blood cell extracts (WBCex) were examined for anticancer activity in HeLa cell lines using the MTT assay. The percentage viability of HeLa cells significantly deceased after treatment with WBCex in a dose- and time-dependent manner. The IC50 dose was suggested to be approximately 225 μg/mL protein. Apoptotic cell death occurred in a time-dependent manner based on investigation by flow cytometry using annexin V-FITC and PI staining. DAPI nucleic acid staining indicated increased chromatin condensation. Caspase-3, -8 and -9 activities also increased, suggesting the induction of the caspase-dependent apoptotic pathway. Furthermore, the mitochondrial membrane potential (ΔΨm ) of HeLa cells was lost as a result of increasing levels of Bax and reduced levels of Bcl-2, Bcl-XL, Bcl-Xs, and XIAP. The decreased ΔΨm led to the release of cytochrome c and the activation of caspase-9 and -3. Apoptosis-inducing factor translocated into the nuclei, and endonuclease G (Endo G) was released from the mitochondria. These results suggest that anticancer agents in WBCex can induce apoptosis in HeLa cells via both caspase-dependent and -independent pathways. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 986-997, 2016. © 2015 Wiley Periodicals, Inc.

  10. Antiproliferative and apoptotic effects of diffractaic acid in A549 and AGS cancer cells

    NASA Astrophysics Data System (ADS)

    Kızıl, Hamit Emre; Aǧar, Güleray

    2017-04-01

    In this study, we determined the antiproliferative and apoptotic effects of diffractaic acid by measuring the gene expression changes of topo II α, caspase-3 and p53 on A549 and AGS cancer cells. Real time PCR assay was used to measure the change folds. It was determined that concentrations of 12,5, 50 and 100 µg / ml were antiproliferative and apoptotic for the A549 cancer cell line and 50 µg / ml for the AGS cell line.

  11. Effects of culture media on the susceptibility of cells to apoptotic cell death.

    PubMed

    Anai, Chikara; Kawaguchi, Masatoshi; Eto, Ko

    2014-09-01

    Whether responses of cells to extracellular environments affect the induction of apoptotic cell death is poorly understood. The current study aimed to unravel the different effects of culture media employed in vitro as extracellular environments on the susceptibility of cells to apoptosis. We found that apoptosis is stimulated to the higher levels by culturing human HeLa cells in Opti-MEM with unknown components, a medium that is specifically used for transfections, than by culturing cells in Dulbecco's modified Eagle's medium, a medium that is generally used for maintenance of cells. We showed that apoptosis is suppressed partially by culturing cells in heat-treated Opti-MEM, implicating a heat-sensitive component(s) in stimulating the apoptotic response of cells. Thus, different extracellular environments may contribute to different responses of cells to apoptosis, and this should be considered to evaluate the incidences of apoptotic cell death and could be applied to develop an efficient treatment for curing diseases such as cancer.

  12. Inhibition of radiation-induced apoptosis by dexamethasone in cervical carcinoma cell lines depends upon increased HPV E6/E7

    PubMed Central

    Kamradt, M C; Mohideen, N; Krueger, E; Walter, S; Vaughan, A T M

    2000-01-01

    Through a glucocorticoid-responsive promoter, glucocorticoids can regulate the transcription of the human papillomavirus (HPV) E6 and E7 viral genes which target the tumour suppressor proteins p53 and Rb respectively. In C4-1 cells, the glucocorticoid dexamethasone up-regulated HPV E6/E7 mRNA and decreased radiation-induced apoptosis. In contrast, dexamethasone had no effect on apoptosis of cells that either lack the HPV genome (C33-a) or in which HPV E6/E7 transcription is repressed by dexamethasone (SW756). Irradiated C4-1 cells showed increased p53 expression, while dexamethasone treatment prior to irradiation decreased p53 protein expression. In addition, p21 mRNA was regulated by irradiation and dexamethasone in accordance with the observed changes in p53. Overall, glucocorticoids decreased radiation-induced apoptosis in cervical carcinoma cells which exhibit increased HPV E6/E7 transcription and decreased p53 expression. Therefore, in HPV-infected cervical epithelial cells, p53-dependent apoptosis appears to depend upon the levels of HPV E6/E7 mRNA. © 2000 Cancer Research Campaign PMID:10817508

  13. Phototherapy-treated apoptotic tumor cells induce pro-inflammatory cytokines production in macrophage

    NASA Astrophysics Data System (ADS)

    Lu, Cuixia; Wei, Yanchun; Xing, Da

    2014-09-01

    Our previous studies have demonstrated that as a mitochondria-targeting cancer phototherapy, high fluence low-power laser irradiation (HF-LPLI) induces mitochondrial superoxide anion burst, resulting in oxidative damage to tumor cells. In this study, we further explored the immunological effects of HF-LPLI-induced apoptotic tumor cells. When macrophages were co-incubated with apoptotic cells induced by HF-LPLI, we observed the increased levels of TNF-α secretion and NO production in macrophages. Further experiments showed that NF-κB was activated in macrophages after co-incubation with HF-LPLI-induced apoptotic cells, and inhibition of NF-κB activity by pyrrolidinedithiocarbamic acid (PDTC) reduced the elevated levels of TNF-α secretion and NO production. These data indicate that HF-LPLI-induced apoptotic tumor cells induce the secretion of pro-inflammatory cytokines in macrophages, which may be helpful for better understanding the biological effects of cancer phototherapy.

  14. Isolation of cell type-specific apoptotic bodies by fluorescence-activated cell sorting

    PubMed Central

    Atkin-Smith, Georgia K.; Paone, Stephanie; Zanker, Damien J.; Duan, Mubing; Phan, Than K.; Chen, Weisan; Hulett, Mark D.; Poon, Ivan K. H.

    2017-01-01

    Apoptotic bodies (ApoBDs) are membrane-bound extracellular vesicles that can mediate intercellular communication in physiological and pathological settings. By combining recently developed analytical strategies with fluorescence-activated cell sorting (FACS), we have developed a method that enables the isolation of ApoBDs from cultured cells to 99% purity. In addition, this approach also enables the identification and isolation of cell type-specific ApoBDs from tissue, bodily fluid and blood-derived samples. PMID:28057919

  15. Regulation of cell death receptor S-nitrosylation and apoptotic signaling by Sorafenib in hepatoblastoma cells.

    PubMed

    Rodríguez-Hernández, A; Navarro-Villarán, E; González, R; Pereira, S; Soriano-De Castro, L B; Sarrias-Giménez, A; Barrera-Pulido, L; Álamo-Martínez, J M; Serrablo-Requejo, A; Blanco-Fernández, G; Nogales-Muñoz, A; Gila-Bohórquez, A; Pacheco, D; Torres-Nieto, M A; Serrano-Díaz-Canedo, J; Suárez-Artacho, G; Bernal-Bellido, C; Marín-Gómez, L M; Barcena, J A; Gómez-Bravo, M A; Padilla, C A; Padillo, F J; Muntané, J

    2015-12-01

    Nitric oxide (NO) plays a relevant role during cell death regulation in tumor cells. The overexpression of nitric oxide synthase type III (NOS-3) induces oxidative and nitrosative stress, p53 and cell death receptor expression and apoptosis in hepatoblastoma cells. S-nitrosylation of cell death receptor modulates apoptosis. Sorafenib is the unique recommended molecular-targeted drug for the treatment of patients with advanced hepatocellular carcinoma. The present study was addressed to elucidate the potential role of NO during Sorafenib-induced cell death in HepG2 cells. We determined the intra- and extracellular NO concentration, cell death receptor expression and their S-nitrosylation modifications, and apoptotic signaling in Sorafenib-treated HepG2 cells. The effect of NO donors on above parameters has also been determined. Sorafenib induced apoptosis in HepG2 cells. However, low concentration of the drug (10nM) increased cell death receptor expression, as well as caspase-8 and -9 activation, but without activation of downstream apoptotic markers. In contrast, Sorafenib (10 µM) reduced upstream apoptotic parameters but increased caspase-3 activation and DNA fragmentation in HepG2 cells. The shift of cell death signaling pathway was associated with a reduction of S-nitrosylation of cell death receptors in Sorafenib-treated cells. The administration of NO donors increased S-nitrosylation of cell death receptors and overall induction of cell death markers in control and Sorafenib-treated cells. In conclusion, Sorafenib induced alteration of cell death receptor S-nitrosylation status which may have a relevant repercussion on cell death signaling in hepatoblastoma cells. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Suppression of Sclerostin Alleviates Radiation-Induced Bone Loss by Protecting Bone-Forming Cells and Their Progenitors Through Distinct Mechanisms

    PubMed Central

    Chandra, Abhishek; Lin, Tiao; Young, Tiffany; Tong, Wei; Ma, Xiaoyuan; Tseng, Wei-Ju; Kramer, Ina; Kneissel, Michaela; Levine, Michael A; Zhang, Yejia; Cengel, Keith; Liu, X. Sherry; Qin, Ling

    2017-01-01

    Focal radiotherapy is frequently associated with skeletal damage within the radiation field. Our previous in vitro study showed that activation of Wnt/β-catenin pathway can overcome radiation-induced DNA damage and apoptosis of osteoblastic cells. Neutralization of circulating sclerostin with a monoclonal antibody (Scl-Ab) is an innovative approach for treating osteoporosis by enhancing Wnt/β-catenin signaling in bone. Together with the fact that focal radiation increases sclerostin amount in bone, we sought to determine whether weekly treatment with Scl-Ab would prevent focal radiotherapy-induced osteoporosis in mice. Micro-CT and histomorphometric analyses demonstrated that Scl-Ab blocked trabecular bone structural deterioration after radiation by partially preserving osteoblast number and activity. Consistently, trabecular bone in sclerostin null mice was resistant to radiation via the same mechanism. Scl-Ab accelerated DNA repair in osteoblasts after radiation by reducing the number of γ-H2AX foci, a DNA double-strand break marker, and increasing the amount of Ku70, a DNA repair protein, thus protecting osteoblasts from radiation-induced apoptosis. In osteocytes, apart from using similar DNA repair mechanism to rescue osteocyte apoptosis, Scl-Ab restored the osteocyte canaliculi structure that was otherwise damaged by radiation. Using a lineage tracing approach that labels all mesenchymal lineage cells in the endosteal bone marrow, we demonstrated that radiation damage to mesenchymal progenitors mainly involves shifting their fate to adipocytes and arresting their proliferation ability but not inducing apoptosis, which are different mechanisms from radiation damage to mature bone forming cells. Scl-Ab treatment partially blocked the lineage shift but had no effect on the loss of proliferation potential. Taken together, our studies provide proof-of-principle evidence for a novel use of Scl-Ab as a therapeutic treatment for radiation-induced osteoporosis and

  17. Effect of captopril on radiation-induced TGF-β1 secretion in EA.Hy926 human umbilical vein endothelial cells.

    PubMed

    Wei, Jingni; Xu, Hui; Liu, Yinyin; Li, Baiyu; Zhou, Fuxiang

    2017-02-15

    The pathophysiological mechanism involved in the sustained endothelial secretion of cytokines that leads to fibrosis 6-16 months after radiotherapy remains unclear. Angiotensin II (Ang II) is produced by the endothelium in response to stressing stimuli, like radiation, and may induce the synthesis of TGF-β, a profibrotic cytokine. In this study we tested the hypothesis that captopril, an angiotensin-converting enzyme (ACE) inhibitor, inhibits or attenuates radiation-induced endothelial TGF-β1 secretion. The human endothelial hybrid cell line EA.HY926 was irradiated with split doses of x-rays (28 Gy delivered in 14 fractions of 2 Gy). TGF-β1 mRNA, TNF-α mRNA and TGF-β1 protein levels were evaluated by RT-PCR and western blotting each month until the fifth month post radiation. Ang II was detected using radioimmunoassays, NF-κB activity was examined using EMSA, and western blotting was used to detect the expression of Iκ-Bα. To explore the role of Ang II on radiation-induced TGF-β1 release and Iκ-Bα expression, captopril was added to cultured cells before, during, or after irradiation. Sustained strong expression of TGF-β1 was observed after conventional fractionated irradiation. TNF-α, Ang II, and NF-κB activity were also increased in EA.Hy926 cells after radiation. Captopril decreased Ang II expression, inhibited the NF-κB pathway and reduced TGF-β1 expression. These data suggest that captopril might protect the endothelium from radiation-induced injury.

  18. Suppression of Sclerostin Alleviates Radiation-Induced Bone Loss by Protecting Bone-Forming Cells and Their Progenitors Through Distinct Mechanisms.

    PubMed

    Chandra, Abhishek; Lin, Tiao; Young, Tiffany; Tong, Wei; Ma, Xiaoyuan; Tseng, Wei-Ju; Kramer, Ina; Kneissel, Michaela; Levine, Michael A; Zhang, Yejia; Cengel, Keith; Liu, X Sherry; Qin, Ling

    2017-02-01

    Focal radiotherapy is frequently associated with skeletal damage within the radiation field. Our previous in vitro study showed that activation of Wnt/β-catenin pathway can overcome radiation-induced DNA damage and apoptosis of osteoblastic cells. Neutralization of circulating sclerostin with a monoclonal antibody (Scl-Ab) is an innovative approach for treating osteoporosis by enhancing Wnt/β-catenin signaling in bone. Together with the fact that focal radiation increases sclerostin amount in bone, we sought to determine whether weekly treatment with Scl-Ab would prevent focal radiotherapy-induced osteoporosis in mice. Micro-CT and histomorphometric analyses demonstrated that Scl-Ab blocked trabecular bone structural deterioration after radiation by partially preserving osteoblast number and activity. Consistently, trabecular bone in sclerostin null mice was resistant to radiation via the same mechanism. Scl-Ab accelerated DNA repair in osteoblasts after radiation by reducing the number of γ-H2AX foci, a DNA double-strand break marker, and increasing the amount of Ku70, a DNA repair protein, thus protecting osteoblasts from radiation-induced apoptosis. In osteocytes, apart from using similar DNA repair mechanism to rescue osteocyte apoptosis, Scl-Ab restored the osteocyte canaliculi structure that was otherwise damaged by radiation. Using a lineage tracing approach that labels all mesenchymal lineage cells in the endosteal bone marrow, we demonstrated that radiation damage to mesenchymal progenitors mainly involves shifting their fate to adipocytes and arresting their proliferation ability but not inducing apoptosis, which are different mechanisms from radiation damage to mature bone forming cells. Scl-Ab treatment partially blocked the lineage shift but had no effect on the loss of proliferation potential. Taken together, our studies provide proof-of-principle evidence for a novel use of Scl-Ab as a therapeutic treatment for radiation-induced osteoporosis and

  19. Die another way – non-apoptotic mechanisms of cell death

    PubMed Central

    Tait, Stephen W. G.; Ichim, Gabriel; Green, Douglas R.

    2014-01-01

    ABSTRACT Regulated, programmed cell death is crucial for all multicellular organisms. Cell death is essential in many processes, including tissue sculpting during embryogenesis, development of the immune system and destruction of damaged cells. The best-studied form of programmed cell death is apoptosis, a process that requires activation of caspase proteases. Recently it has been appreciated that various non-apoptotic forms of cell death also exist, such as necroptosis and pyroptosis. These non-apoptotic cell death modalities can be either triggered independently of apoptosis or are engaged should apoptosis fail to execute. In this Commentary, we discuss several regulated non-apoptotic forms of cell death including necroptosis, autophagic cell death, pyroptosis and caspase-independent cell death. We outline what we know about their mechanism, potential roles in vivo and define outstanding questions. Finally, we review data arguing that the means by which a cell dies actually matters, focusing our discussion on inflammatory aspects of cell death. PMID:24833670

  20. Growth arrest and induction of apoptotic and non-apoptotic programmed cell death by, Physalis minima L. chloroform extract in human ovarian carcinoma Caov-3 cells.

    PubMed

    Ooi, Kheng Leong; Muhammad, Tengku Sifzizul Tengku; Sulaiman, Shaida Fariza

    2010-03-02

    The decoction of the whole plant of Physalis minima L. is traditionally consumed to treat cancer. Its anticancer property has been previously verified (using in vitro cytotoxicity assays) against NCI-H23 lung, CORL23 lung and MCF7 breast cancer cell lines but the mechanism underlying the anticancer potency towards ovarian carcinoma cells remain unclear. The present study is aimed to systematically determine the cytotoxicity and possible cell death mechanism elicited by the chloroform extract of Physalis minima in human ovarian Caov-3 carcinoma. Cytotoxicity of the extract was measured using the methylene blue assay. The mechanism of cell death was determined using four independent methods, namely DeadEnd assay to label the DNA fragmentation nuclei cells, RT-PCR analysis to determine the mRNA expression level of three apoptotic genes (c-myc, p53 and caspase-3 genes), Transmission Electron Microscope (TEM) analysis to describe the ultra structural characteristics and annexin V and propidium iodide staining to confirm the types and stages of cell deaths. Cytotoxicity screening of the extract on Caov-3 cells exhibited concentration- and time-dependent inhibitory effects. A combination of apoptotic and autophagic programmed cell death was detected. The apoptotic characteristic was initially determined by DNA fragmentation followed by the expression of c-myc and p53 genes that was much earlier than caspase-3. Apoptotic ultra structural changes (including clumping and magination of chromatin, blebbing and convolution of nucleus membrane and formation of apoptotic bodies) and autophagy (Type II non-apoptotic programmed cell death) with distinct vacuolated morphology were detected in TEM analysis. The existence of these programmed cell deaths was then corroborated using annexin V and propidium iodide staining. The chloroform extract of Physalis minima exerted anticancer effect due to a combination of apoptotic and autophagic cell death mechanisms on Caov-3 cells. The

  1. Effect of pretreatment with cysteamine on gamma-radiation-induced sister chromatid exchanges in mouse bone marrow cells in vivo

    SciTech Connect

    Mendiola-Cruz, M.T.; Morales-Ramirez, P.

    1989-04-01

    The effect of pretreatment with cysteamine on gamma-radiation-induced sister chromatid exchanges (SCEs) and on the mitotic index and average generation time was determined. Groups of mice were treated in one of the following regimens: (1) irradiated, (2) treated with cysteamine and irradiated, (3) treated with cysteamine only, or (4) left untreated. Intraperitoneal administration of cysteamine preceding gamma-radiation exposure protected against SCE induction. However, radioprotection was not reflected by change in the mitotic index or in the average generation time. The results suggest that, under the experimental conditions of this study, the SCEs are caused by free radicals produced by gamma radiation, but not the additional damage indices measured.

  2. Helicobacter pylori infection inhibits phagocyte clearance of apoptotic gastric epithelial cells.

    PubMed

    Bimczok, Diane; Smythies, Lesley E; Waites, Ken B; Grams, Jayleen M; Stahl, Richard D; Mannon, Peter J; Peter, Shajan; Wilcox, C Mel; Harris, Paul R; Das, Soumita; Ernst, Peter B; Smith, Phillip D

    2013-06-15

    Increased apoptotic death of gastric epithelial cells is a hallmark of Helicobacter pylori infection, and altered epithelial cell turnover is an important contributor to gastric carcinogenesis. To address the fate of apoptotic gastric epithelial cells and their role in H. pylori mucosal disease, we investigated phagocyte clearance of apoptotic gastric epithelial cells in H. pylori infection. Human gastric mononuclear phagocytes were analyzed for their ability to take up apoptotic epithelial cells (AECs) in vivo using immunofluorescence analysis. We then used primary human gastric epithelial cells induced to undergo apoptosis by exposure to live H. pylori to study apoptotic cell uptake by autologous monocyte-derived macrophages. We show that HLA-DR(+) mononuclear phagocytes in human gastric mucosa contain cytokeratin-positive and TUNEL-positive AEC material, indicating that gastric phagocytes are involved in AEC clearance. We further show that H. pylori both increased apoptosis in primary gastric epithelial cells and decreased phagocytosis of the AECs by autologous monocyte-derived macrophages. Reduced macrophage clearance of apoptotic cells was mediated in part by H. pylori-induced macrophage TNF-α, which was expressed at higher levels in H. pylori-infected, compared with uninfected, gastric mucosa. Importantly, we show that H. pylori-infected gastric mucosa contained significantly higher numbers of AECs and higher levels of nonphagocytosed TUNEL-positive apoptotic material, consistent with a defect in apoptotic cell clearance. Thus, as shown in other autoimmune and chronic inflammatory diseases, insufficient phagocyte clearance may contribute to the chronic and self-perpetuating inflammation in human H. pylori infection.

  3. Oncogenic Properties of Apoptotic Tumor Cells in Aggressive B Cell Lymphoma

    PubMed Central

    Ford, Catriona A.; Petrova, Sofia; Pound, John D.; Voss, Jorine J.L.P.; Melville, Lynsey; Paterson, Margaret; Farnworth, Sarah L.; Gallimore, Awen M.; Cuff, Simone; Wheadon, Helen; Dobbin, Edwina; Ogden, Carol Anne; Dumitriu, Ingrid E.; Dunbar, Donald R.; Murray, Paul G.; Ruckerl, Dominik; Allen, Judith E.; Hume, David A.; van Rooijen, Nico; Goodlad, John R.; Freeman, Tom C.; Gregory, Christopher D.

    2015-01-01

    Summary Background Cells undergoing apoptosis are known to modulate their tissue microenvironments. By acting on phagocytes, notably macrophages, apoptotic cells inhibit immunological and inflammatory responses and promote trophic signaling pathways. Paradoxically, because of their potential to cause death of tumor cells and thereby militate against malignant disease progression, both apoptosis and tumor-associated macrophages (TAMs) are often associated with poor prognosis in cancer. We hypothesized that, in progression of malignant disease, constitutive loss of a fraction of the tumor cell population through apoptosis could yield tumor-promoting effects. Results Here, we demonstrate that apoptotic tumor cells promote coordinated tumor growth, angiogenesis, and accumulation of TAMs in aggressive B cell lymphomas. Through unbiased “in situ transcriptomics” analysis—gene expression profiling of laser-captured TAMs to establish their activation signature in situ—we show that these cells are activated to signal via multiple tumor-promoting reparatory, trophic, angiogenic, tissue remodeling, and anti-inflammatory pathways. Our results also suggest that apoptotic lymphoma cells help drive this signature. Furthermore, we demonstrate that, upon induction of apoptosis, lymphoma cells not only activate expression of the tumor-promoting matrix metalloproteinases MMP2 and MMP12 in macrophages but also express and process these MMPs directly. Finally, using a model of malignant melanoma, we show that the oncogenic potential of apoptotic tumor cells extends beyond lymphoma. Conclusions In addition to its profound tumor-suppressive role, apoptosis can potentiate cancer progression. These results have important implications for understanding the fundamental biology of cell death, its roles in malignant disease, and the broader consequences of apoptosis-inducing anti-cancer therapy. PMID:25702581

  4. Oncogenic properties of apoptotic tumor cells in aggressive B cell lymphoma.

    PubMed

    Ford, Catriona A; Petrova, Sofia; Pound, John D; Voss, Jorine J L P; Melville, Lynsey; Paterson, Margaret; Farnworth, Sarah L; Gallimore, Awen M; Cuff, Simone; Wheadon, Helen; Dobbin, Edwina; Ogden, Carol Anne; Dumitriu, Ingrid E; Dunbar, Donald R; Murray, Paul G; Ruckerl, Dominik; Allen, Judith E; Hume, David A; van Rooijen, Nico; Goodlad, John R; Freeman, Tom C; Gregory, Christopher D

    2015-03-02

    Cells undergoing apoptosis are known to modulate their tissue microenvironments. By acting on phagocytes, notably macrophages, apoptotic cells inhibit immunological and inflammatory responses and promote trophic signaling pathways. Paradoxically, because of their potential to cause death of tumor cells and thereby militate against malignant disease progression, both apoptosis and tumor-associated macrophages (TAMs) are often associated with poor prognosis in cancer. We hypothesized that, in progression of malignant disease, constitutive loss of a fraction of the tumor cell population through apoptosis could yield tumor-promoting effects. Here, we demonstrate that apoptotic tumor cells promote coordinated tumor growth, angiogenesis, and accumulation of TAMs in aggressive B cell lymphomas. Through unbiased "in situ transcriptomics" analysis-gene expression profiling of laser-captured TAMs to establish their activation signature in situ-we show that these cells are activated to signal via multiple tumor-promoting reparatory, trophic, angiogenic, tissue remodeling, and anti-inflammatory pathways. Our results also suggest that apoptotic lymphoma cells help drive this signature. Furthermore, we demonstrate that, upon induction of apoptosis, lymphoma cells not only activate expression of the tumor-promoting matrix metalloproteinases MMP2 and MMP12 in macrophages but also express and process these MMPs directly. Finally, using a model of malignant melanoma, we show that the oncogenic potential of apoptotic tumor cells extends beyond lymphoma. In addition to its profound tumor-suppressive role, apoptosis can potentiate cancer progression. These results have important implications for understanding the fundamental biology of cell death, its roles in malignant disease, and the broader consequences of apoptosis-inducing anti-cancer therapy. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  5. Effect of Apoptotic Cell Recognition on Macrophage Polarization and Mycobacterial Persistence

    PubMed Central

    de Oliveira Fulco, Tatiana; Andrade, Priscila Ribeiro; de Mattos Barbosa, Mayara Garcia; Pinto, Thiago Gomes Toledo; Ferreira, Paula Fernandez; Ferreira, Helen; da Costa Nery, José Augusto; Real, Suzana Côrte; Borges, Valéria Matos; Moraes, Milton Ozório; Sarno, Euzenir Nunes; Sampaio, Elizabeth Pereira

    2014-01-01

    Intracellular Mycobacterium leprae infection modifies host macrophage programming, creating a protective niche for bacterial survival. The milieu regulating cellular apoptosis in the tissue plays an important role in defining susceptible and/or resistant phenotypes. A higher density of apoptotic cells has been demonstrated in paucibacillary leprosy lesions than in multibacillary ones. However, the effect of apoptotic cell removal on M. leprae-stimulated cells has yet to be fully elucidated. In this study, we investigated whether apoptotic cell removal (efferocytosis) induces different phenotypes in proinflammatory (Mϕ1) and anti-inflammatory (Mϕ2) macrophages in the presence of M. leprae. We stimulated Mϕ1 and Mϕ2 cells with M. leprae in the presence or absence of apoptotic cells and subsequently evaluated the M. leprae uptake, cell phenotype, and cytokine pattern in the supernatants. In the presence of M. leprae and apoptotic cells, Mϕ1 macrophages changed their phenotype to resemble the Mϕ2 phenotype, displaying increased CD163 and SRA-I expression as well as higher phagocytic capacity. Efferocytosis increased M. leprae survival in Mϕ1 cells, accompanied by reduced interleukin-15 (IL-15) and IL-6 levels and increased transforming growth factor beta (TGF-β) and IL-10 secretion. Mϕ1 cells primed with M. leprae in the presence of apoptotic cells induced the secretion of Th2 cytokines IL-4 and IL-13 in autologous T cells compared with cultures stimulated with M. leprae or apoptotic cells alone. Efferocytosis did not alter the Mϕ2 cell phenotype or cytokine secretion profile, except for TGF-β. Based on these data, we suggest that, in paucibacillary leprosy patients, efferocytosis contributes to mycobacterial persistence by increasing the Mϕ2 population and sustaining the infection. PMID:25024361

  6. Celecoxib Induced Tumor Cell Radiosensitization by Inhibiting Radiation Induced Nuclear EGFR Transport and DNA-Repair: A COX-2 Independent Mechanism

    SciTech Connect

    Dittmann, Klaus H. Mayer, Claus; Ohneseit, Petra A.; Raju, Uma; Andratschke, Nickolaus H.; Milas, Luka; Rodemann, H. Peter

    2008-01-01

    Purpose: The purpose of the study was to elucidate the molecular mechanisms mediating radiosensitization of human tumor cells by the selective cyclooxygenase (COX)-2 inhibitor celecoxib. Methods and Materials: Experiments were performed using bronchial carcinoma cells A549, transformed fibroblasts HH4dd, the FaDu head-and-neck tumor cells, the colon carcinoma cells HCT116, and normal fibroblasts HSF7. Effects of celecoxib treatment were assessed by clonogenic cell survival, Western analysis, and quantification of residual DNA damage by {gamma}H{sub 2}AX foci assay. Results: Celecoxib treatment resulted in a pronounced radiosensitization of A549, HCT116, and HSF7 cells, whereas FaDu and HH4dd cells were not radiosensitized. The observed radiosensitization could neither be correlated with basal COX-2 expression pattern nor with basal production of prostaglandin E2, but was depended on the ability of celecoxib to inhibit basal and radiation-induced nuclear transport of epidermal growth factor receptor (EGFR). The nuclear EGFR transport was strongly inhibited in A549-, HSF7-, and COX-2-deficient HCT116 cells, which were radiosensitized, but not in FaDu and HH4dd cells, which resisted celecoxib-induced radiosensitization. Celecoxib inhibited radiation-induced DNA-PK activation in A549, HSF7, and HCT116 cells, but not in FaDu and HH4dd cells. Consequentially, celecoxib increased residual {gamma}H2AX foci after irradiation, demonstrating that inhibition of DNA repair has occurred in responsive A549, HCT116, and HSF7 cells only. Conclusions: Celecoxib enhanced radiosensitivity by inhibition of EGFR-mediated mechanisms of radioresistance, a signaling that was independent of COX-2 activity. This novel observation may have therapeutic implications such that COX-2 inhibitors may improve therapeutic efficacy of radiation even in patients whose tumor radioresistance is not dependent on COX-2.

  7. Sci—Fri AM: Mountain — 04: Label-free Raman spectroscopy of single tumour cells detects early radiation-induced glycogen synthesis associated with increased radiation resistance

    SciTech Connect

    Matthews, Q; Lum, JJ; Isabelle, M; Harder, S; Jirasek, A; Brolo, AG

    2014-08-15

    Purpose: To use label-free Raman spectroscopy (RS) for early treatment monitoring of tumour cell radioresistance. Methods: Three human tumour cell lines, two radioresistant (H460, SF{sub 2} = 0.57 and MCF7, SF{sub 2} = 0.70) and one radiosensitive (LNCaP, SF{sub 2} = 0.36), were irradiated with single fractions of 2, 4, 6, 8 or 10 Gy. In additional experiments, H460 and MCF7 cells were irradiated under co-treatment with the anti-diabetic drug metformin, a known radiosensitizing agent. Treated and control cultures were analyzed with RS daily for 3 days post-treatment. Single-cell Raman spectra were acquired from 20 live cells per sample, and experiments were repeated in triplicate. The combined data sets were analyzed with principal component analysis using standard algorithms. Cells from each culture were also subjected to standard assays for viability, proliferation, cell cycle, and radiation clonogenic survival. Results: The radioresistant cells (H460, MCF7) exhibited a RS molecular radiation response signature, detectable as early as 1 day post-treatment, of which radiation-induced glycogen synthesis is a significant contributor. The radiosensitive cells (LNCaP) exhibited negligible glycogen synthesis. Co-treatment with metformin in MCF7 cells blocked glycogen synthesis, reduced viability and proliferation, and increased radiosensitivity. Conversely, metformin co-treatment in H460 cells did not produce these same effects; importantly, both radiation-induced synthesis of glycogen and radiosensitivity were unaffected. Conclusions: Label-free RS can detect early glycogen synthesis post-irradiation, a previously undocumented metabolic mechanism associated with tumour cell radioresistance that can be targeted to increase radiosensitivity. RS monitoring of intratumoral glycogen may provide new opportunities for personalized combined modality radiotherapy treatments.

  8. Approaches to augment CAR T-cell therapy by targeting the apoptotic machinery.

    PubMed

    Karlsson, Hannah

    2016-04-15

    Chimaeric antigen receptor (CAR) T-cells have shown impressive results in patients with B-cell leukaemia. Yet, in patients with lymphoma durable responses are still rare and heavy preconditioning required. Apoptosis resistance is considered a hallmark of cancer, often conveyed by a halted apoptosis signalling. Tumours regularly skew the balance of the components of the apoptotic machinery either through up-regulating anti-apoptotic proteins or silencing pro-apoptotic ones. Malignant B-cells frequently up-regulate anti-apoptotic B-cell lymphoma 2 (Bcl-2) family proteins leading to therapy resistance. CAR T-cells kill tumour cells via apoptosis induction and their efficacy may be affected by the level of Bcl-2 family proteins. Hence, there is an interesting possibility to increase the effect of CAR T-cell therapy by combining it with apoptosis inhibitor blockade agents. Compounds that inhibit Bcl-2, B-cell lymphoma extra large (Bcl-xL) and Bcl-2-like protein 2 (Bcl-w), can restore execution of apoptosis in tumour cells or sensitize them to other apoptosis-dependent treatments. Hence, there is a great interest to combine such agents with CAR T-cell therapy to potentiate the effect of CAR T-cell killing. This review will focus on the potential of targeting the apoptotic machinery to sensitize tumour cells to CAR T-cell killing. © 2016 Authors; published by Portland Press Limited.

  9. Nintedanib Compared With Placebo in Treating Against Radiation-Induced Pneumonitis in Patients With Non-small Cell Lung Cancer That Cannot Be Removed by Surgery and Are Undergoing Chemoradiation Therapy

    ClinicalTrials.gov

    2017-07-08

    Radiation-Induced Pneumonitis; Stage IIA Non-Small Cell Lung Carcinoma; Stage IIB Non-Small Cell Lung Carcinoma; Stage IIIA Non-Small Cell Lung Cancer; Stage IIIB Non-Small Cell Lung Cancer; Stage IV Non-Small Cell Lung Cancer

  10. Tolerance to apoptotic cells is regulated by indoleamine 2,3-dioxygenase

    PubMed Central

    Ravishankar, Buvana; Liu, Haiyun; Shinde, Rahul; Chandler, Phillip; Baban, Babak; Tanaka, Masato; Munn, David H.; Mellor, Andrew L.; Karlsson, Mikael C. I.; McGaha, Tracy L.

    2012-01-01

    Tolerance to self-antigens present in apoptotic cells is critical to maintain immune-homeostasis and prevent systemic autoimmunity. However, mechanisms that sustain self-tolerance are poorly understood. Here we show that systemic administration of apoptotic cells to mice induced splenic expression of the tryptophan catabolizing enzyme indoleamine 2,3-dioxygenase (IDO). IDO expression was confined to the splenic marginal zone and was abrogated by depletion of CD169+ cells. Pharmacologic inhibition of IDO skewed the immune response to apoptotic cells, resulting in increased proinflammatory cytokine production and increased effector T-cell responses toward apoptotic cell-associated antigens. Presymptomatic lupus-prone MRLlpr/lpr mice exhibited abnormal elevated IDO expression in the marginal zone and red pulp and inhibition of IDO markedly accelerated disease progression. Moreover, chronic exposure of IDO-deficient mice to apoptotic cells induced a lupus-like disease with serum autoreactivity to double-stranded DNA associated with renal pathology and increased mortality. Thus, IDO limits innate and adaptive immunity to apoptotic self-antigens and IDO-mediated regulation inhibits inflammatory pathology caused by systemic autoimmune disease. PMID:22355111

  11. Investigation of Radiation-induced Transcriptome Profile of Radioresistant Non-small Cell Lung Cancer A549 Cells Using RNA-seq

    PubMed Central

    Yang, Hee Jung; Kim, Namshin; Seong, Ki Moon; Youn, HyeSook; Youn, BuHyun

    2013-01-01

    Radioresistance is a main impediment to effective radiotherapy for non-small cell lung cancer (NSCLC). Despite several experimental and clinical studies of resistance to radiation, the precise mechanism of radioresistance in NSCLC cells and tissues still remains unclear. This result could be explained by limitation of previous researches such as a partial understanding of the cellular radioresistance mechanism at a single molecule level. In this study, we aimed to investigate extensive radiation responses in radioresistant NSCLC cells and to identify radioresistance-associating factors. For the first time, using RNA-seq, a massive sequencing-based approach, we examined whole-transcriptome alteration in radioresistant NSCLC A549 cells under irradiation, and verified significant radiation-altered genes and their chromosome distribution patterns. Also, bioinformatic approaches (GO analysis and IPA) were performed to characterize the radiation responses in radioresistant A549 cells. We found that epithelial–mesenchymal transition (EMT), migration and inflammatory processes could be meaningfully related to regulation of radiation responses in radioresistant A549 cells. Based on the results of bioinformatic analysis for the radiation-induced transcriptome alteration, we selected seven significant radiation-altered genes (SESN2, FN1, TRAF4, CDKN1A, COX-2, DDB2 and FDXR) and then compared radiation effects in two types of NSCLC cells with different radiosensitivity (radioresistant A549 cells and radiosensitive NCI-H460 cells). Interestingly, under irradiation, COX-2 showed the most significant difference in mRNA and protein expression between A549 and NCI-H460 cells. IR-induced increase of COX-2 expression was appeared only in radioresistant A549 cells. Collectively, we suggest that COX-2 (also known as prostaglandin-endoperoxide synthase 2 (PTGS2)) could have possibility as a putative biomarker for radioresistance in NSCLC cells. PMID:23533613

  12. Apoptotic Capacity and Risk of Squamous Cell Carcinoma of the Head and Neck

    PubMed Central

    Liu, Zhensheng; Liu, Hongliang; Han, Peng; Gao, Fengqin; Dahlstrom, Kristina R.; Li, Guojun; Owzar, Kouros; Zevallos, Jose P.; Sturgis, Erich M.; Wei, Qingyi

    2017-01-01

    Background Tobacco smoke and alcohol drinking are the major risk factors for squamous cell carcinoma of the head and neck (SCCHN). Smoking and drinking cause DNA damage leading to apoptosis, and insufficient apoptotic capacity may favor development of cancer because of the dysfunction of removing damaged cells. In the present study, we investigated the association between camptothecin (CPT)-induced apoptotic capacity and risk of SCCHN in a North American population. Methods In a case-control study of 708 SCCHN patients and 685 matched cancer-free controls, we measured apoptotic capacity in cultured peripheral blood lymphocytes (PBLs) in response to in vitro exposure to CPT by using the flow cytometry-based method. Results We found that the mean level of apoptotic capacity in the cases (45.9±23.3%) was significantly lower than that in the controls (49.0±23.1%) (P = 0.002). When we used the median level of apoptotic capacity in the controls as the cutoff value for calculating adjusted odds ratios (ORs), subjects with a reduced apoptotic capacity had an increased risk (adjusted OR = 1.42, 95% confidence interval [CI] = 1.13–1.78, P = 0.002), especially for those who were age ≥57 (1.73, 1.25–2.38, 0.0009), men (1.76, 1.36–2.27, < 0.0001) and ever drinkers (1.67, 1.27–2.21, 0.0003), and these variables significantly interacted with apoptotic capacity (Pinteraction = 0.015, 0.005 and 0.009, respectively). A further fitted prediction model suggested that the inclusion of apoptotic capacity significantly improved in the prediction of SCCHN risk. Conclusion Individuals with a reduced CPT-induced apoptotic capacity may be at an increased risk of developing SCCHN, and apoptotic capacity may be a biomarker for susceptibility to SCCHN. PMID:28033527

  13. Pallbearer and friends: lending a hand in the clearance of apoptotic cells

    PubMed Central

    Elliott, Michael R.; Ravichandran, Kodi S.

    2010-01-01

    Engulfment and prompt removal of apoptotic cells occurs from embryogenesis throughout the lifespan of multi-cellular organisms. A new player, Pallbearer, has recently been identified in Drosophila as being important for efficient engulfment by macrophages. Pallbearer is a component of the SCF E3 ubiquitin ligase complex involved in ubiquitylation of proteins targeted for proteasomal degradation. This work provides the first link between the cellular processes of ubiquitylation/proteasomal degradation and the ability to efficiently clear apoptotic cells. PMID:18280734

  14. Paeoniflorin protects human EA.hy926 endothelial cells against gamma-radiation induced oxidative injury by activating the NF-E2-related factor 2/heme oxygenase-1 pathway.

    PubMed

    Yu, Jing; Zhu, Xiaoyun; Qi, Xin; Che, Juanjuan; Cao, Bangwei

    2013-04-26

    Pulmonary endothelial cells have been demonstrated to have a critical role in the pathogenesis of radiation-induced lung injury. Our preliminary experiments indicated that paeoniflorin protected human EA.hy926 endothelial cells from radiation-induced oxidative injury. This study was designed to confirm the protective effect of paeoniflorin against radiation-induced endothelial cellular damage and to elucidate the underlying mechanisms. Preincubation of EA.hy926 cells with paeoniflorin before γ-radiation resulted in significant inhibition of apoptosis, a decrease in mitochondrial membrane potential and enhanced cell viability. In particular, we showed that paeoniflorin significantly reduced the formation of intracellular reactive oxygen species (ROS), the level of malondialdehyde (MDA) and lactate dehydrogenase (LDH) leakage, and enhanced production of the endogenous antioxidants, glutathione (GSH) and superoxide dismutase (SOD) in EA.hy926 cells. Treatment of these cells with paeoniflorin significantly induced HO-1 expression. Moreover, paeoniflorin promoted the nuclear translocation of nuclear factor erythroid 2 related factor-2 (Nrf-2). The paeoniflorin-induced HO-1 expression was abrogated by Nrf2 siRNA. Furthermore, inhibition of HO-1 with zinc protoporphyrin IX (ZNPP) significantly reversed the protective effect of paeoniflorin against radiation-induced damage in EA.hy926 cells. Our findings confirmed that paeoniflorin protected EA.hy926 cells against radiation-induced injury through the Nrf2/HO-1 pathway.

  15. Dasatinib blocks cetuximab- and radiation-induced nuclear translocation of the epidermal growth factor receptor in head and neck squamous cell carcinoma

    PubMed Central

    Li, Chunrong; Iida, Mari; Dunn, Emily F.; Wheeler, Deric L.

    2010-01-01

    Background and Purpose The aberrant expression of epidermal growth factor receptor (EGFR) has been linked to the etiology of head and neck squamous cell carcinoma (HNSCC). The first major phase III trial combining cetuximab with radiation confirmed a strong survival advantage. However, both cetuximab and radiation can promote EGFR translocation to the nucleus where it enhances resistance to both of these modalities. In this report we sought to determine how to block cetuximab and radiation–induced translocation of EGFR to the nucleus in HNSCC cell lines. Material and Methods We utilized three established HNSCC cell lines, SCC1, SCC6 and SCC1483 and measured nuclear translocation of EGFR after treatment with cetuximab or radiation. We then utilized dasatinib (BMS-354825), a potent, orally bioavailable inhibitor of several tyrosine kinases, including the Src Family Kinases, to determine if SFKs blockade could abrogate cetuximab and radiation-induced nuclear EGFR translocation. Results Cetuximab and radiation treatment of all three HNSCC lines lead to translocation of the EGFR to the nucleus. Blockade of SFKs abrogated cetuximab and radiation-induced EGFR translocation to the nucleus. Conclusions The data presented in this report suggests that both cetuximab and radiation can promote EGFR translocation to the nucleus and dasatinib can inhibit this