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Sample records for cell killing effect

  1. Morphological effect of oscillating magnetic nanoparticles in killing tumor cells

    Science.gov (United States)

    Cheng, Dengfeng; Li, Xiao; Zhang, Guoxin; Shi, Hongcheng

    2014-04-01

    Forced oscillation of spherical and rod-shaped iron oxide magnetic nanoparticles (MNPs) via low-power and low-frequency alternating magnetic field (AMF) was firstly used to kill cancer cells in vitro. After being loaded by human cervical cancer cells line (HeLa) and then exposed to a 35-kHz AMF, MNPs mechanically damaged cell membranes and cytoplasm, decreasing the cell viability. It was found that the concentration and morphology of the MNPs significantly influenced the cell-killing efficiency of oscillating MNPs. In this preliminary study, when HeLa cells were pre-incubated with 100 μg/mL rod-shaped MNPs (rMNP, length of 200 ± 50 nm and diameter of 50 to 120 nm) for 20 h, MTT assay proved that the cell viability decreased by 30.9% after being exposed to AMF for 2 h, while the cell viability decreased by 11.7% if spherical MNPs (sMNP, diameter of 200 ± 50 nm) were used for investigation. Furthermore, the morphological effect of MNPs on cell viability was confirmed by trypan blue assay: 39.5% rMNP-loaded cells and 15.1% sMNP-loaded cells were stained after being exposed to AMF for 2 h. It was also interesting to find that killing tumor cells at either higher (500 μg/mL) or lower (20 μg/mL) concentration of MNPs was less efficient than that achieved at 100 μg/mL concentration. In conclusion, the relatively asymmetric morphological rod-shaped MNPs can kill cancer cells more effectively than spherical MNPs when being exposed to AMF by virtue of their mechanical oscillations.

  2. Bystander Host Cell Killing Effects of Clostridium perfringens Enterotoxin

    Directory of Open Access Journals (Sweden)

    Archana Shrestha

    2016-12-01

    Full Text Available Clostridium perfringens enterotoxin (CPE binds to claudin receptors, e.g., claudin-4, and then forms a pore that triggers cell death. Pure cultures of host cells that do not express claudin receptors, e.g., fibroblasts, are unaffected by pathophysiologically relevant CPE concentrations in vitro. However, both CPE-insensitive and CPE-sensitive host cells are present in vivo. Therefore, this study tested whether CPE treatment might affect fibroblasts when cocultured with CPE-sensitive claudin-4 fibroblast transfectants or Caco-2 cells. Under these conditions, immunofluorescence microscopy detected increased death of fibroblasts. This cytotoxic effect involved release of a toxic factor from the dying CPE-sensitive cells, since it could be reproduced using culture supernatants from CPE-treated sensitive cells. Supernatants from CPE-treated sensitive cells, particularly Caco-2 cells, were found to contain high levels of membrane vesicles, often containing a CPE species. However, most cytotoxic activity remained in those supernatants even after membrane vesicle depletion, and CPE was not detected in fibroblasts treated with supernatants from CPE-treated sensitive cells. Instead, characterization studies suggest that a major cytotoxic factor present in supernatants from CPE-treated sensitive cells may be a 10- to 30-kDa host serine protease or require the action of that host serine protease. Induction of caspase-3-mediated apoptosis was found to be important for triggering release of the cytotoxic factor(s from CPE-treated sensitive host cells. Furthermore, the cytotoxic factor(s in these supernatants was shown to induce a caspase-3-mediated killing of fibroblasts. This bystander killing effect due to release of cytotoxic factors from CPE-treated sensitive cells could contribute to CPE-mediated disease.

  3. Optimization of total body irradiation: the match between (maximal) leukemic cell kill and (minimal) late effects

    NARCIS (Netherlands)

    Harteveld, M.L. van

    2007-01-01

    Optimization of total body irradiation: the match between (maximal) leukemic cell kill and (minimal) late effects: In this thesis, cataract formation and renal dysfunction as late effects of high-dose total body irradiation (TBI) as part of the conditioning before hematological stem cell transplanta

  4. Photocatalytic killing effect of TiO2 nanoparticles on Ls-174-t human colon carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Ai-Ping Zhang; Yan-Ping Sun

    2004-01-01

    AIM: To investigate the photocatalytic killing effect of photoexcited TiO2 nanoparticles on human colon carcinoma cell line (Ls-174-t) and to study the mechanism underlying the action of photoexcited TiO2 nanoparticles on malignant cells.METHODS: Ls-174-t human colon carcinoma cells were cultured in RPMI 1640 medium supplemented with 199 mL/L calf serum in a humidified incubator with an atmosphere of 50 mL/L CO2 at 37 ℃. Viable cells in the samples were measured by using the MlT method. A GGZ-300 W high pressure Hg lamp with a maximum ultraviolet-A (UVA, 320-400 nm) irradiation peak at 365 nm was used as light source in the photocatalytic killing test.RESULTS: The photocatalytic killing of Ls-174-t cells was carried out in vitro with TiO2 nanoparticles. The killing effect was weak by using UVA irradiation without TiO2 nanoparticles.In our studies, the photocatalytic killing effect was correlated with the concentration of TiO2 and illumination time. Once TiO2 was added, Ls-174-t cells were killed at a much higher rate. In the presence of 1 000 μg/mL TiO2, 44% of cells were killed after 10 min of UVA irradiation, and 88% of cells were killed after 30 min of UVA irradiation.CONCLUSION: When the concentration of TiO2 is below 200 μg/mL, the photocatalytic killing effect on human colon carcinoma cells is almost the same as that of UVA irradiation alone. When the concentration of TiO2 is above 200 μg/mL,the remarkable killing effect of photoexcited TiO2 nanoparticles can be found.

  5. Effective cancer cell killing by hydrophobic nanovoid-enhanced cavitation under safe low-energy ultrasound.

    Science.gov (United States)

    Zhao, Yang; Zhu, Yingchun; Fu, Jingke; Wang, Lianzhou

    2014-03-01

    β-Cyclodextrin (β-CD)-capped mesoporous silica nanoparticles with hydrophobic internal nanovoids were prepared and used for effective cancer cell killing in synergistic combination with low-energy ultrasound (≤1.0 W cm(-2) , 1 MHz). The water-dispersible nanoparticles with hydrophobic internal nanovoids can be taken up by cancer cells and subsequently evoke a remarkable cavitation effect under irradiation with mild low-energy ultrasound (≤1.0 W cm(-2) , 1 MHz). A significant cancer cell killing effect was observed in cancer cells and in a mouse xenograft tumor model treated with the nanoagents together with the low-energy ultrasound, showing a distinct dependence on the concentration of nanoagents and ultrasound intensity. By contrast, an antitumor effect was not observed when either low-energy ultrasound or nanoagents were applied alone. These findings are significant as the technique promises a safe, low-cost, and effective treatment for cancer therapy.

  6. Killing effect of coexpressing cytosine deaminase and thymidine kinase on rat vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    曹慧青; 孟宪敏; 刘冬青; 赵秀文; 丁金凤

    2004-01-01

    Background Vascular smooth muscle cell (VSMC) proliferation following arterial injury plays a critical role in a variety of vascular proliferative disorders, such as atherosclerosis and restenosis after balloon angioplasty. Herpes simplex virus-thymidine kinase (HSV-TK)/ganciclovir (GCV) and E.coli cytosine deaminase (CD)/5-fluorocytosine (5-Fc) suicide gene systems have been successfully employed in cardiovascular gene therapy, respectively. We reasoned that coexpression of both HSV-TK with CD suicide genes would lead to increased cell killing. To test this imagine, the adenoviral vectors expressing TK and/or CD genes were developed and tested on vascular smooth muscle cells. Methods Adenoviral vectors, including Ad-EF1α-CD-cytomegolovirus (CMV)-TK coexpressing both CD and TK double suicide genes, Ad-EF1α-CD and Ad-CMV-TK expressing CD and TK respectively, and control vector Ad-CMV-LacZ, were constructed and prepared with homologous recombination in RecA+E.coli cells. Integration and expression of CD and/or TK gene were identified by PCR and Western blot. Primary cultured VSMCs were infected at a multiplicity of infection (MOI) of 20 with exposure to their matching prodrugs 5-Fc and GCV. Cell mortality was measured by methyl thiazolyl tetrazolium (MTT) assays. Flow cytometry analysis was used to detect cell death. Apoptotic cells were analyzed using Hoechst 33342 fluorescence dye as a DNA probe. Genomic DNA cleavage of apoptotic VSMCs was tested by agarose gel electrophoresis. Results Recombinant adenovirus expressing CD and/or TK suicide genes were successfully constructed. Both single and double suicide genes could be integrated into adenoviral genome and expressed. Cytotoxic effects of Ad-EF1α-CD-CMV-TK double suicide genes combined with 5-Fc and GCV were higher than those of Ad-CMV-TK and Ad-EF1α-CD single gene groups. The rate of cell survival was only (9±3)% in the Ad-EF1α-CD-CMV-TK group, but (37±3)% in the Ad-CMV-TK and (46±4)% in the Ad-EF1

  7. Enhancing effects of gamma interferon on phagocytic cell association with and killing of Trypanosoma cruzi

    Science.gov (United States)

    Wirth, J. J.; Kierszenbaum, F.; Sonnenfeld, G.; Zlotnik, A.

    1985-01-01

    Results are reported from a study of the influence gamma interferon (GIFN) and interleukin 2 (IL2) have on the capability of P388D1 cells and mouse resident peritoneal macrophages (MPM) to attach to the blood-resident parasites Trypanosoma cruzi and kill them. Cultures of trypomastigote forms of the Tulahuen strain of T. cruzi grown in bovine serum were introduced into peritoneal cells of mice, along with P388D1 cells incubated with GIFN, IL2 and both. Control cells were also maintained. Statistical analysis were then performed on data on counts of the number of dead T. Cruzi cells. The GIFN enhanced the interaction of MPM and P388D1 cells with the surface of T. Cruzi, provided the interaction was given over 12 hr to take place. A depression of the cytotoxicity of P388D1 cells was attributed to mediation by H2O2, an effect partially offset by incubation with the lymphokine GIFN.

  8. Two-stage model of radon-induced malignant lung tumors in rats: effects of cell killing

    Science.gov (United States)

    Luebeck, E. G.; Curtis, S. B.; Cross, F. T.; Moolgavkar, S. H.

    1996-01-01

    A two-stage stochastic model of carcinogenesis is used to analyze lung tumor incidence in 3750 rats exposed to varying regimens of radon carried on a constant-concentration uranium ore dust aerosol. New to this analysis is the parameterization of the model such that cell killing by the alpha particles could be included. The model contains parameters characterizing the rate of the first mutation, the net proliferation rate of initiated cells, the ratio of the rates of cell loss (cell killing plus differentiation) and cell division, and the lag time between the appearance of the first malignant cell and the tumor. Data analysis was by standard maximum likelihood estimation techniques. Results indicate that the rate of the first mutation is dependent on radon and consistent with in vitro rates measured experimentally, and that the rate of the second mutation is not dependent on radon. An initial sharp rise in the net proliferation rate of initiated cell was found with increasing exposure rate (denoted model I), which leads to an unrealistically high cell-killing coefficient. A second model (model II) was studied, in which the initial rise was attributed to promotion via a step function, implying that it is due not to radon but to the uranium ore dust. This model resulted in values for the cell-killing coefficient consistent with those found for in vitro cells. An "inverse dose-rate" effect is seen, i.e. an increase in the lifetime probability of tumor with a decrease in exposure rate. This is attributed in large part to promotion of intermediate lesions. Since model II is preferable on biological grounds (it yields a plausible cell-killing coefficient), such as uranium ore dust. This analysis presents evidence that a two-stage model describes the data adequately and generates hypotheses regarding the mechanism of radon-induced carcinogenesis.

  9. Killing cells by targeting mitosis.

    Science.gov (United States)

    Manchado, E; Guillamot, M; Malumbres, M

    2012-03-01

    Cell cycle deregulation is a common feature of human cancer. Tumor cells accumulate mutations that result in unscheduled proliferation, genomic instability and chromosomal instability. Several therapeutic strategies have been proposed for targeting the cell division cycle in cancer. Whereas inhibiting the initial phases of the cell cycle is likely to generate viable quiescent cells, targeting mitosis offers several possibilities for killing cancer cells. Microtubule poisons have proved efficacy in the clinic against a broad range of malignancies, and novel targeted strategies are now evaluating the inhibition of critical activities, such as cyclin-dependent kinase 1, Aurora or Polo kinases or spindle kinesins. Abrogation of the mitotic checkpoint or targeting the energetic or proteotoxic stress of aneuploid or chromosomally instable cells may also provide further benefits by inducing lethal levels of instability. Although cancer cells may display different responses to these treatments, recent data suggest that targeting mitotic exit by inhibiting the anaphase-promoting complex generates metaphase cells that invariably die in mitosis. As the efficacy of cell-cycle targeting approaches has been limited so far, further understanding of the molecular pathways modulating mitotic cell death will be required to move forward these new proposals to the clinic.

  10. Killing effect of adenoviral mediated cytosine deaminase gene on human pancreatic cancer cell line PaTu 8988

    Institute of Scientific and Technical Information of China (English)

    PAN Xue; LI Zhao-shen; XU Guo-ming; CUI Long; ZHANG Su-zhen; GONG Yan-fang; TU Zhen-xing

    2001-01-01

    Objective: To evaluate the in vitro killing effects of cytosine deaminase gene mediated by adenovirus vector on human pancreatic carcinoma. Methods: Cytosine Deaminase (CD) gene was cloned into pAdTrack-CMV-CD, pAdTrack-CMV-CD and pAdEasy-1 were recombined in bacteria, and the products containing green fluorescent protein (GFP)were propagated in 293 cells and purified by cesium chloride gradient centrifugation. Human pancreatic carcinoma cell line 8988 were infected with this virus, then 5-FC was added; XTT assay was used to estimate the relative numbers of viable cells. Results: The positive clones were confirmed by using endonuclease digestion, and the titer of the virus containing CD gene was 2 × 1011 pfu/ml. It was found that 5-FC possessed significant cytotoxic activities for CD gene transfected 8988cell line, but had little effects on non-transfected pancreatic carcinoma cells. Conclusion: CD gene mediated by adenovirus has a high infectivity and is efficient for killing cultured pancreatic carcinoma cells, indicating suicide gene may be effective for pancreatic cancer in furure.

  11. EFFECTS OF CARBOXYMETHLY DEXTRAN MAGNETIC NANOPARTICLES CARRIER SYSTEM ASSOCIATED WITH EXTERNAL MAGNETIC FIELDS ON KILLING TUMOR CELLS AND GENE TRANSFECTION

    Institute of Scientific and Technical Information of China (English)

    CAO Zheng-guo; ZHOU Si-wei; LIU Ji-hong

    2005-01-01

    Objective: To investigate the preparation of the carboxymethly dextran iron oxide magnetic nanoparticles (CDMN) and the effects of CDMN carrier system associated with external magnetic fields on killing tumor cells and gene transfection in vitro. Methods: Epirubicin-CDMN (EPI-CDMN) and green fluorescent protein (GFP) plasmid-CDMN (GFP-CDMN) were prepared by the oxidation-reduction procedure and their characters were detected, respectively. The effects of EPI-CDMN associated with external pulsed electromagnetic fields (PEMFs) (10 mT) on killing human bladder cancer BIU-87 cells were studied by MTT assay and Annexin-V/PI double-labeled flow cytometry technique, respectively. And the transfection efficiency of GFP when CDMN were used as gene carrier associated with the external magnetic fields was evaluated under fluorescence microscope in vitro. Results: The diameter of EPI-CDMN and GFP-CDMN were about 8~10 nm and 5~9 nm, respectively, and saturation magnetization were 0.22 emu/g and 0.26 emu/g, respectively. EPI-CDMN associated with PEMFs could significantly inhibit the proliferation of BIU-87 cells and induce cells apoptosis, the growth inhibitory rate and apoptosis rate were (21.82(3.18)% and (16.79(3.37)%, respectively. The transfection efficiency of GFP-CDMN combined with PEMFs was significant higher than that of GFP-CDMN without PEMFs [(45.70(4.32)% vs (35.85(2.16)%, P<0.05]. Conclusion: It seemed that EPI-CDMN associated with external magnetic fields could significantly killed human bladder cancer BIU-87 cells and CDMN could effectively transfer GFP gene into tumors cells with the help of external magnetic fields which provided experimental basis for the magnetic targeting therapy of tumor.

  12. Cell killing by avian leukosis viruses.

    OpenAIRE

    Weller, S K; Temin, H M

    1981-01-01

    Infection of chicken cells with a cytopathic avian leukosis virus resulted in the detachment of killed cells from the culture dish. The detached, dead cells contained more unintegrated viral DNA than the attached cells. These results confirm the hypothesis that cell killing after infection with a cytopathic avian leukosis virus is associated with accumulation of large amounts of unintegrated viral DNA. No accumulation of large amounts of integrated viral DNA was found in cells infected with c...

  13. Killing effect of different doses of preoperative iodine 131 therapy on thyroid cancer cells and its effect on salivary gland function

    Institute of Scientific and Technical Information of China (English)

    Xiao-Dong Zheng; Tao Pu; Yi Luo; Xing-An Zhang; Zu-Mao Li

    2015-01-01

    Objective:To study the killing effect of different doses of preoperative iodine 131 therapy on thyroid cancer cells and its effect on salivary gland function.Methods:Patients diagnosed with thyroid cancer in our hospital from May 2013 to June 2014 were enrolled for study, given preoperative iodine 131 therapy and randomly divided into control group, low dose group, middle dose group and high dose group. Then cell apoptotic rate, cell cycle, cancer promoting gene and cancer suppressor gene expression in thyroid carcinoma tissue as well as salivary gland function were detected.Results: (1) cancer cell killing effect: compared with control group, cell apoptotic rates and number of cells in G0/G1 phase of low dose group, middle dose group and high dose group increased, number of cells in S and G2/M phase decreased, BRAF, Livin, MCM7 and CDK2 expression decreased, CCNG2 and PTEN expression increased; cell killing effect of middle dose group and high dose group were better than that of low dose group, and cell killing effect of middle dose group and high dose group had no differences; (2) salivary gland function: compared with control group, UI and SR in bilateral parotid and bilateral submandibular glands of low dose group, middle dose group and high dose group decreased; salivary gland damage effect of low dose group and middle dose group were weaker than that of high dose group, and salivary gland damage effect of low dose group and middle dose group had no differences.Conclusion:Middle dose of iodine 131 can take the killing effect on cancer cells and the protective effect on salivary glands into account; it’s an ideal dosage for preoperative iodine 131 internal radiation therapy of thyroid cancer patients.

  14. Selective Killing Effects of Cold Atmospheric Pressure Plasma with NO Induced Dysfunction of Epidermal Growth Factor Receptor in Oral Squamous Cell Carcinoma.

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    Jung-Hwan Lee

    Full Text Available The aim of this study is to investigate the effects of cold atmospheric pressure plasma (CAP-induced radicals on the epidermal growth factor receptor (EGFR, which is overexpressed by oral squamous cell carcinoma, to determine the underlying mechanism of selective killing. CAP-induced highly reactive radicals were observed in both plasma plume and cell culture media. The selective killing effect was observed in oral squamous cell carcinoma compared with normal human gingival fibroblast. Degradation and dysfunction of EGFRs were observed only in the EGFR-overexpressing oral squamous cell carcinoma and not in the normal cell. Nitric oxide scavenger pretreatment in cell culture media before CAP treatment rescued above degradation and dysfunction of the EGFR as well as the killing effect in oral squamous cell carcinoma. CAP may be a promising cancer treatment method by inducing EGFR dysfunction in EGFR-overexpressing oral squamous cell carcinoma via nitric oxide radicals.

  15. Selective Killing Effects of Cold Atmospheric Pressure Plasma with NO Induced Dysfunction of Epidermal Growth Factor Receptor in Oral Squamous Cell Carcinoma.

    Science.gov (United States)

    Lee, Jung-Hwan; Om, Ji-Yeon; Kim, Yong-Hee; Kim, Kwang-Mahn; Choi, Eun-Ha; Kim, Kyoung-Nam

    2016-01-01

    The aim of this study is to investigate the effects of cold atmospheric pressure plasma (CAP)-induced radicals on the epidermal growth factor receptor (EGFR), which is overexpressed by oral squamous cell carcinoma, to determine the underlying mechanism of selective killing. CAP-induced highly reactive radicals were observed in both plasma plume and cell culture media. The selective killing effect was observed in oral squamous cell carcinoma compared with normal human gingival fibroblast. Degradation and dysfunction of EGFRs were observed only in the EGFR-overexpressing oral squamous cell carcinoma and not in the normal cell. Nitric oxide scavenger pretreatment in cell culture media before CAP treatment rescued above degradation and dysfunction of the EGFR as well as the killing effect in oral squamous cell carcinoma. CAP may be a promising cancer treatment method by inducing EGFR dysfunction in EGFR-overexpressing oral squamous cell carcinoma via nitric oxide radicals.

  16. GP73-regulated oncolytic adenoviruses possess potent killing effect on human liver cancer stem-like cells

    Science.gov (United States)

    Zhang, Rong; Ma, Buyun; Liu, Tao; Yang, Yu; Xie, Wenjie; Liu, Xianglei; Huang, Fang; Liu, Tao; Zhou, Xiumei; Liu, Xinyuan; Wang, Yigang

    2016-01-01

    Cancer stem cells (CSCs), also known as tumor-initiating cells, are highly metastatic, chemo-resistant and tumorigenic, and are critical for cancer development, maintenance and recurrence. Oncolytic adenovirus could targetedly kill CSCs and has been acted as a promising anticancer agent. Currently, a novel GP73-regulated oncolytic adenovirus GD55 was constructed to specifically treat liver cancer and exhibited obvious cytotoxicity effect. However, there remains to be confirmed that whether GD55 could effectively eliminate liver CSCs. We first utilized the suspension culture to enrich the liver CSCs-like cells, which acquires the properties of liver CSCs in self-renewal, differentiation, quiescence, chemo-resistance and tumorigenicity. The results indicated that GD55 elicited more significant cytotoxicity and stronger oncolytic effect in liver CSC-like cells compared to common oncolytic virus ZD55. Additionally, GD55 possessed the greater efficacy in suppressing the growth of implanted tumors derived from liver CSC-like cells than ZD55. Furthermore, GD55 induced remarkable apoptosis of liver CSC-like cells in vitro and in vivo, and inhibited the propogation of cells and angiogenesis in xenograft tumor tissues. Thus, GD55 may virtually represent an attractive therapeutic agent for targeting liver CSCs to achieve better clinical outcomes for HCC patients. PMID:27121064

  17. Fiber mediated receptor masking in non-infected bystander cells restricts adenovirus cell killing effect but promotes adenovirus host co-existence.

    Directory of Open Access Journals (Sweden)

    Johan Rebetz

    Full Text Available The basic concept of conditionally replicating adenoviruses (CRAD as oncolytic agents is that progenies generated from each round of infection will disperse, infect and kill new cancer cells. However, CRAD has only inhibited, but not eradicated tumor growth in xenograft tumor therapy, and CRAD therapy has had only marginal clinical benefit to cancer patients. Here, we found that CRAD propagation and cancer cell survival co-existed for long periods of time when infection was initiated at low multiplicity of infection (MOI, and cancer cell killing was inefficient and slow compared to the assumed cell killing effect upon infection at high MOI. Excessive production of fiber molecules from initial CRAD infection of only 1 to 2% cancer cells and their release prior to the viral particle itself caused a tropism-specific receptor masking in both infected and non-infected bystander cells. Consequently, the non-infected bystander cells were inefficiently bound and infected by CRAD progenies. Further, fiber overproduction with concomitant restriction of adenovirus spread was observed in xenograft cancer therapy models. Besides the CAR-binding Ad4, Ad5, and Ad37, infection with CD46-binding Ad35 and Ad11 also caused receptor masking. Fiber overproduction and its resulting receptor masking thus play a key role in limiting CRAD functionality, but potentially promote adenovirus and host cell co-existence. These findings also give important clues for understanding mechanisms underlying the natural infection course of various adenoviruses.

  18. SENSITIZATION OF ACNU KILLING EFFECTS ON HeLa S3 CELLS BY MGMT ANTISENSE RNA TRANSFECTION

    Institute of Scientific and Technical Information of China (English)

    季守平; 由英; 吴英; 陈建敏; 杨军; 章扬培

    1998-01-01

    O’-methylguanlne-DNA-msthybransferase(MGMT)plays a very important role in the ceUular resistsnce to uitrosoureas drugs. Inhibition of MGMT might be a useful approach in tumor chemotherapy. In this study, the depletlon vii MGMT activity hy retroviral-mediated antisense RNA transfectkm were reported. Three retroviral vectors expressing MGMT antisense RNA were constructed and transfected into HeLa S3 cells. The difference of MGMT mRNA, MGMT activity as well as cellular resistance to ACNU before and after transtecfion were ohserved. It was found that antisense RNA targeting 5''region and whole length of MGMT mRNA could partially deplete MGMT activity and enhance killing effects of ACNU.However, 3'' region antisense RNA had no effect on MGMT modulation.

  19. Killing effect of Chinese hamster V79 cells exposed to accelerated carbon ions and RBE determination

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Survival curves of Chinese hamster V79 cells exposed to accclerated carbon ions with linear energy transfers of 125.5, 200 and 700 keV/μm were measured, respectively. Inactivation cross sections corresponding to the irradiation above were deduced from the V79 cell survival curves. They are 7.86±0.17, 10.44±1.11 and 32.32±3.58 μm2 in turn. With the surviving response of V79 cells to 60Co γ-rays as a reference value, relative biological effectiveness at 10%, 20%, 50% and 80% survival levels were given for the accelerated carbon ions. The results showed that carbon ions with LET of 125.5 keV/μm had a higher value of RBE at all the four survival levels than the carbon ions with other LETs. It was prompted that the maximum value of RBE for the V79 cell surviving as the biological endpoint emerged at the LET below 200 keV/μm for carbon ions.

  20. Killing effect of Chinese hamster V79 cells exposed to accelerated carbon ions and RBE determination

    Institute of Scientific and Technical Information of China (English)

    LIQiang; ZHOUGuang-Ming; 等

    2002-01-01

    Survival curves of Chinese hamster V79 cells exposed to accelerated carbon ions with linear energy transfers of 125.5,200 and 700keV/um were measured,respectively,Inactivation cross sections corresponding to the irradiation above were deduced from the V79 cell survival curves.They are 7.86±0.17,10.44±1.11 and 32.32±3.59um2 in turn.With the surviving response of V79 cells to 60Co γ-rays as a reference value,relative biological effectiveness at 10%,20%,50%and 80% survival levels were given for the accelerated carbon ions,The results showed that carbon ions with LET of 125.5keV/um had a higher value of RBE at all the four survival levels than the carbon ions with other LETs.It was prompted that the maximum value of RBE for the V79 cell surviving as the biological endpoint emerged at the LET below 200keV/um for carbon ions.

  1. The Effects of Arolycoricidine and Narciprimine on Tumor Cell Killing and Topoisomerase Activity

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    Buket Bozkurt Sarikaya

    2012-07-01

    Full Text Available In this study, narciprimine and arolycoricidine were isolated from G. rizehensis Stern (Amaryllidaceae. The structures of the alkaloids were elucidated by spectroscopic methods (1D NMR, EI-MS. Due to the previous reports on anti-cancer activity of this group of alkaloids, we investigated their effects on DNA topoisomerase reactions, which are known as the cellular targets of a number of chemotherapeutical drugs. The results revealed that arolycoricidine and narciprimine were effective in both type I and type II DNA topoisomerase reactions in a dose-dependent manner. Topoisomerase-interfering ability of these alkaloids partially correlated with cytostaticity assays, using HeLa (cervix adenocarcinoma, MCF7 (breast adenocarcinoma and A431 (skin epidermoid carcinoma cells. Our results are discussed in relation to the potential significance of these alkaloids in the course of drug-development studies.

  2. Immune Killing Activity of Lymphocytes on Hela Cells Expressing Interleukin-12 In Vitro

    Institute of Scientific and Technical Information of China (English)

    Huiyan WANG; Suhua CHEN

    2008-01-01

    The killing effects of lymphocytes on Hela cells expressing intedeukin-12 (IL-12) in vitro were explored. By using gene transfection technique, full length IL-12 gene was transfected into Hela cells. The expression of IL-12 in Hela cells was detected quantitatively by ELISA; Changes in killing effects of lymphocytes on Hela cells expressing IL-12 were observed by MTT. It was found that Hela cells could express IL-12 between 24h and 72h after transfection. Killing activity of lymphocytes on Hela cells expressing IL-12 was significantly enhanced. It was concluded by cell transfection technique, Hela cells could express IL-12 and were more easily killed by lymphocytes.

  3. Comparison of the killing effects between nitrogen-doped and pure TiO2 on HeLa cells with visible light irradiation

    Science.gov (United States)

    Li, Zheng; Pan, Xiaobo; Wang, Tianlong; Wang, Pei-Nan; Chen, Ji-Yao; Mi, Lan

    2013-02-01

    The killing effect of nitrogen-doped titanium dioxide (N-TiO2) nanoparticles on human cervical carcinoma (HeLa) cells by visible light photodynamic therapy (PDT) was higher than that of TiO2 nanoparticles. To study the mechanism of the killing effect, the reactive oxygen species produced by the visible-light-activated N-TiO2 and pure-TiO2 were evaluated and compared. The changes of the cellular parameters, such as the mitochondrial membrane potential (MMP), intracellular Ca2+, and nitrogen monoxide (NO) concentrations after PDT were measured and compared for N-TiO2- and TiO2-treated HeLa cells. The N-TiO2 resulted in more loss of MMP and higher increase of Ca2+ and NO in HeLa cells than pure TiO2. The cell morphology changes with time were also examined by a confocal microscope. The cells incubated with N-TiO2 exhibited serious distortion and membrane breakage at 60 min after the PDT.

  4. Targeted cytotoxic therapy kills persisting HIV infected cells during ART.

    Science.gov (United States)

    Denton, Paul W; Long, Julie M; Wietgrefe, Stephen W; Sykes, Craig; Spagnuolo, Rae Ann; Snyder, Olivia D; Perkey, Katherine; Archin, Nancie M; Choudhary, Shailesh K; Yang, Kuo; Hudgens, Michael G; Pastan, Ira; Haase, Ashley T; Kashuba, Angela D; Berger, Edward A; Margolis, David M; Garcia, J Victor

    2014-01-01

    Antiretroviral therapy (ART) can reduce HIV levels in plasma to undetectable levels, but rather little is known about the effects of ART outside of the peripheral blood regarding persistent virus production in tissue reservoirs. Understanding the dynamics of ART-induced reductions in viral RNA (vRNA) levels throughout the body is important for the development of strategies to eradicate infectious HIV from patients. Essential to a successful eradication therapy is a component capable of killing persisting HIV infected cells during ART. Therefore, we determined the in vivo efficacy of a targeted cytotoxic therapy to kill infected cells that persist despite long-term ART. For this purpose, we first characterized the impact of ART on HIV RNA levels in multiple organs of bone marrow-liver-thymus (BLT) humanized mice and found that antiretroviral drug penetration and activity was sufficient to reduce, but not eliminate, HIV production in each tissue tested. For targeted cytotoxic killing of these persistent vRNA(+) cells, we treated BLT mice undergoing ART with an HIV-specific immunotoxin. We found that compared to ART alone, this agent profoundly depleted productively infected cells systemically. These results offer proof-of-concept that targeted cytotoxic therapies can be effective components of HIV eradication strategies.

  5. Killing effect of TNF-related apoptosis inducing ligand regulated by tetracycline on gastric cancer cell line NCI-N87

    Institute of Scientific and Technical Information of China (English)

    Xiao-Chao Wei; Xin-Juan Wang; Kai-Chen; Lei Zhang; Yu Liang; Xin-Li Lin

    2001-01-01

    AIM To clone the cDNA fragment of human TRAIL (TNFrelated apoptosis inducing ligand) into a tetracyclineregulated gene expression system, the RevTet-On system, transduce expression vectors into a gastric carcinoma cell line-NCl-N87 and examine the effects of controlled expression of TRAIL in vitro on the gastric carcinoma cells. METHODS The full-length cDNA of TRAIL was inserted into a vector under the control of the tetracyclineresponsive element (TRE) to obtain the plasmid pRevTRETRAIL, which was transfected into a packaging cell line PT67. In addition, vector pRev-Tet- On and pRevTRE were also transfected into PT67 separately. After hygromycin and G418 selection, the viral titer was determined. The medium containing retroviral vectors was collected and used to transduce a gastric carcinoma cell line NCI-N87.The resulting cell line NCI-N87-Tet-On-TRE-TRAIL and a control cell line, NCI-N87-Tet-On-TRE, were established.TRAIL expression in the cell line was induced by incubating cells with doxycycline (Dox), which is a tetracycline analogue. The killing effect on gastric carcinoma cells was analyzed after induction. RESULTS The recombinant plasmid pRev-TRE-TRAIL was constructed. After hygromycin or G418 selection, the producer cell lines PT67-TRE, PT67-TRE-TRAIL and PT67TetOn were obtained, with titers of about 108CFU@ L-1 By transducing NCI-N87 cells with retroviral vectors from these cell lines, stable cell lines NCI-N87-Tet-On-TRETRAIL (NN3T) and control cell line NCI-N87-Tet-On-TRE (NN2T) were established. The growth curves of the selected cell lines were the same with the wild type NCIN87. When Dox was added, cell death was obvious in the test groups (29% -77%), whereas no difference was observed in control and wild type cell lines. With the addition of a medium from the test group, human leukemia cell line Jurkat was activated till death (83%), indicating the secretion of active TRAIL proteins from the test cells to the medium. CONCLUSION With the use of the

  6. Pseudomonas Exotoxin A: optimized by evolution for effective killing

    Directory of Open Access Journals (Sweden)

    Marta eMichalska

    2015-09-01

    Full Text Available Pseudomonas Exotoxin A (PE is the most toxic virulence factor of the pathogenic bacterium Pseudomonas aeruginosa. This review describes current knowledge about the intoxication pathways of PE. Moreover, PE represents a remarkable example for pathoadaptive evolution, how bacterial molecules have been structurally and functionally optimized under evolutionary pressure to effectively impair and kill their host cells.

  7. HIV transcription is induced with some forms of cell killing

    Energy Technology Data Exchange (ETDEWEB)

    Woloschak, G.E. [Argonne National Lab., IL (United States); Schreck, S. [Argonne National Lab., IL (United States)][South Carolina Univ., Columbia, SC (United States). Dept. of Chemistry; Panozzo, J. [Loyola Univ. Medical Center, Maywood, IL (United States); Chang-Liu, C.-M. [Argonne National Lab., IL (United States); Libertin, C.R. [Loyola Univ. Medical Center, Maywood, IL (United States)

    1996-11-01

    Using HeLa cells stably transfected with an HIV-LTR-CAT construct`, we demonstrated a peak in CAT induction that occurs in viable (but not necessarily cell-division-competent) cells 24 h following exposure to some cell-killing agents. {Gamma} rays were the only cell-killing agent which did not induce HIV transcription; this can be attributed to the fact that {gamma}-ray-induced apoptotic death requires function p53, which is missing in HeLa cells. For all other agents, HIV-LTR induction was dose-dependent and correlated with the amount of cell killing that occurred in the culture.

  8. Effects of the oral administration of viable and heat-killed Streptococcus bovis HC5 cells to pre-sensitized BALB/c mice.

    Directory of Open Access Journals (Sweden)

    Aline D Paiva

    Full Text Available Antimicrobial peptides have been suggested as an alternative to classical antibiotics in livestock production and bacteriocin-producing bacteria could be added to animal feeds to deliver bacteriocins in the gastrointestinal (GI tract of ruminant and monogastric animals. In this study, viable (V and heat-killed (HK Streptococcus bovis HC5 cells were orally administered to pre-sensitized mice in order to assess the effects of a bacteriocin-producing bacteria on histological parameters and the immune response of the GI tract of monogastric animals. The administration of V and HK S. bovis HC5 cells during 58 days to BALB/c mice did not affect weight gain, but an increase in gut permeability was detected in animals receiving the HK cells. Viable and heat killed cells caused similar morphological alterations in the GI tract of the animals, but the most prominent effects were detected in the small intestine. The oral administration of S. bovis HC5 also influenced cytokine production in the small intestine, and the immune-mediated activity differed between V and HK cells. The relative expression of IL-12 and INF-γ was significantly higher in the small intestine of mice treated with V cells, while an increase in IL-5, IL-13 and TNF-α expression was only detected in mice treated with HK cells. Considering that even under a condition of severe challenge (pre-sensitization followed by daily exposure to the same bacterial immunogen the general health of the animals was maintained, it appears that oral administration of S. bovis HC5 cells could be a useful route to deliver bacteriocin in the GI tract of livestock animals.

  9. Double suicide genes selectively kill human umbilical vein endothelial cells

    Directory of Open Access Journals (Sweden)

    Liu Lunxu

    2011-02-01

    Full Text Available Abstract Background To construct a recombinant adenovirus containing CDglyTK double suicide genes and evaluate the killing effect of the double suicide genes driven by kinase domain insert containing receptor (KDR promoter on human umbilical vein endothelial cells. Methods Human KDR promoter, Escherichia coli (E. coli cytosine deaminase (CD gene and the herpes simplex virus-thymidine kinase (TK gene were cloned using polymerase chain reaction (PCR. Plasmid pKDR-CDglyTK was constructed with the KDR promoter and CDglyTK genes. A recombinant adenoviral plasmid AdKDR-CDglyTK was then constructed and transfected into 293 packaging cells to grow and harvest adenoviruses. KDR-expressing human umbilical vein endothelial cells (ECV304 and KDR-negative liver cancer cell line (HepG2 were infected with the recombinant adenoviruses at different multiplicity of infection (MOI. The infection rate was measured by green fluorescent protein (GFP expression. The infected cells were cultured in culture media containing different concentrations of prodrugs ganciclovir (GCV and/or 5-fluorocytosine (5-FC. The killing effects were measured using two different methods, i.e. annexin V-FITC staining and terminal transferase-mediated dUTP nick end-labeling (TUNEL staining. Results Recombinant adenoviruses AdKDR-CDglyTK were successfully constructed and they infected ECV304 and HepG2 cells efficiently. The infection rate was dependent on MOI of recombinant adenoviruses. ECV304 cells infected with AdKDR-CDglyTK were highly sensitive to GCV and 5-FC. The cell survival rate was dependent on both the concentration of the prodrugs and the MOI of recombinant adenoviruses. In contrast, there were no killing effects in the HepG2 cells. The combination of two prodrugs was much more effective in killing ECV304 cells than GCV or 5-FC alone. The growth of transgenic ECV304 cells was suppressed in the presence of prodrugs. Conclusion AdKDR-CDglyTK/double prodrog system may be a useful

  10. Cell killing and mutation induction on Chinese hamster cells by photoradiations

    Energy Technology Data Exchange (ETDEWEB)

    Lam, C.K.C.

    1982-11-01

    Applying radiation directly on cells, far-uv is more effective than black light, and black light is more effective than white light in inducing proliferative death and in inducing resistance to 6-thioguanine (6-TG), ouabain and diptheria toxin (DT). Gold light has no killing and mutagenic effects on CHO (Chinese hamster ovary) cells. Use of filters showed that a small percentage of shorter wavelengths in the far-uv region is responsible for most of the killing and mutagenic effects in the unfiltered broad spectra of black and white light.

  11. Predicting In Silico Which Mixtures of the Natural Products of Plants Might Most Effectively Kill Human Leukemia Cells?

    Directory of Open Access Journals (Sweden)

    Hany A. El-Shemy

    2013-01-01

    Full Text Available The aim of the analysis of just 13 natural products of plants was to predict the most likely effective artificial mixtures of 2-3 most effective natural products on leukemia cells from over 364 possible mixtures. The natural product selected included resveratrol, honokiol, chrysin, limonene, cholecalciferol, cerulenin, aloe emodin, and salicin and had over 600 potential protein targets. Target profiling used the Ontomine set of tools for literature searches of potential binding proteins, binding constant predictions, binding site predictions, and pathway network pattern analysis. The analyses indicated that 6 of the 13 natural products predicted binding proteins which were important targets for established cancer treatments. Improvements in effectiveness were predicted for artificial combinations of 2 or 3 natural products. That effect might be attributed to drug synergism rather than increased numbers of binding proteins bound (dose effects. Among natural products, the combinations of aloe emodin with mevinolin and honokiol were predicted to be the most effective combination for AML-related predicted binding proteins. Therefore, plant extracts may in future provide more effective medicines than the single purified natural products of modern medicine, in some cases.

  12. Nanoassemblies Based on Supramolecular Complexes of Nonionic Amphiphilic Cyclodextrin and Sorafenib as Effective Weapons to Kill Human HCC Cells.

    Science.gov (United States)

    Bondì, Maria Luisa; Scala, Angela; Sortino, Giuseppe; Amore, Erika; Botto, Chiara; Azzolina, Antonina; Balasus, Daniele; Cervello, Melchiorre; Mazzaglia, Antonino

    2015-12-14

    Sorafenib (Sor), an effective chemiotherapeutic drug utilized against hepatocellular carcinoma (HCC), robustly interacts with nonionic amphiphilic cyclodextrin (aCD, SC6OH), forming, in aqueous solution, supramolecular complexes that behave as building blocks of highly water-dispersible colloidal nanoassemblies. SC6OH/Sor complex has been characterized by complementary spectroscopic techniques, such as UV-vis, steady-state fluorescence and anisotropy, resonance light scattering and (1)H NMR. The spectroscopic evidences and experiments carried out in the presence of an adamantane derivative, which competes with drug for CD cavity, agree with the entrapment of Sor in aCD, pointing out the role of the aCD cavity in the interaction between drug and amphiphile. Nanoassemblies based on SC6OH/Sor display size of ∼200 nm, negative zeta-potential (ζ = -11 mV), and both maximum loading capacity (LC ∼ 17%) and entrapment efficiency (EE ∼ 100%). Kinetic release profiles show a slower release of Sor from nanoassemblies with respect to the free drug. SC6OH/Sor nanoassemblies have very low hemolytic activity and high efficiency in vitro in decreasing cell growth and viability of HCC cell lines, such as HepG2, Hep3B, and PLC/PRF/5, opening promising chances to their in vivo applications.

  13. A compound that inhibits the HOP-Hsp90 complex formation and has unique killing effects in breast cancer cell lines.

    Science.gov (United States)

    Pimienta, Genaro; Herbert, Kristina M; Regan, Lynne

    2011-12-05

    The chaperone Hsp90 is required for the correct folding and maturation of certain "client proteins" within all cells. Hsp90-mediated folding is particularly important in cancer cells, because upregulated or mutant oncogenic proteins are often Hsp90 clients. Hsp90 inhibitors thus represent a route to anticancer agents that have the potential to be active against several different types of cancer. Currently, various Hsp90 inhibitors that bind to Hsp90 at its ATP-binding site are in preclinical and clinical trials. Some of the most promising Hsp90 ATP-binding site inhibitors are the well characterized geldanamycin derivative 17-AAG and the recently described compounds PU-H71 and NVP-AUY922. An undesirable characteristic of these compounds is the transcriptional upregulation of Hsp70 that has prosurvival effects. Here we characterize the activity of a new type of chaperone inhibitor, 1,6-dimethyl-3-propylpyrimido[5,4-e][1,2,4]triazine-5,7-dione (named C9 for simplicity). Using purified protein components in vitro, C9 prevents Hsp90 from interacting with the cochaperone HOP and is thus expected to impair the Hsp90-dependent folding pathway in vivo. We show that this compound is effective in killing various breast cancer cell lines including the highly metastatic MDA-MB-231. An important property of this compound is that it does not induce the transcriptional upregulation of Hsp70. Moreover, when cells are treated with a combination of C9 and either 17-AAG or NVP-AUY922, the overexpression of Hsp70 is counteracted considerably and C9's lethal-IC50 decreases compared to its value when added alone.

  14. IAP antagonists sensitize murine osteosarcoma cells to killing by TNFα

    Science.gov (United States)

    Shekhar, Tanmay M.; Miles, Mark A.; Gupte, Ankita; Taylor, Scott; Tascone, Brianna; Walkley, Carl R.; Hawkins, Christine J.

    2016-01-01

    Outcomes for patients diagnosed with the bone cancer osteosarcoma have not improved significantly in the last four decades. Only around 60% of patients and about a quarter of those with metastatic disease survive for more than five years. Although DNA-damaging chemotherapy drugs can be effective, they can provoke serious or fatal adverse effects including cardiotoxicity and therapy-related cancers. Better and safer treatments are therefore needed. We investigated the anti-osteosarcoma activity of IAP antagonists (also known as Smac mimetics) using cells from primary and metastatic osteosarcomas that arose spontaneously in mice engineered to lack p53 and Rb expression in osteoblast-derived cells. The IAP antagonists SM-164, GDC-0152 and LCL161, which efficiently target XIAP and cIAPs, sensitized cells from most osteosarcomas to killing by low levels of TNFα but not TRAIL. RIPK1 expression levels and activity correlated with sensitivity. RIPK3 levels varied considerably between tumors and RIPK3 was not required for IAP antagonism to sensitize osteosarcoma cells to TNFα. IAP antagonists, including SM-164, lacked mutagenic activity. These data suggest that drugs targeting XIAP and cIAP1/2 may be effective for osteosarcoma patients whose tumors express abundant RIPK1 and contain high levels of TNFα, and would be unlikely to provoke therapy-induced cancers in osteosarcoma survivors. PMID:27129149

  15. Killing effect of CIK cells and DC-CIK cells on neuroblastoma cells%CIK、DC-CIK细胞对神经母细胞瘤细胞杀伤作用的研究

    Institute of Scientific and Technical Information of China (English)

    邢准; 王秋实; 佟海侠

    2014-01-01

    Objective:To study the killing effects on neuroblastoma( NB)cells of cytokine-induced killer cells ( CIK)co-cultured with dendritic cells( DC). Methods:Peripheral blood mononuelear cells( PBMC)were isolated from healthy subjects and tumor patients. DC and CIK cells were induced by different cytokines. The phenotypes of CIK and DC cells before and after culture were determined by flow cytometry. The killing activity of different CIK groups against NB was determined by MTT. Results:The percentage of CD3+CD56+ cells as well as the cytotoxicity activity of CIK cells originated from PBMC of peripheral blood of healthy people were significantly higher than those of tumor patients(P<0. 05). In addition,compared with CIK cells,the DC-CIK cells presented a significantly higher killing effects on NB cells(P<0. 05). Conclusion:DC-CIK cell is a kind of immune cell which has a higher cyto-toxicity than that in CIK cell. CIK cells generated from PBMC of healthy subjects and tumor patients have significant differences,and provide further experimental basis for improving the clinical curative effect of CIK cells.%目的:研究细胞因子诱导的杀伤细胞( CIK)与树突状细胞( DC)共培养后对神经母细胞瘤( neuro-blastoma,NB)细胞株的杀伤作用。方法:取健康人和肿瘤患者外周血单个核细胞( PBMC),加入不同的细胞因子分别诱导出DC和CIK细胞,用流式细胞术测定诱导培养前后DC和CIK细胞的表型,MTT法测定不同组CIK细胞对NB细胞株的杀伤活性。结果:流式细胞仪检测健康人PBMC培养后CD3+CD56+淋巴细胞百分比以及对NB细胞株的杀伤活性均显著高于肿瘤患者( P<0.05)。此外,与单纯CIK细胞相比,DC-CIK细胞具有更强的杀伤NB细胞株的活性( P<0.05)。结论:DC-CIK细胞是一种细胞毒作用高于单纯CIK细胞的免疫活性细胞。健康人和肿瘤患者的PBMC经诱导培养获得的CIK细胞有显著差别,为临床进

  16. Antibacterial activity of silver-killed bacteria: the "zombies" effect

    Science.gov (United States)

    Wakshlak, Racheli Ben-Knaz; Pedahzur, Rami; Avnir, David

    2015-04-01

    We report a previously unrecognized mechanism for the prolonged action of biocidal agents, which we denote as the zombies effect: biocidally-killed bacteria are capable of killing living bacteria. The concept is demonstrated by first killing Pseudomonas aeruginosa PAO1 with silver nitrate and then challenging, with the dead bacteria, a viable culture of the same bacterium: Efficient antibacterial activity of the killed bacteria is observed. A mechanism is suggested in terms of the action of the dead bacteria as a reservoir of silver, which, due to Le-Chatelier's principle, is re-targeted to the living bacteria. Langmuirian behavior, as well as deviations from it, support the proposed mechanism.

  17. Comparative efficiency of HIV-1-infected T cell killing by NK cells, monocytes and neutrophils.

    Science.gov (United States)

    Smalls-Mantey, Adjoa; Connors, Mark; Sattentau, Quentin J

    2013-01-01

    HIV-1 infected cells are eliminated in infected individuals by a variety of cellular mechanisms, the best characterized of which are cytotoxic T cell and NK cell-mediated killing. An additional antiviral mechanism is antibody-dependent cellular cytotoxicity. Here we use primary CD4(+) T cells infected with the BaL clone of HIV-1 as target cells and autologous NK cells, monocytes, and neutrophils as effector cells, to quantify the cytotoxicity mediated by the different effectors. This was carried out in the presence or absence of HIV-1-specific antiserum to assess antibody-dependent cellular cytotoxicity. We show that at the same effector to target ratio, NK cells and monocytes mediate similar levels of both antibody-dependent and antibody-independent killing of HIV-1-infected T cells. Neutrophils mediated significant antibody-dependent killing of targets, but were less effective than monocytes or NK cells. These data have implications for acquisition and control of HIV-1 in natural infection and in the context of vaccination.

  18. Retargeting T cells for HER2-positive tumor killing by a bispecific Fv-Fc antibody.

    Directory of Open Access Journals (Sweden)

    Lei Wang

    Full Text Available To exploit the biological and pharmacological properties of immunoglobulin constant domain Fc fragment and increase the killing efficacy of T cells, a single chain variable fragment specific to CD3 was fused with Fcab (Fc antigen binding, a mutant Fc fragment with specificity against Human epidermal growth factor receptor 2 (HER2 developed by F-star. The bispecific fusion named as FcabCD3 was expressed by transient transfection in HEK-293T cells and purified by affinity chromatography. Specific cytolytic activity of retargeted T cells to kill HER2 positive SKBR3 cell line was evaluated in vitro. FcabCD3 was able to retarget T cells to kill both Herceptin insensitive Colo205-luc cell line and HER2 low expression MDA-MB-231-luc cell line. Furthermore, FcabCD3 was effective in eliminating the Colo205 tumor established on BALB/c nu/nu mice.

  19. Kinase domain insert containing receptor promotor controlled suicide gene system kills human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Zong-Hai Huang; Wen-Yu Yang; Qi Cheng; Jing-Long Yu; Zhou Li; Zong-Yan Tong; Hui-Juan Song; Xiao-Yan Che

    2005-01-01

    AIM: To evaluate the killing effect of double suicide gene mediated by adenovirus and regulated under kinase domain insert containing receptor (KDR) promoter on human umbilical vein endothelial cells. METHODS: By PCR technology, human KDR promoter gene, Escherichia coli(E. coli) cytosine deaminase (CD) gene and the herpes simple virus-thymidine kinase (TK) gene were cloned. Plasmid pKDR-CDglyTK was constructed with them. Then, a recombinant adenoviral plasmid pAdKDRCDglyTK was constructed in a "two-step transformation protocol". The newly constructed plasmids were transfected to 293 packaging cells to grow adenoviruses, which were further propagated and purified. Human umbilical vein endothelial cells (HUVEC) were infected with a different multiplicity of infection (MOI) of resultant recombinant adenovirus, the infection rate was measured with the aid of (GFP) expression. Infected cells were cultured in culture media containing different concentrations of (GCV) and/or 5-(FC), and the killing effects were measured.RESULTS: Recombinant adenoviruses AdKDR-CDglyTK were successfully constructed, and they infected HUVEC cells efficiently. Our data indicated that the infection rate was relevant to MOI of recombinant adenoviruses. HUVEC cells infected with AdKDR-CDglyTK were highly sensitive to the prodrugs, their survival rate correlated to both the concentration of the prodrugs and the MOI of recombinant adenoviruses. Our data also indicated that the two prodrugs used in combination were much more effective on killing transgeneic cells than GCV or 5-FC used alone. CONCLUSION: Prodrug/KDR-CDglyTK system is effective on killing HUVEC cells, its killing effect correlates to the concentration of prodrugs and recombinant adenovirus' MOI. Combined use of the two prodrugs confers better killing effects on transgeneic cells.

  20. Characterization of cell lysis in Pseudomonas putida induced upon expression of heterologous killing genes

    DEFF Research Database (Denmark)

    Ronchel, M.C.; Molina, L.; Witte, A.;

    1998-01-01

    Active biological containment systems are based on the controlled expression of killing genes. These systems are of interest for the Pseudomonadaceae because of the potential applications of these microbes as bioremediation agents and biopesticides, The physiological effects that lead to cell death...... upon the induction of expression of two different heterologous killing genes in nonpathogenic Pseudomonas putida KT2440 derivatives have been analyzed, P. putida CMC4 and CMC12 carry in their chromosomes a fusion of the PAl-04/03 promoter to the Escherichia coli gef gene and the phi X174 lysis gene E......, respectively. Expression of the killing genes is controlled by the LacI protein, whose expression is initiated from the XylS-dependent Pm promoter. Under induced conditions, killing of P. putida CMC12 cells mediated by phi X174 lysis protein E was faster than that observed for P. putida CMC4, for which the Gef...

  1. Combination of sulindac and dichloroacetate kills cancer cells via oxidative damage.

    Science.gov (United States)

    Ayyanathan, Kasirajan; Kesaraju, Shailaja; Dawson-Scully, Ken; Weissbach, Herbert

    2012-01-01

    Sulindac is an FDA-approved non-steroidal anti-inflammatory drug with documented anticancer activities. Our recent studies showed that sulindac selectively enhanced the killing of cancer cells exposed to oxidizing agents via production of reactive oxygen species (ROS) resulting in mitochondrial dysfunction. This effect of sulindac and oxidative stress on cancer cells could be related to the defect in respiration in cancer cells, first described by Warburg 50 years ago, known as the Warburg effect. We postulated that sulindac might enhance the selective killing of cancer cells when combined with any compound that alters mitochondrial respiration. To test this hypothesis we have used dichloroacetate (DCA), which is known to shift pyruvate metabolism away from lactic acid formation to respiration. One might expect that DCA, since it stimulates aerobic metabolism, could stress mitochondrial respiration in cancer cells, which would result in enhanced killing in the presence of sulindac. In this study, we have shown that the combination of sulindac and DCA enhances the selective killing of A549 and SCC25 cancer cells under the conditions used. As predicted, the mechanism of killing involves ROS production, mitochondrial dysfunction, JNK signaling and death by apoptosis. Our results suggest that the sulindac-DCA drug combination may provide an effective cancer therapy.

  2. Combination of sulindac and dichloroacetate kills cancer cells via oxidative damage.

    Directory of Open Access Journals (Sweden)

    Kasirajan Ayyanathan

    Full Text Available Sulindac is an FDA-approved non-steroidal anti-inflammatory drug with documented anticancer activities. Our recent studies showed that sulindac selectively enhanced the killing of cancer cells exposed to oxidizing agents via production of reactive oxygen species (ROS resulting in mitochondrial dysfunction. This effect of sulindac and oxidative stress on cancer cells could be related to the defect in respiration in cancer cells, first described by Warburg 50 years ago, known as the Warburg effect. We postulated that sulindac might enhance the selective killing of cancer cells when combined with any compound that alters mitochondrial respiration. To test this hypothesis we have used dichloroacetate (DCA, which is known to shift pyruvate metabolism away from lactic acid formation to respiration. One might expect that DCA, since it stimulates aerobic metabolism, could stress mitochondrial respiration in cancer cells, which would result in enhanced killing in the presence of sulindac. In this study, we have shown that the combination of sulindac and DCA enhances the selective killing of A549 and SCC25 cancer cells under the conditions used. As predicted, the mechanism of killing involves ROS production, mitochondrial dysfunction, JNK signaling and death by apoptosis. Our results suggest that the sulindac-DCA drug combination may provide an effective cancer therapy.

  3. Newcastle disease virus selectively kills human tumor cells.

    Science.gov (United States)

    Reichard, K W; Lorence, R M; Cascino, C J; Peeples, M E; Walter, R J; Fernando, M B; Reyes, H M; Greager, J A

    1992-05-01

    Newcastle disease virus (NDV), strain 73-T, has previously been shown to be cytolytic to mouse tumor cells. In this study, we have evaluated the ability of NDV to replicate in and kill human tumor cells in culture and in athymic mice. Plaque assays were used to determine the cytolytic activity of NDV on six human tumor cell lines, fibrosarcoma (HT1080), osteosarcoma (KHOS), cervical carcinoma (KB8-5-11), bladder carcinoma (HCV29T), neuroblastoma (IMR32), and Wilm's tumor (G104), and on nine different normal human fibroblast lines. NDV formed plaques on all tumor cells tested as well as on chick embryo cells (CEC), the native host for NDV. Plaques did not form on any of the normal fibroblast lines. To detect NDV replication, virus yield assays were performed which measured virus particles in infected cell culture supernatants. Virus yield increased 10,000-fold within 24 hr in tumor and CEC supernatants. Titers remained near zero in normal fibroblast supernatants. In vivo tumoricidal activity was evaluated in athymic nude Balb-c mice by subcutaneous injection of 9 x 10(6) tumor cells followed by intralesional injection of either live or heat-killed NDV (1.0 x 10(6) plaque forming units [PFU]), or medium. After live NDV treatment, tumor regression occurred in 10 out of 11 mice bearing KB8-5-11 tumors, 8 out of 8 with HT-1080 tumors, and 6 out of 7 with IMR-32 tumors. After treatment with heat-killed NDV no regression occurred (P less than 0.01, Fisher's exact test). Nontumor-bearing mice injected with 1.0 x 10(8) PFU of NDV remained healthy. These results indicate that NDV efficiently and selectively replicates in and kills tumor cells, but not normal cells, and that intralesional NDV causes complete tumor regression in athymic mice with a high therapeutic index.

  4. How Taxol/paclitaxel kills cancer cells.

    Science.gov (United States)

    Weaver, Beth A

    2014-09-15

    Taxol (generic name paclitaxel) is a microtubule-stabilizing drug that is approved by the Food and Drug Administration for the treatment of ovarian, breast, and lung cancer, as well as Kaposi's sarcoma. It is used off-label to treat gastroesophageal, endometrial, cervical, prostate, and head and neck cancers, in addition to sarcoma, lymphoma, and leukemia. Paclitaxel has long been recognized to induce mitotic arrest, which leads to cell death in a subset of the arrested population. However, recent evidence demonstrates that intratumoral concentrations of paclitaxel are too low to cause mitotic arrest and result in multipolar divisions instead. It is hoped that this insight can now be used to develop a biomarker to identify the ∼50% of patients that will benefit from paclitaxel therapy. Here I discuss the history of paclitaxel and our recently evolved understanding of its mechanism of action.

  5. Galactose/N-acetylgalactosamine lectin: the coordinator of host cell killing

    Indian Academy of Sciences (India)

    Douglas R Boettner; Christopher Huston; William A Petri Jr

    2002-11-01

    Entamoeba histolytica is an enteric parasite that can kill host cells via a contact-dependent mechanism. This killing involves the amoebic surface protein referred to as the Gal/GalNAc lectin. The Gal/GalNAc lectin binds galactose and N-acetylgalactosamine allowing the adherence of amoebas to host cells. Involvement of the lectin in the pathogenesis of E. histolytica infection will be reviewed in this paper. The lectin has been shown to have very specific and substantial effects on adherence, cytotoxicity, and encystation. There is also possible involvement of the lectin in phagocytosis and caspase activation in host cells.

  6. Modeling the Effects of Vorinostat In Vivo Reveals both Transient and Delayed HIV Transcriptional Activation and Minimal Killing of Latently Infected Cells.

    Science.gov (United States)

    Ke, Ruian; Lewin, Sharon R; Elliott, Julian H; Perelson, Alan S

    2015-10-01

    Recent efforts to cure human immunodeficiency virus type-1 (HIV-1) infection have focused on developing latency reversing agents as a first step to eradicate the latent reservoir. The histone deacetylase inhibitor, vorinostat, has been shown to activate HIV RNA transcription in CD4+ T-cells and alter host cell gene transcription in HIV-infected individuals on antiretroviral therapy. In order to understand how latently infected cells respond dynamically to vorinostat treatment and determine the impact of vorinostat on reservoir size in vivo, we have constructed viral dynamic models of latency that incorporate vorinostat treatment. We fitted these models to data collected from a recent clinical trial in which vorinostat was administered daily for 14 days to HIV-infected individuals on suppressive ART. The results show that HIV transcription is increased transiently during the first few hours or days of treatment and that there is a delay before a sustained increase of HIV transcription, whose duration varies among study participants and may depend on the long term impact of vorinostat on host gene expression. Parameter estimation suggests that in latently infected cells, HIV transcription induced by vorinostat occurs at lower levels than in productively infected cells. Furthermore, the estimated loss rate of transcriptionally induced cells remains close to baseline in most study participants, suggesting vorinostat treatment does not induce latently infected cell killing and thus reduce the latent reservoir in vivo.

  7. Modeling the Effects of Vorinostat In Vivo Reveals both Transient and Delayed HIV Transcriptional Activation and Minimal Killing of Latently Infected Cells.

    Directory of Open Access Journals (Sweden)

    Ruian Ke

    2015-10-01

    Full Text Available Recent efforts to cure human immunodeficiency virus type-1 (HIV-1 infection have focused on developing latency reversing agents as a first step to eradicate the latent reservoir. The histone deacetylase inhibitor, vorinostat, has been shown to activate HIV RNA transcription in CD4+ T-cells and alter host cell gene transcription in HIV-infected individuals on antiretroviral therapy. In order to understand how latently infected cells respond dynamically to vorinostat treatment and determine the impact of vorinostat on reservoir size in vivo, we have constructed viral dynamic models of latency that incorporate vorinostat treatment. We fitted these models to data collected from a recent clinical trial in which vorinostat was administered daily for 14 days to HIV-infected individuals on suppressive ART. The results show that HIV transcription is increased transiently during the first few hours or days of treatment and that there is a delay before a sustained increase of HIV transcription, whose duration varies among study participants and may depend on the long term impact of vorinostat on host gene expression. Parameter estimation suggests that in latently infected cells, HIV transcription induced by vorinostat occurs at lower levels than in productively infected cells. Furthermore, the estimated loss rate of transcriptionally induced cells remains close to baseline in most study participants, suggesting vorinostat treatment does not induce latently infected cell killing and thus reduce the latent reservoir in vivo.

  8. HAMLET kills tumor cells by apoptosis: structure, cellular mechanisms, and therapy.

    Science.gov (United States)

    Gustafsson, Lotta; Hallgren, Oskar; Mossberg, Ann-Kristin; Pettersson, Jenny; Fischer, Walter; Aronsson, Annika; Svanborg, Catharina

    2005-05-01

    New cancer treatments should aim to destroy tumor cells without disturbing normal tissue. HAMLET (human alpha-lactalbumin made lethal to tumor cells) offers a new molecular approach to solving this problem, because it induces apoptosis in tumor cells but leaves normal differentiated cells unaffected. After partial unfolding and binding to oleic acid, alpha-lactalbumin forms the HAMLET complex, which enters tumor cells and freezes their metabolic machinery. The cells proceed to fragment their DNA, and they disintegrate with apoptosis-like characteristics. HAMLET kills a wide range of malignant cells in vitro and maintains this activity in vivo in patients with skin papillomas. In addition, HAMLET has striking effects on human glioblastomas in a rat xenograft model. After convection-enhanced delivery, HAMLET diffuses throughout the brain, selectively killing tumor cells and controlling tumor progression without apparent tissue toxicity. HAMLET thus shows great promise as a new therapeutic with the advantage of selectivity for tumor cells and lack of toxicity.

  9. Recombinant scorpion insectotoxin AaIT kills specifically insect cells but not human cells

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The nucleotide sequence deduced from the amino acid sequence of the scorpion insectotoxin AaIT was chemically synthesized and was expressed in Escherichia coli. The authenticity of this in vitro expressed peptide was confirmed by N-terminal peptide sequencing. Two groups of bioassays, artificial diet incorporation assay and contact insecticidal effect assay, were carried out separately to verify the toxicity of this recombinant toxin. At the end of a 24 h experimental period, more than 60% of the testing diamondback moth (Plutella xylostella) larvae were killed in both groups with LCs0 value of 18.4 uM and 0.70 μM respectively. Cytotoxicity assay using cultured Sf9 insect cells and MCF-7 human cells demonstrated that the toxin AaIT had specific toxicity against insect cells but not human cells. Only 0.13 μM recombinant toxin was needed to kill 50% of cultured insect cells while as much as 1.3μM toxin had absolutely no effect on human cells. Insect cells produced obvious intrusions from their plasma membrane before broken up. We infer that toxin AaIT bind to a putative sodium channel in these insect cells and open the channel persistently, which would result in Na+ influx and finally cause destruction of insect cells.

  10. ABT-737 synergizes with Bortezomib to kill melanoma cells

    Directory of Open Access Journals (Sweden)

    Steven N. Reuland

    2012-02-01

    The BH3 mimetic ABT-737 is a potent inhibitor of the anti-apoptotic proteins Bcl-2, Bcl-XL, and Bcl-w. The Bcl-2 family modulates sensitivity to anticancer drugs in many cancers, including melanomas. In this study, we examined whether ABT-737 is effective in killing melanoma cells either alone or in combination with a proteasome inhibitor already in clinical use (Bortezomib in vitro and in vivo, and further evaluated the mechanisms of action. Results showed that ABT-737 alone induced modest cytotoxicity in melanoma cells, but only at higher doses. Knock-down of the anti-apoptotic proteins Bcl-2, Bcl-XL, or Mcl-1 with siRNAs demonstrated that Mcl-1 is the critical mediator of melanoma's resistance to ABT-737 treatment. However, ABT-737 displayed strong synergistic lethality when combined with Bortezomib. Immunoblot analyses demonstrated that Bortezomib increased expression of Noxa, a pro-apoptotic Bcl-2 member that antagonizes Mcl-1. Additionally, siRNA-mediated inhibition of Noxa expression protected melanoma cells from cytotoxicity induced by the combination treatment. These results demonstrate that Bortezomib synergizes with ABT-737 by neutralizing Mcl-1's function via increased levels of Noxa. In a xenograft mouse model, although drug doses were limited due to toxicity, ABT-737 or Bortezomib slowed melanoma tumor growth compared to the control, and the drug combination significantly decreased growth compared to either drug alone. These data imply that less toxic drugs fulfilling a function similar to Bortezomib to neutralize Mcl-1 are promising candidates for combination with ABT-737 for treating melanomas.

  11. Antiproliferative and proapoptotic effects of viable or heat-killed Lactobacillus paracasei IMPC2.1 and Lactobacillus rhamnosus GG in HGC-27 gastric and DLD-1 colon cell lines.

    Science.gov (United States)

    Orlando, A; Refolo, M G; Messa, C; Amati, L; Lavermicocca, P; Guerra, V; Russo, F

    2012-01-01

    Data from literature suggest the possible use of probiotics as chemopreventive agents against colon cancer, but few investigations are available on their effects on gastric cancer proliferation. In our previous study, a specific Lactobacillus, strain L. paracasei IMPC2.1, was demonstrated to colonize the human gut and positively affect fecal bacteria and biochemical parameters. The aims of the present study were to investigate the effects of L. paracasei IMPC2.1, comparing them with those of Lactobacillus rhamnosus GG (L.GG), either as viable or heat-killed cells, on cell proliferation and apoptosis in a gastric cancer (HGC-27) and a colorectal cancer cell line (DLD-1). Both the gastric and colon cancer cells were sensitive to the growth inhibition and apoptosis induction by both viable or heat-killed cells from L. paracasei IMPC2.1 and L.GG. These findings suggest the possibility for a food supplement, based on dead probiotics, including L. paracasei IMPC2.1 cells, which could represent an effective component of a functional food strategy for cancer growth inhibition, with potential for cancer prevention.

  12. EGFR-targeted magnetic nanoparticle heaters kill cancer cells without a perceptible temperature rise.

    Science.gov (United States)

    Creixell, Mar; Bohórquez, Ana C; Torres-Lugo, Madeline; Rinaldi, Carlos

    2011-09-27

    It is currently believed that magnetic nanoparticle heaters (MNHs) can kill cancer cells only when the temperature is raised above 43 °C due to energy dissipation in an alternating magnetic field. On the other hand, simple heat conduction arguments indicate that in small tumors or single cells the relative rates of energy dissipation and heat conduction result in a negligible temperature rise, thus limiting the potential of MNHs in treating small tumors and metastatic cancer. Here we demonstrate that internalized MNHs conjugated to epidermal growth factor (EGF) and which target the epidermal growth factor receptor (EGFR) do result in a significant (up to 99.9%) reduction in cell viability and clonogenic survival in a thermal heat dose dependent manner, without the need for a perceptible temperature rise. The effect appears to be cell type specific and indicates that magnetic nanoparticles in alternating magnetic fields may effectively kill cancer cells under conditions previously considered as not possible.

  13. [Killing Effect of Carpesium abrotanoides on Taenia asiatica Cysticercus].

    Science.gov (United States)

    Liu, Xiao-yan; Guo, Guang-wu; Wang, Heng

    2015-06-01

    The cysticerci of Taenia asiatica were cultured in vitro with different concentrations of water decoction of Carpesium abrotanoides (20, 40, and 60 mg/ml). The killing effect of C. abrotanoides on T. asiatica and the morphological change of cysticerci were observed under microscope 24 hours post-culture. The water decoction of C. abrotanoides showed significant killing effect on the cysticerci. The mortality of the parasites(95.0%, 57/60) was highest in 60 mg/ml group. The dead body of cysticercus shows shrunken with the enlarged scolex, and sucker tissue degenerated.

  14. Effect of Shark Liver Oil on Peritoneal Murine Macrophages in Responses to Killed-Candida albicans

    Directory of Open Access Journals (Sweden)

    Monire Hajimoradi

    2009-09-01

    Full Text Available Objective(sShark Liver Oil (SLO is an immunomodulator. Macrophages play a key role in host defense against pathogens like fungi. Candida albicans have mechanisms to escape immune system. We determined the effect of killed-Candida on the in vitro viability of macrophages and the effect of SLO on augmentation of this potency.Materials and MethodsPeritoneal macrophages were separated and cultured (3×105/well. At first, the effect of killed-Candida (200 cells/well on macrophage viability was evaluated, using MTT test. Then, MTT was performed on macrophages stimulated with killed-Candida in the presence of SLO. ResultsKilled-Candida suppressed the ability of MTT reduction and hence macrophages viability (P=0.026, but addition of SLO (100 mg/ml significantly enhanced cell viability (P=0.00. So, SLO could neutralize the inhibitory effect of Candida.ConclusionSimultaneous with cytotoxic effect of killed-Candida cells on macrophages viability, SLO augment macrophages viability. So, it can be applied in candidiasis as a complement.

  15. Mechanistic insights into selective killing of OXPHOS-dependent cancer cells by arctigenin.

    Science.gov (United States)

    Brecht, Karin; Riebel, Virginie; Couttet, Philippe; Paech, Franziska; Wolf, Armin; Chibout, Salah-Dine; Pognan, Francois; Krähenbühl, Stephan; Uteng, Marianne

    2017-04-01

    Arctigenin has previously been identified as a potential anti-tumor treatment for advanced pancreatic cancer. However, the mechanism of how arctigenin kills cancer cells is not fully understood. In the present work we studied the mechanism of toxicity by arctigenin in the human pancreatic cell line, Panc-1, with special emphasis on the mitochondria. A comparison of Panc-1 cells cultured in glucose versus galactose medium was applied, allowing assessments of effects in glycolytic versus oxidative phosphorylation (OXPHOS)-dependent Panc-1 cells. For control purposes, the mitochondrial toxic response to treatment with arctigenin was compared to the anti-cancer drug, sorafenib, which is a tyrosine kinase inhibitor known for mitochondrial toxic off-target effects (Will et al., 2008). In both Panc-1 OXPHOS-dependent and glycolytic cells, arctigenin dissipated the mitochondrial membrane potential, which was demonstrated to be due to inhibition of the mitochondrial complexes II and IV. However, arctigenin selectively killed only the OXPHOS-dependent Panc-1 cells. This selective killing of OXPHOS-dependent Panc-1 cells was accompanied by generation of ER stress, mitochondrial membrane permeabilization and caspase activation leading to apoptosis and aponecrosis.

  16. NK cell-mediated killing of AML blasts. Role of histamine, monocytes and reactive oxygen metabolites

    Energy Technology Data Exchange (ETDEWEB)

    Brune, M.; Mellqvist, U.H. [Sahlgren`s Univ. Hospital, Dept. of Medicine, Haematology Section, Goeteborg (Sweden); Hansson, M.; Hermodsson, S.; Hellstrand, K. [Sahlgren`s Univ. Hospital, Dept. of Virology, Goeteborg (Sweden)

    1996-10-01

    Blasts recovered from patients with acute myelogenous leukaemia (AML) were lysed by heterologeous natural killer (NK) cells treated with NK cell-activating cytokine-induced killing of AML blasts was inhibited by monocytes, recovered from peripheral blood by counterflow centrifugal elutriation. Histamine, at concentrations exceeding 0.1 {mu}M, abrogated the monocyte-induced inhibition of NK cells; thereby, histamine and IL-2 or histamine and IFN-{alpha} synergistically induced NK cell-mediated destruction of AML blasts. The effect of histamine was completely blocked by the histamine H2-receptor (H2R) antagonist ranitidine but not by its chemical control AH20399AA. Catalase, a scavenger of reactive oxygen metabolites (ROM), reversed the monocyte-induced inhibition of NK cell-mediated killing of blast cells, indicating that the inhibitory signal was mediated by products of the respiratory burst of monocytes. It is concluded that (i) monocytes inhibit anti-leukemic properties of NK cells, (ii) the inhibition is conveyed by monocyte-derived ROM, and (iii) histamine reverses the inhibitory signal and, thereby, synergizes with NK cell-activating cytokines to induce killing of AML blasts. (au) 19 refs.

  17. Combined effects of lactoferrin and lysozyme on Streptococcus pneumoniae killing.

    Science.gov (United States)

    André, G O; Politano, W R; Mirza, S; Converso, T R; Ferraz, L F C; Leite, L C C; Darrieux, M

    2015-12-01

    Streptococcus pneumoniae is a common colonizer of the human nasopharynx, which can occasionally spread to sterile sites, causing diseases such as otitis media, sinusitis, pneumonia, meningitis and bacteremia. Human apolactoferrin (ALF) and lysozyme (LZ) are two important components of the mucosal innate immune system, exhibiting lytic effects against a wide range of microorganisms. Since they are found in similar niches of the host, it has been proposed that ALF and LZ could act synergistically in controlling bacterial spread throughout the mucosa. The combination of ALF and LZ has been shown to enhance killing of different pathogens in vitro, with ALF facilitating the latter action of LZ. The aim of the present work was to investigate the combined effects of ALF and LZ on S pneumoniae. Concomitant addition of ALF and LZ had a synergistic killing effect on one of the pneumococci tested. Furthermore, the combination of ALF and ALZ was more bactericidal than lysozyme alone in all pneumococcal strains. Pneumococcal surface protein A (PspA), an important vaccine candidate, partially protects pneumococci from ALF mediated killing, while antibodies against one PspA enhance killing of the homologous strain by ALF. However, the serological variability of this molecule could limit the effect of anti-PspA antibodies on different pneumococci. Therefore, we investigated the ability of anti-PspA antibodies to increase ALF-mediated killing of strains that express different PspAs, and found that antisera to the N-terminal region of PspA were able to increase pneumococcal lysis by ALF, independently of the sequence similarities between the molecule expressed on the bacterial surface and that used to produce the antibodies. LF binding to the pneumococcal surface was confirmed by flow cytometry, and found to be inhibited in presence of anti-PspA antibodies. On a whole, the results suggest a contribution of ALF and LZ to pneumococcal clearance, and confirm PspA's ability to interact

  18. Mechanisms of Enhanced Cell Killing at Low Doses: Implications for Radiation Risk

    Energy Technology Data Exchange (ETDEWEB)

    Dr. Peter J. Johnston; Dr. George D. Wilson

    2003-10-15

    We have shown that cell lethality actually measured after exposure to low-doses of low-LET radiation, is markedly enhanced relative to the cell lethality previously expected by extrapolation of the high-dose cell-killing response. Net cancer risk is a balance between cell transformation and cell kill and such enhanced lethality may more than compensate for transformation at low radiation doses over a least the first 10 cGy of low-LET exposure. This would lead to a non-linear, threshold, dose-risk relationship. Therefore our data imply the possibility that the adverse effects of small radiation doses (<10 cGy) could be overestimated in specific cases. It is now important to research the mechanisms underlying the phenomenon of low-dose hypersensitivity to cell killing, in order to determine whether this can be generalized to safely allow an increase in radiation exposure limits. This would have major cost-reduction implications for the whole EM program.

  19. Polysaccharide nano-vesicular multidrug carriers for synergistic killing of cancer cells

    Science.gov (United States)

    Pramod, P. S.; Shah, Ruchira; Chaphekar, Sonali; Balasubramanian, Nagaraj; Jayakannan, Manickam

    2014-09-01

    Multi-drug delivery based on polymer nano-scaffolds is an essential protocol to be developed for better administration of anticancer drugs to enhance their therapeutic efficacies against cancer cells. Here, we report dual delivery polysaccharide nano-vesicles that are capable of loading and delivering both water soluble and water insoluble drugs together in a single polymer scaffold. The selective rupture of the nano-vesicular assembly under intracellular enzyme conditions allowed the simultaneous delivery of a hydrophobic drug camptothecin (CPT) and hydrophilic drug doxorubicin (DOX) supporting their synergistic killing of breast and colon cancer cells. The polysaccharide nano-vesicles have allowed us to address a few important questions regarding the need for multiple drug administration in cancer cells including (a) the role of simultaneous drug release, (b) antagonistic versus synergistic effects of drug combinations and (c) how these are affected by the ratio of drugs. Further, evaluation of the role of caveolae in endocytosis of these polymer scaffolds was also made. The vesicular scaffolds were found to preserve and deliver DOX resulting in 50-60% better killing of cancer cells than the free drug. Additionally, dual loaded nano-vesicles when compared to drug cocktails with individual drugs in separate nano-vesicles (at comparable molar ratios) suggest the relative drug concentration following release and mode of delivery to be both important in cancer cell killing. Results from these experiments have revealed newly developed polysaccharide nano-vesicles loaded with DOX and CPT drugs as potential candidates for improved breast cancer cell killing. Thus, these custom-designed polysaccharide nano-vesicles provide a new perspective on multi-anticancer drug delivery systems and their efficacy.Multi-drug delivery based on polymer nano-scaffolds is an essential protocol to be developed for better administration of anticancer drugs to enhance their therapeutic

  20. Photothermal-triggered control of sub-cellular drug accumulation using doxorubicin-loaded single-walled carbon nanotubes for the effective killing of human breast cancer cells

    Science.gov (United States)

    Oh, Yunok; Jin, Jun-O.; Oh, Junghwan

    2017-03-01

    Single-walled carbon nanotubes (SWNTs) are often the subject of investigation as effective photothermal therapy (PTT) agents owing to their unique strong optical absorption. Doxorubicin (DOX)-loaded SWNTs (SWNTs-DOX) can be used as an efficient therapeutic agent for combined near infrared (NIR) cancer photothermal and chemotherapy. However, SWNTs-DOX-mediated induction of cancer cell death has not been fully investigated, particularly the reaction of DOX inside cancer cells by PTT. In this study, we examined how the SWNTs-DOX promoted effective MDA-MB-231 cell death compared to DOX and PTT alone. We successfully synthesized the SWNTs-DOX. The SWNTs-DOX exhibited a slow DOX release, which was accelerated by NIR irradiation. Furthermore, DOX released from the SWNTs-DOX accumulated inside the cells at high concentration and effectively localized into the MDA-MB-231 cell nucleus. A combination of SWNTs-DOX and PTT promoted an effective MDA-MB-231 cell death by mitochondrial disruption and ROS generation. Thus, SWNTs-DOX can be utilized as an excellent anticancer agent for early breast cancer treatment.

  1. Studies of colicin action on wall-less stable L-forms of Escherichia coli. I. Degree of attachment and of killing effect on rods and stable L-form cells.

    Science.gov (United States)

    Smarda, J; Schuhmann, E

    1979-01-01

    Escherichia coli strains B and K12 W 1655 F+ are able to bind more lethal units of colicins E2, E3, G, H, Ia, and K+ X per one stable L-form cell (of the protoplast type) than per one rod cell; colicin D is bound in a higher amount on E. coli B rods. This pattern remains unchanged, if the same colicins are attached on chloroform-killed cells of both forms. Rods of both E. coli strains are more sensitive to colicins D, E2, E3, K + X (as--in the strain B--to colicin Ia) than cells of the respective L-forms. In the strain W 1655 F+ both cell forms are equally highly sensitive to colicin Ia. The stable L-forms of both strains are much more sensitive to colicins G and H than the rods. Thus the Gram-negative cell wall decreases the probability of a colicin molecule to get attached to its receptor in the cytoplasmic membrane. On the other hand, in E. coli cells the attachment of most colicin molecules to the wall receptors increases the probability of their biological effect. There is no such effect of the wall-attachment on the action of colicins G or H. The strain B is tolerant to colicin E2, while being resistant to E3; thus the cytoplasmic membrane receptor sites for them are not identical.

  2. Killing of glucose-deprived hypoxic cells with moderate hyperthermia. [HeLa cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, J.H.; Kim, S.H.; Hahn, E.W.

    1978-08-01

    Investigations were carried out to determine the importance of glucose as a modifying factor for hyperthermic cellular damage. HeLa S-3 cells were heated under oxic and hypoxic conditions, in the presence and absence of D-glucose. Temperatures as low as 41/sup 0/C selectively enhanced killing of the glucose deprived hypoxic cell. Glucose deprivation (37/sup 0/C, 2 hr) or heat alone (41/sup 0/C for 2 hr in regular medium with glucose) produced minimal cell kill under oxic or hypoxic conditions. No enhancement was seen with hyperthermia under oxic condition in the absence of glucose. The result suggests that the interplay between glucose concentration is important for hyperthermic cell kill.

  3. Merocyanine 540 and Photofrin II as photosensitizers for in vitro killing of duck hepatitis B virus and human hepatoma cells

    Science.gov (United States)

    Lin, Tsung-I.; Shien, Yong-Shau; Kao, Ming-Chien

    1994-03-01

    The feasibility of using merocyanine 540 (MC 540) and Photofrin II (PII) as effective photodynamic therapeutic (PDT) agents for killing hepatoma cells and duck hepatitis B virus (DHBV) in vitro was investigated. Cultured duck hepatocytes infected with DHBV and hepatoma cells, Hep 3B and HCC 36, were used as models. MC 540 and PII effectively inhibits the DHBV growth by 90 - 99% in a dose- and light-dependent manner. Photodynamic killing of MC 540 in the two hepatoma cell lines results in 94 - 99% growth inhibition. However, both photosensitizers exhibit dark cytotoxicity (37 - 56%). The present results suggest that MC 540 and PII could be promising and effective photodynamic agents for killing HBV and hepatoma cells.

  4. Artesunate Exerts a Direct Effect on Endothelial Cell Activation and NF-κB Translocation in a Mechanism Independent of Plasmodium Killing

    Directory of Open Access Journals (Sweden)

    Mariana C. Souza

    2012-01-01

    Full Text Available Artemisinin and its derivates are an important class of antimalarial drug and are described to possess immunomodulatory activities. Few studies have addressed the effect of artesunate in the murine malaria model or its effect on host immune response during malaria infection. Herein, we study the effect of artesunate treatment and describe an auxiliary mechanism of artesunate in modulating the inflammatory response during experimental malaria infection in mice. Treatment with artesunate did not reduce significantly the parasitemia within 12 h, however, reduced BBB breakdown and TNF-α mRNA expression in the brain tissue of artesunate-treated mice. Conversely, mefloquine treatment was not able to alter clinical features. Notably, artesunate pretreatment failed to modulate the expression of LFA-1 in splenocytes stimulated with parasitized red blood cells (pRBCs in vitro; however, it abrogated the expression of ICAM-1 in pRBC-stimulated endothelial cells. Accordingly, a cytoadherence in vitro assay demonstrated that pRBCs did not adhere to artesunate-treated vascular endothelial cells. In addition, NF-κB nuclear translocation in endothelial cells stimulated with pRBCs was impaired by artesunate treatment. Our results suggest that artesunate is able to exert a protective effect against the P. berghei-induced inflammatory response by inhibiting NF-κB nuclear translocation and the subsequent expression of ICAM-1.

  5. Combination of anti-retroviral drugs and radioimmunotherapy specifically kills infected cells from HIV infected individuals

    Directory of Open Access Journals (Sweden)

    Dina Tsukrov

    2016-09-01

    Full Text Available Eliminating virally infected cells is an essential component of any HIV eradication strategy. Radioimmunotherapy (RIT, a clinically established method for killing cells using radiolabeled antibodies, was recently applied to target HIV-1 gp41 antigen expressed on the surface of infect-ed cells. Since gp41 expression by infected cells is likely down-regulated in patients on an-tiretroviral therapy (ART, we evaluated the ability of RIT to kill ART-treated infected cells us-ing both in vitro models and lymphocytes isolated from HIV-infected subjects. Human peripheral blood mononuclear cells (PBMCs were infected with HIV and cultured in the presence of two clinically relevant ART combinations. Scatchard analysis of the 2556 human monoclonal anti-body to HIV gp41 binding to the infected and ART-treated cells demonstrated sufficient residual expression of gp41 on the cell surface to warrant subsequent RIT. This is the first time the quantification of gp41 post-ART is being reported. Cells were then treated with Bismuth-213-labeled 2556 antibody. conjugated to the human monoclonal antibody 2556, which binds to HIV gp41. Cell survival was quantified by Trypan blue and residual viremia by p24 ELISA. Cell surface gp41 expression was assessed by Scatchard analysis. The experiments were repeated using PBMCs isolated from blood specimens obtained from 15 HIV-infected individuals: ten on ART and five ART-naive. We found that 213Bi-2556 killed ART-treated infected PBMCs and reduced viral production to undetectable levels. ART and RIT co-treatment was more effective at reducing viral load in vitro than either therapy alone, indicating that gp41 expression under ART was sufficient to allow 213Bi-2556 to deliver cytocidal doses of radiation to infected cells. This study provides proof of concept that 213Bi-2556 may represent an innovative and effective targeting method for killing HIV-infected cells treated with ART, and supports continued development of 213Bi

  6. Repurposing a Prokaryotic Toxin-Antitoxin System for the Selective Killing of Oncogenically Stressed Human Cells.

    Science.gov (United States)

    Preston, Mark A; Pimentel, Belén; Bermejo-Rodríguez, Camino; Dionne, Isabelle; Turnbull, Alice; de la Cueva-Méndez, Guillermo

    2016-07-15

    Prokaryotes express intracellular toxins that pass unnoticed to carrying cells until coexpressed antitoxin partners are degraded in response to stress. Although not evolved to function in eukaryotes, one of these toxins, Kid, induces apoptosis in mammalian cells, an effect that is neutralized by its cognate antitoxin, Kis. Here we engineered this toxin-antitoxin pair to create a synthetic system that becomes active in human cells suffering a specific oncogenic stress. Inspired by the way Kid becomes active in bacterial cells, we produced a Kis variant that is selectively degraded in human cells expressing oncoprotein E6. The resulting toxin-antitoxin system functions autonomously in human cells, distinguishing those that suffer the oncogenic insult, which are killed by Kid, from those that do not, which remain protected by Kis. Our results provide a framework for developing personalized anticancer strategies avoiding off-target effects, a challenge that has been hardly tractable by other means thus far.

  7. Pan-Bcl-2 inhibitor AT-101 enhances tumor cell killing by EGFR targeted T cells.

    Directory of Open Access Journals (Sweden)

    Archana Thakur

    Full Text Available Pancreatic cancer is a deadly disease and has the worst prognosis among almost all cancers and is in dire need of new and improved therapeutic strategies. Conditioning of tumor cells with chemotherapeutic drug has been shown to enhance the anti-tumor effects of cancer vaccines and adoptive cell therapy. In this study, we investigated the immunomodulatory effects of pan-Bcl-2 inhibitor AT-101 on pancreatic cancer (PC cell cytotoxicity by activated T cells (ATC. The effects of AT-101 on cytotoxicity, early apoptosis, and Granzyme B (GrzB and IFN-γ signaling pathways were evaluated during EGFR bispecific antibody armed ATC (aATC-mediated killing of L3.6pl and MiaPaCa-2 PC cells pre-sensitized with AT-101. We found that pretreatment of tumor cells with AT-101 enhanced susceptibility of L3.6pl and MiaPaCa-2 tumor cells to ATC and aATC-mediated cytotoxicity, which was in part mediated via enhanced release of cytolytic granule GrzB from ATC and aATC. AT-101-sensitized L3.6pl cells showed up-regulation of IFN-γ-mediated induction in the phosphorylation of Ser(727-Stat1 (pS(727-Stat1, and IFN-γ induced dephosphorylation of phospho-Tyr(705-Stat3 (pY(705-Stat3. Priming (conditioning of PC cells with AT-101 can significantly enhance the anti-tumor activity of EGFRBi armed ATC through increased IFN-γ induced activation of pS(727-Stat1 and inhibition of pY(705-Stat3 phosphorylation, and resulting in increased ratio of pro-apoptotic to anti-apoptotic proteins. Our results verify enhanced cytotoxicity after a novel chemotherapy conditioning strategy against PC that warrants further in vivo and clinical investigations.

  8. Biodegradable polymeric micelle-encapsulated doxorubicin suppresses tumor metastasis by killing circulating tumor cells

    Science.gov (United States)

    Deng, Senyi; Wu, Qinjie; Zhao, Yuwei; Zheng, Xin; Wu, Ni; Pang, Jing; Li, Xuejing; Bi, Cheng; Liu, Xinyu; Yang, Li; Liu, Lei; Su, Weijun; Wei, Yuquan; Gong, Changyang

    2015-03-01

    Circulating tumor cells (CTCs) play a crucial role in tumor metastasis, but it is rare for any chemotherapy regimen to focus on killing CTCs. Herein, we describe doxorubicin (Dox) micelles that showed anti-metastatic activity by killing CTCs. Dox micelles with a small particle size and high encapsulation efficiency were obtained using a pH-induced self-assembly method. Compared with free Dox, Dox micelles exhibited improved cytotoxicity, apoptosis induction, and cellular uptake. In addition, Dox micelles showed a sustained release behavior in vitro, and in a transgenic zebrafish model, Dox micelles exhibited a longer circulation time and lower extravasation from blood vessels into surrounding tissues. Anti-tumor and anti-metastatic activities of Dox micelles were investigated in transgenic zebrafish and mouse models. In transgenic zebrafish, Dox micelles inhibited tumor growth and prolonged the survival of tumor-bearing zebrafish. Furthermore, Dox micelles suppressed tumor metastasis by killing CTCs. In addition, improved anti-tumor and anti-metastatic activities were also confirmed in mouse tumor models, where immunofluorescent staining of tumors indicated that Dox micelles induced more apoptosis and showed fewer proliferation-positive cells. There were decreased side effects in transgenic zebrafish and mice after administration of Dox micelles. In conclusion, Dox micelles showed stronger anti-tumor and anti-metastatic activities and decreased side effects both in vitro and in vivo, which may have potential applications in cancer therapy.

  9. Cryptococcus gattii is killed by dendritic cells, but evades adaptive immunity by failing to induce dendritic cell maturation.

    Science.gov (United States)

    Huston, Shaunna M; Li, Shu Shun; Stack, Danuta; Timm-McCann, Martina; Jones, Gareth J; Islam, Anowara; Berenger, Byron M; Xiang, Richard F; Colarusso, Pina; Mody, Christopher H

    2013-07-01

    During adaptive immunity to pathogens, dendritic cells (DCs) capture, kill, process, and present microbial Ags to T cells. Ag presentation is accompanied by DC maturation driven by appropriate costimulatory signals. However, current understanding of the intricate regulation of these processes remains limited. Cryptococcus gattii, an emerging fungal pathogen in the Pacific Northwest of Canada and the United States, fails to stimulate an effective immune response in otherwise healthy hosts leading to morbidity or death. Because immunity to fungal pathogens requires intact cell-mediated immunity initiated by DCs, we asked whether C. gattii causes dysregulation of DC functions. C. gattii was efficiently bound and internalized by human monocyte-derived DCs, trafficked to late phagolysosomes, and killed. Yet, even with this degree of DC activation, the organism evaded pathways leading to DC maturation. Despite the ability to recognize and kill C. gattii, immature DCs failed to mature; there was no increased expression of MHC class II, CD86, CD83, CD80, and CCR7, or decrease of CD11c and CD32, which resulted in suboptimal T cell responses. Remarkably, no increase in TNF-α was observed in the presence of C. gattii. However, addition of recombinant TNF-α or stimulation that led to TNF-α production restored DC maturation and restored T cell responses. Thus, despite early killing, C. gattii evades DC maturation, providing a potential explanation for its ability to infect immunocompetent individuals. We have also established that DCs retain the ability to recognize and kill C. gattii without triggering TNF-α, suggesting independent or divergent activation pathways among essential DC functions.

  10. A general functional response of cytotoxic T lymphocyte-mediated killing of target cells.

    Science.gov (United States)

    Gadhamsetty, Saikrishna; Marée, Athanasius F M; Beltman, Joost B; de Boer, Rob J

    2014-04-15

    Cytotoxic T lymphocytes (CTLs) kill virus-infected cells and tumor cells, and play a critical role in immune protection. Our knowledge of how the CTL killing efficiency varies with CTL and target cell numbers is limited. Here, we simulate a region of lymphoid tissue using a cellular Potts model to characterize the functional response of CTL killing of target cells, and find that the total killing rate saturates both with the CTL and the target cell densities. The relative saturation in CTL and target cell densities is determined by whether a CTL can kill multiple target cells at the same time, and whether a target cell can be killed by many CTLs together. We find that all the studied regimes can be well described by a double-saturation (DS) function with two different saturation constants. We show that this DS model can be mechanistically derived for the cases where target cells are killed by a single CTL. For the other cases, a biological interpretation of the parameters is still possible. Our results imply that this DS function can be used as a tool to predict the cellular interactions in cytotoxicity data.

  11. Killing effect of peppermint vapor against pink-slime forming microorganisms.

    Science.gov (United States)

    Ihara, Nozomi; Sakamoto, Jin; Yoshida, Munehiro; Tsuchido, Tetsuaki

    2015-01-01

    The killing effect of peppermint vapor (PMV) against pink-slime forming microorganisms, Methylobacterium mesophilicum as a bacterium and Rhodotorula mucilaginosa as a yeast, was investigated by the agar vapor assay. In this method, microbial cells were spread over the agar surface exposed to PMV in a petri dish, and then transferred into a recovery liquid. When 60μl of the peppermint liquid was added to a paper disc, a marked killing effect of PMV was observed after 48h against M. mesophilicum and after 168h against R. mucilaginosa. M. mesophilicum and R. mucilaginosa were found to be more resistant to PMV than Escherichia coli and Candida albicans, used as reference microorganisms, respectively. With the addition of 0.03% sodium pyruvate as a hydrogen peroxide scavenger in agar, the killing effect of PMV against E. coli and C. albicans was decreased, whereas it was little changed against M. mesophilicum and R. mucilaginosa. In fact, the properties of the killing effect of hydrogen peroxide solution at 0.2-1.0mM was in accord with those of PMV. M. mesophilicum and R. mucilaginosa were more resistant to the oxidant than E. coli and C. albicans, respectively. Results obtained suggested that reactive oxygen species (ROS) may be involved in the killing action of PMV and therefore pink-slime formers are more resistant to PMV than non-pink-slime formers because of the presence of carotenoids as an antioxidant in cells. We also suggest that the use of PMV appeared to be a potential tool for the control of pink-slime forming microorganisms occurring in wet areas of houses such as the bathroom and washing room.

  12. A numerical investigation of the electric and thermal cell kill distributions in electroporation-based therapies in tissue.

    Directory of Open Access Journals (Sweden)

    Paulo A Garcia

    Full Text Available Electroporation-based therapies are powerful biotechnological tools for enhancing the delivery of exogeneous agents or killing tissue with pulsed electric fields (PEFs. Electrochemotherapy (ECT and gene therapy based on gene electrotransfer (EGT both use reversible electroporation to deliver chemotherapeutics or plasmid DNA into cells, respectively. In both ECT and EGT, the goal is to permeabilize the cell membrane while maintaining high cell viability in order to facilitate drug or gene transport into the cell cytoplasm and induce a therapeutic response. Irreversible electroporation (IRE results in cell kill due to exposure to PEFs without drugs and is under clinical evaluation for treating otherwise unresectable tumors. These PEF therapies rely mainly on the electric field distributions and do not require changes in tissue temperature for their effectiveness. However, in immediate vicinity of the electrodes the treatment may results in cell kill due to thermal damage because of the inhomogeneous electric field distribution and high current density during the electroporation-based therapies. Therefore, the main objective of this numerical study is to evaluate the influence of pulse number and electrical conductivity in the predicted cell kill zone due to irreversible electroporation and thermal damage. Specifically, we simulated a typical IRE protocol that employs ninety 100-µs PEFs. Our results confirm that it is possible to achieve predominant cell kill due to electroporation if the PEF parameters are chosen carefully. However, if either the pulse number and/or the tissue conductivity are too high, there is also potential to achieve cell kill due to thermal damage in the immediate vicinity of the electrodes. Therefore, it is critical for physicians to be mindful of placement of electrodes with respect to critical tissue structures and treatment parameters in order to maintain the non-thermal benefits of electroporation and prevent

  13. A numerical investigation of the electric and thermal cell kill distributions in electroporation-based therapies in tissue.

    Science.gov (United States)

    Garcia, Paulo A; Davalos, Rafael V; Miklavcic, Damijan

    2014-01-01

    Electroporation-based therapies are powerful biotechnological tools for enhancing the delivery of exogeneous agents or killing tissue with pulsed electric fields (PEFs). Electrochemotherapy (ECT) and gene therapy based on gene electrotransfer (EGT) both use reversible electroporation to deliver chemotherapeutics or plasmid DNA into cells, respectively. In both ECT and EGT, the goal is to permeabilize the cell membrane while maintaining high cell viability in order to facilitate drug or gene transport into the cell cytoplasm and induce a therapeutic response. Irreversible electroporation (IRE) results in cell kill due to exposure to PEFs without drugs and is under clinical evaluation for treating otherwise unresectable tumors. These PEF therapies rely mainly on the electric field distributions and do not require changes in tissue temperature for their effectiveness. However, in immediate vicinity of the electrodes the treatment may results in cell kill due to thermal damage because of the inhomogeneous electric field distribution and high current density during the electroporation-based therapies. Therefore, the main objective of this numerical study is to evaluate the influence of pulse number and electrical conductivity in the predicted cell kill zone due to irreversible electroporation and thermal damage. Specifically, we simulated a typical IRE protocol that employs ninety 100-µs PEFs. Our results confirm that it is possible to achieve predominant cell kill due to electroporation if the PEF parameters are chosen carefully. However, if either the pulse number and/or the tissue conductivity are too high, there is also potential to achieve cell kill due to thermal damage in the immediate vicinity of the electrodes. Therefore, it is critical for physicians to be mindful of placement of electrodes with respect to critical tissue structures and treatment parameters in order to maintain the non-thermal benefits of electroporation and prevent unnecessary damage to

  14. Cytotoxic T-lymphocyte-mediated killing of human pancreatic islet cells in vitro.

    Science.gov (United States)

    Campbell, Peter D; Estella, Eugene; Dudek, Nadine L; Jhala, Gaurang; Thomas, Helen E; Kay, Thomas W H; Mannering, Stuart I

    2008-09-01

    Cytotoxic T lymphocytes (CTL) are believed to play an essential role in beta-cell destruction leading to development of type 1 diabetes and allogeneic islet graft failure. We aimed to identify the mechanisms used by CTL to kill human beta cells. CTL clones that recognize epitopes from influenza virus and Epstein-Barr virus restricted by human leukocyte antigen (HLA)-A0201 and -B0801, respectively, were used to investigate the susceptibility of human beta cells to CTL. In a short-term (5-hour) assay, CTL killed human islet cells of the appropriate major histocompatibility complex (MHC) class I type that had been pulsed with viral peptides. Killing was increased by pretreating islets with interferon gamma that increases MHC class I on target cells. Killing was abolished by incubation of CTL with the perforin inhibitor concanamycin A. The Fas pathway did not contribute to killing because blocking with neutralizing anti-Fas ligand antibody did not significantly reduce beta-cell killing. In conclusion, we report a novel way of investigating the interaction between CTL and human islets. Human islets were rapidly killed in vitro by MHC class I-restricted CTL predominantly by the granule exocytosis pathway.

  15. Synthesis of multivalent glycoconjugates containing the immunoactive LELTE peptide: effect of glycosylation on cellular activation and natural killing by human peripheral blood mononuclear cells.

    Science.gov (United States)

    Renaudet, Olivier; Krenek, Karel; Bossu, Isabelle; Dumy, Pascal; Kádek, Alan; Adámek, David; Vanek, Ondrej; Kavan, Daniel; Gazák, Radek; Sulc, Miroslav; Bezouska, Karel; Kren, Vladimír

    2010-05-19

    Pentapeptide diacidic sequence LELTE, derived from the mycobacterial heat shock protein hsp65, has been recently identified as a "danger" signal of the immune system effective via specific binding to the universal leukocyte triggering receptor CD69. This sequence is not active per se, only after its presentation within the multivalent environment of its parent protein, or after artificial dimerization using a standard bifunctional reagents. Here we describe an entirely new way of presenting of this peptide based on its attachment to a cyclopeptide RAFT scaffold (K-K-K-P-G)(2) through the epsilon-amino group of lysine residues, alone or in combination with the carbohydrate epitope alphaGalNAc. The ability of such RAFT scaffolds to precipitate the target CD69 receptor or to activate CD69-positive cells is enhanced in compounds 2 and 4 possessing combined peptide/carbohydrate expression. Compounds 2 and 4 are highly efficient activators of natural killer lymphocytes, but they are completely inactive from the point of view of activation-induced apoptosis of lymphocytes by the target cells. These unique properties make the combined peptide/carbohydrate RAFTs highly suitable for future evaluation in animal tumor therapies in vivo and predict them to be readily available and efficient immunoactivators.

  16. Advanced application of bovine intestinal epithelial cell line for evaluating regulatory effect of lactobacilli against heat-killed enterotoxigenicEscherichia coli-mediated inflammation

    OpenAIRE

    Takanashi, Naoya; Tomosada, Yohsuke; Villena, Julio Cesar; Murata, Kozue; Takahashi, Takuya; Chiba, Eriko; Tohno, Masanori; Tomoyuki Shimazu; Aso, Hisashi; Suda, Yoshihito; Ikegami, Shuji; Itoh, Hiroyuki; Kawai, Yasushi; Tadao Saito; Alvarez, Gladis Susana

    2015-01-01

    Background: Previously, a bovine intestinal epithelial cell line (BIE cells) was successfully established. This work hypothesized that BIE cells are useful in vitro model system for the study of interactions of microbial- or pathogenassociated molecular patterns (MAMPs or PAMPs) with bovine intestinal epithelial cells and for the selection of immunoregulatory lactic acid bacteria (LAB). Results: All toll-like receptor (TLR) genes were expressed in BIE cells, being TLR4 one of the most strong...

  17. Sulindac enhances the killing of cancer cells exposed to oxidative stress.

    Directory of Open Access Journals (Sweden)

    Maria Marchetti

    Full Text Available BACKGROUND: Sulindac is an FDA-approved non-steroidal anti-inflammatory drug (NSAID that affects prostaglandin production by inhibiting cyclooxygenases (COX 1 and 2. Sulindac has also been of interest for more than decade as a chemopreventive for adenomatous colorectal polyps and colon cancer. PRINCIPAL FINDINGS: Pretreatment of human colon and lung cancer cells with sulindac enhances killing by an oxidizing agent such as tert-butyl hydroperoxide (TBHP or hydrogen peroxide. This effect does not involve cyclooxygenase (COX inhibition. However, under the conditions used, there is a significant increase in reactive oxygen species (ROS within the cancer cells and a loss of mitochondrial membrane potential, suggesting that cell death is due to apoptosis, which was confirmed by Tunel assay. In contrast, this enhanced killing was not observed with normal lung or colon cells. SIGNIFICANCE: These results indicate that normal and cancer cells handle oxidative stress in different ways and sulindac can enhance this difference. The combination of sulindac and an oxidizing agent could have therapeutic value.

  18. Different Length (DL qPCR for Quantification of Cell Killing by UV-induced DNA Damage

    Directory of Open Access Journals (Sweden)

    Guro Skjefstad

    2010-08-01

    Full Text Available We describe the different length (DL qPCR method for quantification of UV induced DNA damage in cell killing. The principle of DL qPCR is that DNA damage inhibits PCR. Applications with different lengths can therefore be used to detect different levels of UV-induced DNA damage. The assay was evaluated on three strains of Escherichia coli exposed to varying levels of ultraviolet (UV radiation. We show that DL qPCR sensitivity and reproducibility are within the range of practical application to detect the effect of UV cell killing.

  19. Interactions between neutrophils and macrophages promote macrophage killing of rat muscle cells in vitro

    Science.gov (United States)

    Nguyen, Hal X.; Tidball, James G.

    2003-01-01

    Current evidence indicates that the physiological functions of inflammatory cells are highly sensitive to their microenvironment, which is partially determined by the inflammatory cells and their potential targets. In the present investigation, interactions between neutrophils, macrophages and muscle cells that may influence muscle cell death are examined. Findings show that in the absence of macrophages, neutrophils kill muscle cells in vitro by superoxide-dependent mechanisms, and that low concentrations of nitric oxide (NO) protect against neutrophil-mediated killing. In the absence of neutrophils, macrophages kill muscle cells through a NO-dependent mechanism, and the presence of target muscle cells causes a three-fold increase in NO production by macrophages, with no change in the concentration of inducible nitric oxide synthase. Muscle cells that are co-cultured with both neutrophils and macrophages in proportions that are observed in injured muscle show cytotoxicity through a NO-dependent, superoxide-independent mechanism. Furthermore, the concentration of myeloid cells that is necessary for muscle killing is greatly reduced in assays that use mixed myeloid cell populations, rather than uniform populations of neutrophils or macrophages. These findings collectively show that the magnitude and mechanism of muscle cell killing by myeloid cells are modified by interactions between muscle cells and neutrophils, between muscle cells and macrophages and between macrophages and neutrophils.

  20. 3,4-二苯基香豆素衍生物的合成及其促自然杀伤细胞活性研究%Synthesis of 3,4-Biphenyl Coumarin Derivatives and Their Promoting Effects on the Killing Activity of Natural Killing Cell

    Institute of Scientific and Technical Information of China (English)

    陈华; 周利凯; 李帅; 姚玉超; 谷云景; 李春晓; 李娜; 孟明; 李小六

    2013-01-01

    以水杨酸为起始原料,经Fridel-Crafts酰基化反应、微波促进的Perkin反应和两步亲核取代反应,合成了具有氨基烷氧侧链的3,4-二苯基香豆素衍生物1a~1e.初步生物活性测试表明,化合物1a~1e能显著促进自然杀伤细胞杀伤活性,具有较强的免疫调节活性.%A series of novel 3,4-biphenyl coumarin derivatives containing an amino side chain 1a~1e were synthesized using salicylic acid as starting material via Fridel-Crafts acylation, then the microwave-assisted Perkin reaction and double SN2 substitution reactions. The preliminary biological evaluation showed that compounds 1a~1e had significant promoting effects on the killing activity of natural killer cell, and hold potential as immunomodulating agents.

  1. The Smac Mimetic BV6 Improves NK Cell-Mediated Killing of Rhabdomyosarcoma Cells by Simultaneously Targeting Tumor and Effector Cells

    Science.gov (United States)

    Fischer, Kyra; Tognarelli, Sara; Roesler, Stefanie; Boedicker, Cathinka; Schubert, Ralf; Steinle, Alexander; Klingebiel, Thomas; Bader, Peter; Fulda, Simone; Ullrich, Evelyn

    2017-01-01

    Rhabdomyosarcoma (RMS), the most common cancer of connective tissues in pediatrics, is often resistant to conventional therapies. One underlying mechanism of this resistance is the overexpression of Inhibitor of Apoptosis (IAP) proteins, leading to a dysfunctional cell death program within tumor cells. Smac mimetics (SM) are small molecules that can reactivate the cell death program by antagonizing IAP proteins and thereby compensating their overexpression. Here, we report that SM sensitize two RMS cell lines (RD and RH30) toward natural killer (NK) cell-mediated killing on the one hand, and increase the cytotoxic potential of NK cells on the other. The SM-induced sensitization of RH30 cells toward NK cell-mediated killing is significantly reduced through blocking tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on NK cells prior to coculture. In addition, the presence of zVAD.fmk, a pancaspase inhibitor, rescues tumor cells from the increase in killing, indicating an apoptosis-dependent cell death. On the NK cell side, the presence of SM in addition to IL-2 during the ex vivo expansion leads to an increase in their cytotoxic activity against RH30 cells. This effect is mainly TNFα-dependent and partially mediated by NK cell activation, which is associated with transcriptional upregulation of NF-κB target genes such as IκBα and RelB. Taken together, our findings implicate that SM represent a novel double-hit strategy, sensitizing tumor and activating NK cells with one single drug.

  2. VP22-CD基因工程化载体对胶质瘤细胞体外杀伤作用的研究%Effect of VP22-CD genetically engineered vectors on killing glioma cells in vitro

    Institute of Scientific and Technical Information of China (English)

    米蕊芳; 金贵善; 张俊文; 周益强; 徐恒周; 程森; 刘福生

    2016-01-01

    目的 构建CD以及VP22-CD基因工程化载体,并探讨其在体外对胶质瘤细胞的杀伤作用.方法 利用聚合酶链式反应(PCR)得到CD以及VP22基因片段;利用In-fusion基因克隆方法将得到的基因片段插入到pEGFP-N1载体中,构建CD以及VP22-CD基因工程化载体;利用脂质体转染的方法将构建的基因工程化载体转染到胶质瘤细胞C6及U87细胞中,并通过体外光镜及荧光显微镜观察转染率;利用MTT比色法评价加入前药5-氟胞嘧啶后对转染后胶质瘤细胞的杀伤率.结果 (1)将CD基因或VP22-CD融合基因插入到pEGFP-CD载体中,构建得到pEGFP-CD以及pEGFP-VP22-CD载体.(2)pEGFP-CD及pEGFP-VP22-CD载体可转染入C6或U87胶质瘤细胞,转染率分别为2.5%和3.7%以及2.0%和4.1%.(3)在C6或U87细胞中转染pEGFP-CD或pEGFP-VP22-CD载体并加入前药5-氟胞嘧啶后,细胞的存活率分别为(30.36±0.63)%和(11.75±1.01)%以及(28.78±3.62)%和(18.26±2.27)%,与转染pEGFP-N1组相比差异均有统计学意义(均P<0.05).(4)在C6或U87细胞中加入前药5-氟胞嘧啶后,转染pEGFP-VP22-CD组与转染pEGFP-CD组相比差异均有统计学意义(均P<0.05).结论 CD/5-FC系统在体外对恶性胶质瘤细胞具有显著的杀伤作用和旁观者效应;并且穿梭蛋白VP22可进一步促进系统对细胞的杀伤作用.%Objectives To construct CD and VP22-CD genetically engineered vectors and to investigate their killing effects on glioma cells in vitro.Methods CD and VP22 gene segments were obtain using polymerase chain reaction (PCR).The obtained gene fragments were inserted into pEGFP-N1 vectors using in-fusion gene cloning method.CD and VP22-CD gene engineering vectors were constructed.The constructed gene engineering vectors were transfected into the glioma cells C6 and U87 cells using the liposome transfection method,and the transfection efficiency was observed through in vitro light microscope and fluorescence microscope.The kill

  3. T cells kill bacteria captured by transinfection from dendritic cells and confer protection in mice.

    Science.gov (United States)

    Cruz-Adalia, Aránzazu; Ramirez-Santiago, Guillermo; Calabia-Linares, Carmen; Torres-Torresano, Mónica; Feo, Lidia; Galán-Díez, Marta; Fernández-Ruiz, Elena; Pereiro, Eva; Guttmann, Peter; Chiappi, Michele; Schneider, Gerd; Carrascosa, José López; Chichón, Francisco Javier; Martínez Del Hoyo, Gloria; Sánchez-Madrid, Francisco; Veiga, Esteban

    2014-05-14

    Dendritic cells (DCs) phagocytose, process, and present bacterial antigens to T lymphocytes to trigger adaptive immunity. In vivo, bacteria can also be found inside T lymphocytes. However, T cells are refractory to direct bacterial infection, leaving the mechanisms by which bacteria invade T cells unclear. We show that T cells take up bacteria from infected DCs by the process of transinfection, which requires direct contact between the two cells and is enhanced by antigen recognition. Prior to transfer, bacteria localize to the immunological synapse, an intimate DC/T cell contact structure that activates T cells. Strikingly, T cells efficiently eliminate the transinfecting bacteria within the first hours after infection. Transinfected T cells produced high levels of proinflammatory cytokines and were able to protect mice from bacterial challenge following adoptive transfer. Thus, T lymphocytes can capture and kill bacteria in a manner reminiscent of innate immunity.

  4. Poliovirus protease 3C(pro) kills cells by apoptosis.

    Science.gov (United States)

    Barco, A; Feduchi, E; Carrasco, L

    2000-01-20

    The tetracycline-based Tet-Off expression system has been used to analyze the effects of poliovirus protease 3C(pro) on human cells. Stable HeLa cell clones that express this poliovirus protease under the control of an inducible, tightly regulated promoter were obtained. Tetracycline removal induces synthesis of 3C protease, followed by drastic morphological alterations and cellular death. Degradation of cellular DNA in nucleosomes and generation of apoptotic bodies are observed from the second day after 3C(pro) induction. The cleavage of poly(ADP-ribose) polymerase, an enzyme involved in DNA repair, occurs after induction of 3C(pro), indicating caspase activation by this poliovirus protease. The 3C(pro)-induced apoptosis is blocked by the caspase inhibitor z-VAD-fmk. Our findings suggest that the protease 3C is responsible for triggering apoptosis in poliovirus-infected cells by a mechanism that involves caspase activation.

  5. Protection of Candida parapsilosis from neutrophil killing through internalization by human endothelial cells.

    Science.gov (United States)

    Glass, Kyle A; Longley, Sarah J; Bliss, Joseph M; Shaw, Sunil K

    2015-01-01

    Candida parapsilosis is a fungal pathogen that is associated with hematogenously disseminated disease in premature neonates, acutely ill or immunocompromised patients. In cell culture, C. parapsilosis cells are actively and avidly endocytosed by endothelial cells via actin polymerization mediated by N-WASP. Here we present evidence that C. parapsilosis that were internalized by endothelial cells remained alive, and avoided being acidified or otherwise damaged via the host cell. Internalized fungal cells reproduced intracellularly and eventually burst out of the host endothelial cell. When neutrophils were added to endothelium and C. parapsilosis, they patrolled the endothelial surface and efficiently killed most adherent fungal cells prior to endocytosis. But after endocytosis by endothelial cells, internalized fungal cells evaded neutrophil killing. Silencing endothelial N-WASP blocked endocytosis of C. parapsilosis and left fungal cells stranded on the cell surface, where they were susceptible to neutrophil killing. These observations suggest that for C. parapsilosis to escape from the bloodstream, fungi may adhere to and be internalized by endothelial cells before being confronted and phagocytosed by a patrolling leukocyte. Once internalized by endothelial cells, C. parapsilosis may safely replicate to cause further rounds of infection. Immunosurveillance of the intravascular lumen by leukocytes crawling on the endothelial surface and rapid killing of adherent yeast may play a major role in controlling C. parapsilosis dissemination and infected endothelial cells may be a significant reservoir for fungal persistence.

  6. Cell kill by megavoltage protons with high LET

    Science.gov (United States)

    Kuperman, Vadim Y.

    2016-07-01

    The aim of the current study is to develop a radiobiological model which describes the effect of linear energy transfer (LET) on cell survival and relative biological effectiveness (RBE) of megavoltage protons. By assuming the existence of critical sites within a cell, analytical expression for cell survival S as a function of LET is derived. The obtained results indicate that in cases where dose per fraction is small, \\ln (S) is a linear-quadratic (LQ) function of dose while both alpha and beta radio-sensitivities are non-linearly dependent on LET. In particular, in the current model alpha increases with increasing LET while beta decreases. Conversely, in the case of large dose per fraction, the LQ dependence of \\ln (S) on dose is invalid. The proposed radiobiological model predicts cell survival probability and RBE which, in general, deviate from the results obtained by using conventional LQ formalism. The differences between the LQ model and that described in the current study are reflected in the calculated RBE of protons.

  7. HER2-Specific T Lymphocytes Kill both Trastuzumab-Resistant and Trastuzumab-Sensitive Breast Cell Lines In Vitro

    Institute of Scientific and Technical Information of China (English)

    Xiao-lin Lin; Xu Liang; Tao Shen; Jun Ren; Xiao-li Wang; Bo Ma; Jun Jia; Ying Yan; Li-jun Di; Yan-hua Yuan; Feng-ling Wan; Yuan-li Lu

    2012-01-01

    Objective:Although the development of trastuzumab has improved the outlook for women with human epidermal growth factor receptor 2 (HER2)-positive breast cancer,the resistance to anti-HER2 therapy is a growing clinical dilemma.We aim to determine whether HER2-specific T cells generated from dendritic cells (DCs) modified with HER2 gene could effectively kill the HER2-positive breast cancer cells,especially the trastuzumab-resistant cells.Methods:The peripheral blood mononuclear cells (PBMCs) from healthy donors,whose HLA haplotypes were compatible with the tumor cell lines,were transfected with reconstructive human adeno-association virus (rhAAV/HER2) to obtain the specific killing activities of T cells,and were evaluated by lactate dehydrogenase (LDH)releasing assay.Results:Trastuzumab produced a significant inhibiting effect on SK-BR-3,the IC50 was 100ng/ml.MDA-MB-453 was resistant to trastuzumab even at a concentration of 10,000 ng/ml in vitro.HER2-specific T lymphocytes killed effectively SK-BR-3 [(69.86±13.41)%] and MDA-MB-453 [(78.36±10.68)%] at 40:1 (effector:target ratio,E:T),but had no significant cytotoxicity against HER2-negative breast cancer cell lines MDA-MB-231 or MCF-7 (less than 10%).Conclusion:The study showed that HER2-specific T lymphocytes generated from DCs modified by rhAAV/HER2 could kill HER2-positive breast cancer cell lines in a HER2-dependent manner,and result in significantly high inhibition rates on the intrinsic trastuzumab-resistant cell line MDA-MB-453 and the tastuzumab-sensitive cell line SK-BR-3.These results imply that this immunotherapy might be a potential treatment to HER2-positive breast cancer.

  8. Ascorbic acid kills Epstein-Barr virus positive Burkitt lymphoma cells and Epstein-Barr virus transformed B-cells in vitro, but not in vivo.

    Science.gov (United States)

    Shatzer, Amber N; Espey, Michael Graham; Chavez, Mayra; Tu, Hongbin; Levine, Mark; Cohen, Jeffrey I

    2013-05-01

    Ascorbic acid has been shown to kill various cancer cell lines at pharmacologic concentrations. We found that Epstein-Barr virus (EBV)-positive Burkitt lymphoma (BL) cells were more susceptible to ascorbic acid-induced cell killing than EBV-negative BL cells or EBV-transformed lymphoblastoid cells (LCLs). Ascorbic acid did not induce apoptosis in any of the tested cells but did induce the production of reactive oxygen species and cell death. Previously, we showed that bortezomib, a proteasome inhibitor, induces cell death in LCLs and EBV-positive BL cells. We found that ascorbic acid is strongly antagonistic for bortezomib-induced cell death in LCLs and EBV-positive BL cells. Finally, ascorbic acid did not prolong survival of severe combined immunodefiency mice inoculated with LCLs either intraperitoneally or subcutaneously. Thus, while ascorbic acid was highly effective at killing EBV-positive BL cells and LCLs in vitro, it antagonized cell killing by bortezomib and was ineffective in an animal model.

  9. Surface expression of HSP72 by LPS-stimulated neutrophils facilitates gammadeltaT cell-mediated killing.

    Science.gov (United States)

    Hirsh, Mark I; Hashiguchi, Naoyuki; Chen, Yu; Yip, Linda; Junger, Wolfgang G

    2006-03-01

    During inflammation and sepsis, accumulation of activated neutrophils causes lung tissue damage and organ failure. Effective clearance of neutrophils reduces the risk of organ failure; however, its mechanisms are poorly understood. Because lungs are rich in gammadeltaT cells, we investigated the physiological role of these cells in the protection of lung tissue from infiltrating neutrophils. In a mouse model of sepsis, we found that the lungs of survivors contained significantly higher numbers of gammadeltaT cells than those of mice that died from sepsis. The number of gammadeltaT cells correlated inversely with the number of neutrophils in the lungs and with the degree of lung tissue damage. LPS rapidly elicited the expression of heat shock protein (HSP) 72 on the surface of human neutrophils. Inhibitors of transcription, protein synthesis, and intracellular protein transport blocked HSP72 expression, indicating that de novo synthesis is required. gammadeltaT cells targeted and rapidly killed LPS-treated neutrophils through direct cell-to-cell contact. Pre-treatment with neutralizing antibodies to HSP72 diminished neutrophil killing. Our data indicate that HSP72 expression on the cell surface predisposes inflamed neutrophils to killing by gammadeltaT cells. This intercellular exchange may allow gammadeltaT cells to resolve inflammation and limit host tissue damage during sepsis.

  10. Strong synergy of heat and modulated electromagnetic field in tumor cell killing

    Energy Technology Data Exchange (ETDEWEB)

    Andocs, Gabor [Frederic Joliot Curie National Research Inst. for Radiobiology and Radiohygiene, Budapest (Hungary)]|[St. Istvan Univ., Budapest (Hungary). Dept. of Pharmacology and Toxicology; Renner, Helmut [Klinikum Nuernberg (Germany). Clinic of Radiooncology; Balogh, Lajos [Frederic Joliot Curie National Research Inst. for Radiobiology and Radiohygiene, Budapest (Hungary); Fonyad, Laszlo [Semmelweis Univ., Budapest (Hungary). 1. Dept. of of Pathology and Experimental Cancer Research; Jakab, Csaba [St. Istvan Univ., Budapest (Hungary). Dept. of Pathology; Szasz, Andras [St. Istvan Univ., Goedoelloe (Hungary). Biotechnics Dept.

    2009-02-15

    Hyperthermia is an emerging complementary method in radiooncology. Despite many positive studies and comprehensive reviews, the method is not widely accepted as a combination to radiotherapy. Modulated electrohyperthermia (mEHT; capacitive, electric field modulated, 13.56 MHz) has been used in clinical practice for almost 2 decades in Germany, Austria and Hungary. This in vivo study in nude mice xenograft tumors compares mEHT with 'classic' radiative hyperthermia (radHT). Nude mice were xenografted with HT29 human colorectal carcinoma cells. 28 mice in four groups with seven animals each and two tumors per animal (totally 56 tumors) were included in the present study: group 1 as untreated control; group 2 treated with radHT at 42 C; group 3 treated with mEHT at identical 42 C; group 4 treated with mEHT at 38 C (by intensively cooling down the tumor). 24 h after treatment, animals were sacrificed and the tumor cross sections studied by precise morphological methods for the respective relative amount of 'dead' tumor cells. The effect of mEHT established a double effect as a synergy between the purely thermal (temperature-dependent) and nonthermal (not directly temperature-dependent) effects. The solely thermal enhancement ratio (TER) of cell killing was shown to be 2.9. The field enhancement ratio (FER) at a constant temperature of 42 C was measured as 3.2. Their complex application significantly increased the therapeutic enhancement to 9.4. mEHT had a remarkable cancer cell-killing effect in a nude mice xenograft model. (orig.)

  11. Killing of Brain Tumor Cells by Hypoxia-Responsive Element Mediated Expression of BAX

    Directory of Open Access Journals (Sweden)

    Hangjun Ruan

    1999-11-01

    Full Text Available The presence of radioresistant hypoxic cells in human brain tumors limits the overall effectiveness of conventional fractionated radiation therapy. Tumor-specific therapies that target hypoxic cells are clearly needed. We have investigated the expression of suicide genes under hypoxia by a hypoxia-responsive element (HRE, which can be activated through hypoxia-inducible factor-1 (HIF-1. We transfected plasmids containing multiple copies of HIRE into U-87 MG and U-251 MG-NCI human brain tumor cells and tested their ability to induce LacZ gene expression under anoxia. Gene expression under anoxia versus oxia was increased about 12-fold for U-87 MG cells and about fourfold for U-251 MG-NCI cells. At intermediate hypoxic conditions, increased LacZ gene expression in U-87 MG cells was induced by the plasmid that contained three HREs, but not by the plasmid with two HREs. Lastly, when we placed a suicide gene BAX under the control of HREs, cells transfected with the BAX plasmids were preferentially killed through apoptosis under anoxia. Our studies demonstrate that HRE-regulated gene expression is active in brain tumor cells, and that the amount of increased gene expression obtained is dependent on the cell line, the HIRE copy number, and the degree of hypoxia.

  12. Syrosingopine sensitizes cancer cells to killing by metformin

    Science.gov (United States)

    Benjamin, Don; Colombi, Marco; Hindupur, Sravanth K.; Betz, Charles; Lane, Heidi A.; El-Shemerly, Mahmoud Y. M.; Lu, Min; Quagliata, Luca; Terracciano, Luigi; Moes, Suzette; Sharpe, Timothy; Wodnar-Filipowicz, Aleksandra; Moroni, Christoph; Hall, Michael N.

    2016-01-01

    We report that the anticancer activity of the widely used diabetic drug metformin is strongly potentiated by syrosingopine. Synthetic lethality elicited by combining the two drugs is synergistic and specific to transformed cells. This effect is unrelated to syrosingopine’s known role as an inhibitor of the vesicular monoamine transporters. Syrosingopine binds to the glycolytic enzyme α-enolase in vitro, and the expression of the γ-enolase isoform correlates with nonresponsiveness to the drug combination. Syrosingopine sensitized cancer cells to metformin and its more potent derivative phenformin far below the individual toxic threshold of each compound. Thus, combining syrosingopine and codrugs is a promising therapeutic strategy for clinical application for the treatment of cancer. PMID:28028542

  13. A license to kill : The evolution of NK cell receptors

    NARCIS (Netherlands)

    Carrillo Bustamante, N.P.

    2015-01-01

    Natural killer (NK) cells innate immune cells that play a crucial role against viral infections and tumors. To be tolerant against healthy tissue and simultaneously attack infected cells, the activity of NK cells must be tightly regulated. Unlike B and T cells, NK cell do not undergo DNA rearrangeme

  14. Targeted killing of virally infected cells by radiolabeled antibodies to viral proteins.

    Directory of Open Access Journals (Sweden)

    Ekaterina Dadachova

    2006-11-01

    Full Text Available BACKGROUND: The HIV epidemic is a major threat to health in the developing and western worlds. A modality that targets and kills HIV-1-infected cells could have a major impact on the treatment of acute exposure and the elimination of persistent reservoirs of infected cells. The aim of this proof-of-principle study was to demonstrate the efficacy of a therapeutic strategy of targeting and eliminating HIV-1-infected cells with radiolabeled antibodies specific to viral proteins in vitro and in vivo. METHODS AND FINDINGS: Antibodies to HIV-1 envelope glycoproteins gp120 and gp41 labeled with radioisotopes bismuth 213 ((213Bi and rhenium 188 ((188Re selectively killed chronically HIV-1-infected human T cells and acutely HIV-1-infected human peripheral blood mononuclear cells (hPBMCs in vitro. Treatment of severe combined immunodeficiency (SCID mice harboring HIV-1-infected hPBMCs in their spleens with a (213Bi- or (188Re-labeled monoclonal antibody (mAb to gp41 resulted in a 57% injected dose per gram uptake of radiolabeled mAb in the infected spleens and in a greater than 99% elimination of HIV-1-infected cells in a dose-dependent manner. The number of HIV-1-infected thymocytes decreased 2.5-fold in the human thymic implant grafts of SCID mice treated with the (188Re-labeled antibody to gp41 compared with those treated with the (188Re-control mAb. The treatment did not cause acute hematologic toxicity in the treated mice. CONCLUSIONS: The current study demonstrates the effectiveness of HIV-targeted radioimmunotherapy and may provide a novel treatment option in combination with highly active antiretroviral therapy for the eradication of HIV.

  15. Specific killing effect of diphtheria toxin A fragment under control of DF3 promotor on human breast cancer cells%DF3调控下的白喉毒素A片段对人乳腺癌细胞的特异性杀伤作用

    Institute of Scientific and Technical Information of China (English)

    Ming Cai; Wenguang Huang; Wei Luo; Sheng Pan; Tao Yin

    2007-01-01

    Objective:To study the effects of recombinant expression vector containing human breast cancer DF3 promotor and diphtheria toxin A fragment on human breast cancer cells. Methods:Constructing recombinant expression vector PGL3-DF3-DTA and transfecting it into human breast cancer cells of DF3 positive and negative. By means of RT-PCR to measure the expression of DTA in human breast cancer cells. MTT color-imetry was used to examine the effect of PGL3-DF3-DTA on growth of human breast cancer calls. By experiment on nude mice to observe the killing effect of PGL3-DF3-DTA on human breast cancer cells. Results:Recombinant expression vector PGL3-DF3-DTA was highly expressed in human breast cancer cell line of DF3 positive, and it could kill the human breast cancer cells not only in vitro but also in vivo. Conclusion:Recombinant expression vector PGL3-DF3-DTA could produce specific killing effect on human breast cancer cell line of DF3 positive.

  16. A novel transferrin receptor-targeted hybrid peptide disintegrates cancer cell membrane to induce rapid killing of cancer cells

    Directory of Open Access Journals (Sweden)

    Kawamoto Megumi

    2011-08-01

    D cancer cells just in 10 min, to effectively kill these cells and induce approximately 80% apoptotic cell death but not in normal cells. The intravenous administration of TfR-lytic peptide in the athymic mice model significantly inhibited tumor progression. Conclusions TfR-lytic peptide might provide a potent and selective anticancer therapy for patients.

  17. Silver nanocrystals sensitize magnetic-nanoparticle-mediated thermo-induced killing of cancer cells

    Institute of Scientific and Technical Information of China (English)

    Lianke Liu; Fang Ni; Jianchao Zhang; Xiaoli Jiang; Xiang Lu; Zhirui Guo; Ruizhi Xu

    2011-01-01

    Magnetic nanoparticles (MNPs) can heat up tumor tissues and induce killing of cancer cells under external AC magnetic field. However, magnetic nanoparticles hyperthermia (MNPH) requires high concentration of MNPs that are injected into the tumor in order to obtain clinically needed thermal dose because of the complicated heat transfer in v/vo and the limited heat quality of MNPs. To cut down the dose of MNPs and enhance the effect of this Nanotherapy, we prepared silver nanoparticles (AgNPs) with different sizes and investigated the effects of these AgNPs on cancer cells in MNPH treatment. It was found that AgNPs could enhance thermo-sensitivity of glioma cells and this effect was size dependent. AgNPs could induce cell cycles arrested in G2/M phase and enhanced the apoptosis rate of cancer cells after hyperthermia. In glioma bearing rats model, MNPH combined with AgNPs could enhance Bax expression in cancer cells. Our results suggested that AgNPs could be a potential thermo-sensitizer and could be further developed for the design of Ag nanostructurebased thermal seeds for MNPH therapy.

  18. Synergistically killing activity of aspirin and histone deacetylase inhibitor valproic acid (VPA) on hepatocellular cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiaofei; Zhu, Yanshuang [Department of Infectious Diseases, Yiwu Central Hospita, 519 Nan men Street, Yiwu, Jinhua, Zhejing 322000 (China); He, Huabin [Department of Orthopedics, Yiwu Central Hospita, 519 Nan men Street, Yiwu, Jinhua, Zhejing 322000 (China); Lou, Lianqing; Ye, Weiwei; Chen, Yongxin [Department of Infectious Diseases, Yiwu Central Hospita, 519 Nan men Street, Yiwu, Jinhua, Zhejing 322000 (China); Wang, Jinghe, E-mail: Xiaofeili2000@163.com [Department of Infectious Diseases, Yiwu Central Hospita, 519 Nan men Street, Yiwu, Jinhua, Zhejing 322000 (China)

    2013-06-28

    Highlights: •Novel combination therapy using aspirin and valproic acid (VPA). •Combination of aspirin and VPA elicits synergistic cytotoxic effects. •Combination of aspirin and VPA significantly reduces the drug dosage required alone. •Combination of aspirin and VPA significantly inhibit tumor growth. •Lower dose of aspirin in combination therapy will minimize side effects of aspirin. -- Abstract: Aspirin and valproic acid (VPA) have been extensively studied for inducing various malignancies growth inhibition respectively, despite their severe side effects. Here, we developed a novel combination by aspirin and VPA on hepatocellular cancer cells (HCCs). The viability of HCC lines were analyzed by MTT assay, apoptotic analysis of HepG2 and SMMC-7721 cell was performed. Real time-PCR and Western blotting were performed to determine the expression of apoptosis related genes and proteins such as Survivin, Bcl-2/Bax, Cyclin D1 and p15. Moreover, orthotopic xenograft tumors were challenged in nude mice to establish murine model, and then therapeutic effect was analyzed after drug combination therapy. The viability of HCC lines’ significantly decreased after drug combination treatment, and cancer cell apoptosis in combination group increasingly induced compared with single drug use. Therapeutic effect was significantly enhanced by combination therapy in tumor volume and tumor weight decrease. From the data shown here, aspirin and VPA combination have a synergistic killing effect on hepatocellular cancers cells proliferation and apoptosis.

  19. Impact of prolonged fraction dose-delivery time modeling intensity-modulated radiation therapy on hepatocellular carcinoma cell killing

    Institute of Scientific and Technical Information of China (English)

    Xiao-Kang Zheng; Long-Hua Chen; Xiao Yan; Hong-Mei Wang

    2005-01-01

    AIM: To explore the impact of prolonged fraction dosedelivery time modeling intensity-modulated radiation therapy (IMRT) on cell killing of human hepatocellular carcinoma (HCC) HepG2 and Hep3B cell lines.METHODS: The radiobiological characteristics of human HCC HepG2 and Hep3b cell lines were studied with standard clonogenic assays, using standard linear-quadratic model and incomplete repair model to fit the dose-survival curves. The identical methods were also employed to investigate the biological effectiveness of irradiation protocols modeling clinical conventional fractionated external beam radiotherapy (EBRT, fraction delivery time 3 min) and IMRT with different prolonged fraction delivery time (15, 30, and 45 min). The differences of cell surviving fraction irradiated with different fraction delivery time were tested with paired t-test. Factors determining the impact of prolonged fraction delivery time on cell killing were analyzed.RESULTS: The α/β and repair half-time (T1/2) of HepG2and Hep3b were 3.1 and 7.4 Gy, and 22 and 19 min respectively. The surviving fraction of HepG2 irradiated modeling IMRT with different fraction delivery time was significantly higher than irradiated modeling EBRT and the cell survival increased more pronouncedly with the fraction delivery time prolonged from 15 to 45 min,while no significant differences of cell survival in Hep3b were found between different fraction delivery time protocols.CONCLUSION: The prolonged fraction delivery time modeling IMRT significantly decreased the cell killing in HepG2 but not in Hep3b. The capability of sub-lethal damage repair was the predominant factor determining the cell killing decrease. These effects, if confirmed by clinical studies, should be considered in designing IMRT treatments for HCC.

  20. The kinematics of cytotoxic lymphocytes influence their ability to kill target cells.

    Science.gov (United States)

    Bhat, Purnima; Leggatt, Graham; Matthaei, Klaus I; Frazer, Ian H

    2014-01-01

    Cytotoxic lymphocytes (CTL) have been reported to show a range of motility patterns from rapid long-range tracking to complete arrest, but how and whether these kinematics affect their ability to kill target cells is not known. Many in vitro killing assays utilize cell lines and tumour-derived cells as targets, which may be of limited relevance to the kinetics of CTL-mediated killing of somatic cells. Here, live-cell microscopy is used to examine the interactions of CTL and primary murine skin cells presenting antigens. We developed a qualitative and quantitative killing assay using extended-duration fluorescence time-lapse microscopy coupled with large-volume objective software-based data analysis to obtain population data of cell-to-cell interactions, motility and apoptosis. In vivo and ex vivo activated antigen-specific cytotoxic lymphocytes were added to primary keratinocyte targets in culture with fluorometric detection of caspase-3 activation in targets as an objective determinant of apoptosis. We found that activated CTL achieved contact-dependent apoptosis of non-tumour targets after a period of prolonged attachment - on average 21 hours - which was determined by target cell type, amount of antigen, and activation status of CTL. Activation of CTL even without engagement of the T cell receptor was sufficient to mobilise cells significantly above baseline, while the addition of cognate antigen further enhanced their motility. Highly activated CTL showed markedly increased vector displacement, and velocity, and lead to increased antigen-specific target cell death. These data show that the inherent kinematics of CTL correlate directly with their ability to kill non-tumour cells presenting cognate antigen.

  1. The kinematics of cytotoxic lymphocytes influence their ability to kill target cells.

    Directory of Open Access Journals (Sweden)

    Purnima Bhat

    Full Text Available Cytotoxic lymphocytes (CTL have been reported to show a range of motility patterns from rapid long-range tracking to complete arrest, but how and whether these kinematics affect their ability to kill target cells is not known. Many in vitro killing assays utilize cell lines and tumour-derived cells as targets, which may be of limited relevance to the kinetics of CTL-mediated killing of somatic cells. Here, live-cell microscopy is used to examine the interactions of CTL and primary murine skin cells presenting antigens. We developed a qualitative and quantitative killing assay using extended-duration fluorescence time-lapse microscopy coupled with large-volume objective software-based data analysis to obtain population data of cell-to-cell interactions, motility and apoptosis. In vivo and ex vivo activated antigen-specific cytotoxic lymphocytes were added to primary keratinocyte targets in culture with fluorometric detection of caspase-3 activation in targets as an objective determinant of apoptosis. We found that activated CTL achieved contact-dependent apoptosis of non-tumour targets after a period of prolonged attachment - on average 21 hours - which was determined by target cell type, amount of antigen, and activation status of CTL. Activation of CTL even without engagement of the T cell receptor was sufficient to mobilise cells significantly above baseline, while the addition of cognate antigen further enhanced their motility. Highly activated CTL showed markedly increased vector displacement, and velocity, and lead to increased antigen-specific target cell death. These data show that the inherent kinematics of CTL correlate directly with their ability to kill non-tumour cells presenting cognate antigen.

  2. Study of cell killing and morphology on S180 by ultrasound activating hematoporphyrin derivatives

    Institute of Scientific and Technical Information of China (English)

    刘全宏; 孙世惠; 肖娅萍; 齐浩; 尚志远; 张军平; 张金选; 任耀辉; 李萌; 李青

    2003-01-01

    The inhibition of ascitic S180 and induced sarcoma 180 in vivo was studied with the combination of hematoporphyrin derivatives (HpD) and ultrasound (US) at the frequency of 1.1 MHz and different intensities by light microscopy observation, electronic microscopy observation, cytochemical analysis and fluorescence labeling. The present study indicated that the injury of ascitic S180 increased as time passed and the inhibitory effect was stronger in US plus HpD group than that in other groups. Our results also indicated that the changes in cell structure, cytochrome C oxidase activity, the degradation and missing of DNA were the important factors that inhibited the tumor cell growth and even induced celldeath. The phenomenon of apoptosis of tumor cells indicated that cell death andinduced apoptosis exist in the treatment of sonodynamic therapy (SDT). Our study investigated the mechanism underlying the killing effect of S180 induced by USactivating HpD by the observation of cell morphology and dynamic changes from seminal injury to succeeded injury even to death. It would provide rich referencefor the study of SDT.

  3. Both necrosis and apoptosis contribute to HIV-1-induced killing of CD4 cells

    Science.gov (United States)

    Plymale, D. R.; Tang, D. S.; Comardelle, A. M.; Fermin, C. D.; Lewis, D. E.; Garry, R. F.

    1999-01-01

    BACKGROUND: Data currently available on HIV-1-induced cytopathology is unclear regarding the mechanism of cell killing. OBJECTIVE: To clarify the extent to which apoptosis or necrosis is involved in HIV-1-induced cell death in view of conflicting existing data. METHODS: T lymphoblastoid cells or peripheral blood mononuclear cells were infected by various strains of HIV-1 and the numbers of apoptotic or necrotic cells were quantified at various times after infection using video-image analysis techniques; the results were compared with the amount of fragmented DNA using a quantitative method. Measurement of mitochondrial transmembrane potential (deltapsi(m)) and intracellular calcium concentrations [Ca2+]i was performed with fluorescent probes and fluorescence concentration analysis (FCA). RESULTS: Although lymphoblastoid and monocytoid cells acutely infected by HIV-1 had increased levels of fragmented DNA, a marker of apoptotic cell death, few (killing of CD4 cells.

  4. Relative efficiencies of three ultraviolet radiation wavelengths for cell killing and transformation in mouse cells in vitro.

    Science.gov (United States)

    Papadopoulo, D; Muel, B; Latarjet, R

    1983-09-01

    C3H 10 T 1/2 clone 8 mouse cells were irradiated in vitro with three U.V. wavelengths 280, 254, and 230 nm. Two effects were investigated, survival and malignant transformation, and the relative efficiencies were determined for the three radiations. For transformation, these efficiencies were: 280nm:3.9; 254nm:5.1; 230nm:2.3 (transformations produced by 5 Jm-2 of U.V. for 1000 surviving cells). For cell killing the efficiencies were, in relative units, 34, 100, and 50 respectively. These efficiencies are in agreement with the hypothesis that the main chromophore for both effects is the nucleic acid, and not the protein moiety of the genome. This conclusion agrees with that previously reached by other investigators, but our present results obtained with the short wave-length 230 nm provide an especially strong new argument.

  5. Relative efficiencies of three ultraviolet radiation wavelengths for cell killing and transformation in mouse cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Papadopoulo, D.; Muel, B.; Latarjet, R. (Institut du Radium, 75 - Paris (France). Lab. Curie)

    1983-09-01

    C3H 10 T 1/2 clone 8 mouse cells were irradiated in vitro with three U.V. wavelengths 280, 254, and 230 nm. Two effects were investigated, survival and malignant transformation, and the relative efficiences were determined for the three radiation. For transformation, these efficiences were: 280 nm: 3.9; 254 nm: 5.1; 230 nm: 2.3 (transformations produced by 5 J m/sup -2/ of U.V. for 1000 surviving cells). For cell killing the efficiencies were, in relative units, 34, 100, and 50 respectively. These efficiencies are in agreement with the hypothesis that the main chromophore for both effects is the nucleic acid, and not the protein moiety of the genome. This conclusion agrees with that previously reached by other investigators, but our present results obtained with the short wave-length 230 nm provide an especially strong new argument.

  6. 突变型胸苷激酶对鼠C6胶质瘤细胞的杀伤效应%Killing effect of mutant herpes simplex virus type 1 thymidine kinase on rat C6 glioma cells

    Institute of Scientific and Technical Information of China (English)

    徐晓燕; 刘伟国; 杨小锋; 郑学胜; 沈罡

    2009-01-01

    Objective To explore the killing effect of mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) and its wild type (HSV1-tk) on rat C6 glioma cells after Ganciclovir (GCV) treatment. Methods Eukaryotic expressing vector pcDNA3.1 containing HSV1-tk or HSV1-sr39tk was transfected into C6 cells and identified with RT-PCR. GCV killing efficiencies after gene transfer of HSV1-tk or HSV1-sr39tk were observed and compared in vitro and in vivo. Results 308bp DNA fragment was amplified through RT-PCR in both cells transfected with HSV1-tk (C6/tk) and those with HSV1-sr39tk (C6/sr39tk). As the prodrug GCV concentrations increasing from 0 μmol/L to 400 μmol/L, the cell survival rates in C6/sr39tk group declined from (99.96 ± 3.54) % to (4. 75 ± 1.79) %, while in C6/tk group from (100.03±2. 95)% to (59. 16±3.48)% and in C6 group from(100. 29±1.20)% to (83.62 ±7.56)%. There was a significant difference in killing effect among C6、C6/tk and C6/sr39tk group after GCV treatment(P < 0. 05). In vivo experiment, the rate of tumor formation was 100%. After 10 day treatment of GCV following tumor formation, tumors in animals implantated with C6/sr39tk、C6/tk or C6 got an average volume equaling (574. 08±107.72) mm3、(928.47±165.61) mm3 and (1287.24±364. 84) mm3 respectively. The most powerful tumor growth inhibition was observed in C6/sr39tk + GCV group, with an inhibition rate of tumor growth about 55.40%. Conclusion C6 cells transfected with HSV1-sr39tk were more sensitive to GCV than those with HSV1-tk both in vitro and in vivo, and thus HSV1-sr39tk can be considered as a mean to improve the overall efficacy of the HSV1-tk/GCV suicide gene system.%目的 观察比较突变型单纯疱疹胸苷激酶(HSV1-sr39tk)/更昔洛韦(GCV)和野生型HSV1-tk/GCV对鼠C6胶质瘤细胞的杀伤效应.方法 利用真核表达载体转染目的 基因到C6细胞,RT-PCR鉴定.通过MTT实验和活体植瘤,比较各组对GCV敏感性.结果

  7. Doxorubicin loaded 17β-estradiol based SWNT dispersions for target specific killing of cancer cells.

    Science.gov (United States)

    Ghosh, Moumita; Das, Prasanta Kumar

    2016-06-01

    The present work reports the synthesis of a 17β-estradiol based amphiphiles comprising of polyethylene glycol (PEG) moiety linked through succinic acid that non-covalently dispersed (76%) the single walled carbon nanotubes (SWNTs) in water. The superior exfoliation of carbon nanotubes was characterized by microscopic and spectroscopic studies. Significant stability of these SWNT dispersions was observed in the presence of protein in cell culture media and the nanohybrids were highly biocompatible toward mammalian cells. Anticancer drug doxorubicin loaded on these nanohybrids was selectively delivered within estrogen receptor rich cancer cells, MCF7 (breast cancer cell) and A549 (lung cancer cell). Microscopic studies showed the localization of doxorubicin within the cancer cell nucleus whereas no such localization was observed in ER negative cells. Both these ER positive cancer cells were killed by ∼3 fold higher efficiency than that of ER negative MDA-MB-231 (advanced breast cancer cell) and HeLa cells that are deprived of estrogen receptors. Thus, judiciously designed estradiol based nanohybrids proved to be excellent tool for SWNT dispersion and also for selectively killing of ER positive cancer cells. To the best of our knowledge, for the first time non-covalently modified SWNTs by estradiol based amphiphilic dispersing agent have been used for selective killing of ER positive cancer cells by doxorubicin loaded on dispersed SWNTs. It holds immense promise to be exploited as a cancer therapeutic agent.

  8. Cytomegalovirus-Infected Cells Resist T Cell Mediated Killing in an HLA-Recognition Independent Manner.

    Science.gov (United States)

    Proff, Julia; Walterskirchen, Christian; Brey, Charlotte; Geyeregger, Rene; Full, Florian; Ensser, Armin; Lehner, Manfred; Holter, Wolfgang

    2016-01-01

    In order to explore the potential of HLA-independent T cell therapy for human cytomegalovirus (HCMV) infections, we developed a chimeric antigen receptor (CAR) directed against the HCMV encoded glycoprotein B (gB), which is expressed at high levels on the surface of infected cells. T cells engineered with this anti-gB CAR recognized HCMV-infected cells and released cytokines and cytotoxic granules. Unexpectedly, and in contrast to analogous approaches for HIV, Hepatitis B or Hepatitis C virus, we found that HCMV-infected cells were resistant to killing by the CAR-modified T cells. In order to elucidate whether this phenomenon was restricted to the use of CARs, we extended our experiments to T cell receptor (TCR)-mediated recognition of infected cells. To this end we infected fibroblasts with HCMV-strains deficient in viral inhibitors of antigenic peptide presentation and targeted these HLA-class I expressing peptide-loaded infected cells with peptide-specific cytotoxic T cells (CTLs). Despite strong degranulation and cytokine production by the T cells, we again found significant inhibition of lysis of HCMV-infected cells. Impairment of cell lysis became detectable 1 day after HCMV infection and gradually increased during the following 3 days. We thus postulate that viral anti-apoptotic factors, known to inhibit suicide of infected host cells, have evolved additional functions to directly abrogate T cell cytotoxicity. In line with this hypothesis, CAR-T cell cytotoxicity was strongly inhibited in non-infected fibroblasts by expression of the HCMV-protein UL37x1, and even more so by additional expression of UL36. Our data extend the current knowledge on Betaherpesviral evasion from T cell immunity and show for the first time that, beyond impaired antigen presentation, infected cells are efficiently protected by direct blockade of cytotoxic effector functions through viral proteins.

  9. Endocytosis of Cytotoxic Granules Is Essential for Multiple Killing of Target Cells by T Lymphocytes.

    Science.gov (United States)

    Chang, Hsin-Fang; Bzeih, Hawraa; Schirra, Claudia; Chitirala, Praneeth; Halimani, Mahantappa; Cordat, Emmanuelle; Krause, Elmar; Rettig, Jens; Pattu, Varsha

    2016-09-15

    CTLs are serial killers that kill multiple target cells via exocytosis of cytotoxic granules (CGs). CG exocytosis is tightly regulated and has been investigated in great detail; however, whether CG proteins are endocytosed following exocytosis and contribute to serial killing remains unknown. By using primary CTLs derived from a knock-in mouse of the CG membrane protein Synaptobrevin2, we show that CGs are endocytosed in a clathrin- and dynamin-dependent manner. Following acidification, endocytosed CGs are recycled through early and late, but not recycling endosomes. CGs are refilled with granzyme B at the late endosome stage and polarize to subsequent synapses formed between the CTL and new target cells. Importantly, inhibiting CG endocytosis in CTLs results in a significant reduction of their cytotoxic activity. Thus, our data demonstrate that continuous endocytosis of CG membrane proteins is a prerequisite for efficient serial killing of CTLs and identify key events in this process.

  10. Activated allogeneic NK cells preferentially kill poor prognosis B-cell chronic lymphocytic leukemia cells

    Directory of Open Access Journals (Sweden)

    Diego Sanchez-Martinez

    2016-10-01

    Full Text Available Mutational status of TP53 together with expression of wild type (wt IGHV represents the most widely accepted biomarkers, establishing a very poor prognosis in B-cell chronic lymphocytic leukemia (B-CLL patients. Adoptive cell therapy using allogeneic HLA mismatched Natural Killer (NK cells has emerged as an effective and safe alternative in the treatment of acute myeloid and lymphoid leukemias that do not respond to traditional therapies. We have described that allogeneic activated NK cells eliminate hematological cancer cell lines with multidrug resistance acquired by mutations in the apoptotic machinery. This effect depends on the activation protocol, being B-lymphoblastoid cell lines (LCLs the most effective stimulus to activate NK cells. Here we have further analyzed the molecular determinants involved in allogeneic NK cell recognition and elimination of B-CLL cells, including the expression of ligands of the main NK cell activating receptors (NKG2D and NCRs and HLA mismatch. We present preliminary data suggesting that B-CLL susceptibility significantly correlates with HLA mismatch between NK cell donor and B-CLL patient. Moreover, we show that the sensitivity of B-CLL cells to NK cells depends on the prognosis based on TP53 and IGHV mutational status. Cells from patients with worse prognosis (mutated TP53 and wt IGHV are the most susceptible to activated NK cells. Hence, B-CLL prognosis may predict the efficacy of allogenic activated NK cells and, thus, NK cell transfer represents a good alternative to treat poor prognosis B-CLL patients who present a very short life expectancy due to lack of effective treatments.□

  11. Killing Effect of Lentivirus-mediated Double Suicide Genes on Human Breast Cancer Cells in vivo%慢病毒介导双自杀基因对乳腺癌细胞的体内杀伤作用

    Institute of Scientific and Technical Information of China (English)

    孔恒; 黄宗海; 陈海金; 陶霖玉; 齐柯

    2011-01-01

    Objective: To evaluate the killing effect of lentivirus mediated CD/TK fusion gene controlled by kinase insert domain-containing receptor (KDR) promoter on human breast cancer cells in vivo.Methods: Nude mice were used as hosts for cell line MCF-7 xenografts to establish the animal model of breast cancer.The tumor-bearing mice were randomly divided into 4 groups.Group Ⅰ were blank controls with tumor-bearing mice with no treatment.Group Ⅱ was injected with lentivirus and prodrug ( 5-FC+ GCV ).Group Ⅲ was injected with lentivirus.Group Ⅳ was injected with prodrug.Tumor growth rate, tumor size and weight were observed.The expression of double suicide genes in xenograft tumors was examined by RT-PCR.The tumor tissue was analyzed by hematoxylin-eosin staining and PCNA immunohistochemistry.Flow cytometry (FCM) was used for cell cycle analysis.Results: Tumor growth was obviously inhibited in group Ⅱ.But there was no significant difference in tumor growth among group Ⅰ, group Ⅲ, and group Ⅳ.RT-PCR demonstrated the existence of CD/TK gene in genetically modified xenograft tumor cells.The expression of PCNA was lower in tumor tissue of group Ⅱ than in group Ⅰ.Flow cytometry analysis showed that the proportion of G1 phase cells was increased and the proportion of G2-M phase cells was decreased.Conclusion: Combined with the prodrug, FGW-KDRP-CD/TK can significantly inhibit the growth of implanted breast cancer cells in nude mice and cell cycle arrest, which may be related to its inhibitory effect on PCNA expression.%目的:研究慢病毒介导的KDR启动子驱动的胞嘧啶脱氨酶(CD)/胸苷激酶(TK)融合基因系统(FGW-KDRP-CD/TK)对乳腺癌细胞的体内杀伤作用.方法:培养乳腺癌MCF-7细胞,建立裸鼠荷瘤模型.荷瘤后将裸鼠随机分为4组.Ⅰ组:空白对照,荷瘤但不施加任何处理;Ⅱ组:注射慢病毒与前药(5-FC+GCV);Ⅲ组:仅注射慢病毒;Ⅳ组:仅注射前药.观察肿瘤生长速度,测量瘤体大小及

  12. Activated allogeneic NK cells preferentially kill poor prognosis B-cell chronic lymphocytic leukemia cells

    OpenAIRE

    2016-01-01

    Mutational status of TP53 together with expression of wild type (wt) IGHV represents the most widely accepted biomarkers, establishing a very poor prognosis in B-cell chronic lymphocytic leukemia (B-CLL) patients. Adoptive cell therapy using allogeneic HLA mismatched Natural Killer (NK) cells has emerged as an effective and safe alternative in the treatment of acute myeloid and lymphoid leukemias that do not respond to traditional therapies. We have described that allogeneic activated NK cell...

  13. Structural factors and mechanisms underlying the improved photodynamic cell killing with silicon phthalocyanine photosensitizers directed to lysosomes versus mitochondria.

    Science.gov (United States)

    Rodriguez, Myriam E; Zhang, Ping; Azizuddin, Kashif; Delos Santos, Grace B; Chiu, Song-mao; Xue, Liang-yan; Berlin, Jeffery C; Peng, Xinzhan; Wu, Hongqiao; Lam, Minh; Nieminen, Anna-Liisa; Kenney, Malcolm E; Oleinick, Nancy L

    2009-01-01

    The phthalocyanine photosensitizer Pc 4 has been shown to bind preferentially to mitochondrial and endoplasmic reticulum membranes. Upon photoirradiation of Pc 4-loaded cells, membrane components, especially Bcl-2, are photodamaged and apoptosis, as indicated by activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase, is triggered. A series of analogs of Pc 4 were synthesized, and the results demonstrate that Pcs with the aminopropylsiloxy ligand of Pc 4 or a similar one on one side of the Pc ring and a second large axial ligand on the other side of the ring have unexpected properties, including enhanced cell uptake, greater monomerization resulting in greater intracellular fluorescence and three-fold higher affinity constants for liposomes. The hydroxyl-bearing axial ligands tend to reduce aggregation of the Pc and direct it to lysosomes, resulting in four to six times more killing of cells, as defined by loss of clonogenicity, than with Pc 4. Whereas Pc 4-PDT photodamages Bcl-2 and Bcl-xL, Pc 181-PDT causes much less photodamage to Bcl-2 over the same dose-response range relative to cell killing, with earlier cleavage of Bid and slower caspase-3-dependent apoptosis. Therefore, within this series of photosensitizers, these hydroxyl-bearing axial ligands are less aggregated than is Pc 4, tend to localize to lysosomes and are more effective in overall cell killing than is Pc 4, but induce apoptosis more slowly and by a modified pathway.

  14. LuIII parvovirus selectively and efficiently targets, replicates in, and kills human glioma cells.

    Science.gov (United States)

    Paglino, Justin C; Ozduman, Koray; van den Pol, Anthony N

    2012-07-01

    Because productive infection by parvoviruses requires cell division and is enhanced by oncogenic transformation, some parvoviruses may have potential utility in killing cancer cells. To identify the parvovirus(es) with the optimal oncolytic effect against human glioblastomas, we screened 12 parvoviruses at a high multiplicity of infection (MOI). MVMi, MVMc, MVM-G17, tumor virus X (TVX), canine parvovirus (CPV), porcine parvovirus (PPV), rat parvovirus 1A (RPV1A), and H-3 were relatively ineffective. The four viruses with the greatest oncolytic activity, LuIII, H-1, MVMp, and MVM-G52, were tested for the ability, at a low MOI, to progressively infect the culture over time, causing cell death at a rate higher than that of cell proliferation. LuIII alone was effective in all five human glioblastomas tested. H-1 progressively infected only two of five; MVMp and MVM-G52 were ineffective in all five. To investigate the underlying mechanism of LuIII's phenotype, we used recombinant parvoviruses with the LuIII capsid replacing the MVMp capsid or with molecular alteration of the P4 promoter. The LuIII capsid enhanced efficient replication and oncolysis in MO59J gliomas cells; other gliomas tested required the entire LuIII genome to exhibit enhanced infection. LuIII selectively infected glioma cells over normal glial cells in vitro. In mouse models, human glioblastoma xenografts were selectively infected by LuIII when administered intratumorally; LuIII reduced tumor growth by 75%. LuIII also had the capacity to selectively infect subcutaneous or intracranial gliomas after intravenous inoculation. Intravenous or intracranial LuIII caused no adverse effects. Intracranial LuIII caused no infection of mature mouse neurons or glia in vivo but showed a modest infection of developing neurons.

  15. Residual chromatin breaks as biodosimetry for cell killing by carbon ions

    Science.gov (United States)

    Suzuki, M.; Kase, Y.; Nakano, T.; Kanai, T.; Ando, K.

    1998-11-01

    We have studied the relationship between cell killing and the induction of residual chromatin breaks on various human cell lines and primary cultured cells obtained by biopsy from patients irradiated with either X-rays or heavy-ion beams to identify potential bio-marker of radiosensitivity for radiation-induced cell killing. The carbon-ion beams were accelerated with the Heavy Ion Medical Accelerator in Chiba (HIMAC). Six primary cultures obtained by biopsy from 6 patients with carcinoma of the cervix were irradiated with two different mono-LET beams (LET = 13 keV/μm, 76 keV/μm) and 200kV X rays. Residual chromatin breaks were measured by counting the number of non-rejoining chromatin fragments detected by the premature chromosome condensation (PCC) technique after a 24 hour post-irradiation incubation period. The induction rate of residual chromatin breaks per cell per Gy was the highest for 76 keV/μm beams on all of the cells. Our results indicated that cell which was more sensitive to the cell killing was similarly more susceptible to induction of residual chromatin breaks. Furthermore there is a good correlation between these two end points in various cell lines and primary cultured cells. This suggests that the detection of residual chromatin breaks by the PCC technique may be useful as a predictive assay of tumor response to cancer radiotherapy.

  16. Preferential killing of cancer cells using silicon carbide quantum dots.

    Science.gov (United States)

    Mognetti, Barbara; Barberis, Alessandro; Marino, Silvia; Di Carlo, Francesco; Lysenko, Vladimir; Marty, Olivier; Géloën, Alain

    2010-12-01

    Silicon carbide quantum dots are highly luminescent biocompatible nanoparticles whose properties might be of particular interest for biomedical applications. In this study we investigated Silicon Carbide Quantum Dots (3C-SiC QDs) cellular localisation and influence on viability and proliferation on oral squamous carcinoma (AT-84 and HSC) and immortalized cell lines (S-G). They clearly localize into the nuclei, but the presence of 3C-SiC QDs in culture medium provoke morphological changes in cultured cells. We demonstrate that 3C-SiC QDs display dose- and time-dependent selective cytotoxicity on cancer versus immortalized cells in vitro. Since one of the limitations of classical antineoplastic drugs is their lack of selectivity, these results open a new way in the search for antiproliferative drugs.

  17. Effects of Vegetation Structure on the Location of Lion Kill Sites in African Thicket.

    Directory of Open Access Journals (Sweden)

    Andrew B Davies

    Full Text Available Predator-prey relationships are integral to ecosystem stability and functioning. These relationships are, however, difficult to maintain in protected areas where large predators are increasingly being reintroduced and confined. Where predators make kills has a profound influence on their role in ecosystems, but the relative importance of environmental variables in determining kill sites, and how these might vary across ecosystems is poorly known. We investigated kill sites for lions in South Africa's thicket biome, testing the importance of vegetation structure for kill site locations compared to other environmental variables. Kill sites were located over four years using GPS telemetry and compared to non-kill sites that had been occupied by lions, as well as to random sites within lion ranges. Measurements of 3D vegetation structure obtained from Light Detection and Ranging (LiDAR were used to calculate the visible area (viewshed around each site and, along with wind and moonlight data, used to compare kill sites between lion sexes, prey species and prey sexes. Viewshed area was the most important predictor of kill sites (sites in dense vegetation were twice as likely to be kill sites compared to open areas, followed by wind speed and, less so, moonlight. Kill sites for different prey species varied with vegetation structure, and male prey were killed when wind speeds were higher compared to female prey of the same species. Our results demonstrate that vegetation structure is an important component of predator-prey interactions, with varying effects across ecosystems. Such differences require consideration in terms of the ecological roles performed by predators, and in predator and prey conservation.

  18. Irradiation Can Selectively Kill Tumor Cells while Preserving Erythrocyte Viability in a Co-Culture System.

    Directory of Open Access Journals (Sweden)

    Ming Gong

    Full Text Available An understanding of how to safely apply intraoperative blood salvage (IBS in cancer surgery has not yet been obtained. Here, we investigated the optimal dose of 137Cs gamma-ray irradiation for killing human hepatocarcinoma (HepG2, gastrocarcinoma (SGC7901, and colonic carcinoma (SW620 tumor cells while preserving co-cultured erythrocytes obtained from 14 healthy adult volunteers. HepG2, SGC7901, or SW620 cells were mixed into the aliquots of erythrocytes. After the mixed cells were treated with 137Cs gamma-ray irradiation (30, 50, and 100 Gy, tumor cells and erythrocytes were separated by density gradient centrifugation in Percoll with a density of 1.063 g/ml. The viability, clonogenicity, DNA synthesis, tumorigenicity, and apoptosis of the tumor cells were determined by MTT assay, plate colony formation, 5-ethynyl-2'-deoxyuridine (EdU incorporation, subcutaneous xenograft implantation into immunocompromised mice, and annexin V/7-AAD staining, respectively. The ATP concentration, 2,3-DPG level, free Hb concentration, osmotic fragility, membrane phosphatidylserine externalization, blood gas variables, reactive oxygen species levels, and superoxide dismutase levels in erythrocytes were analyzed. We found that 137Cs gamma-ray irradiation at 50 Gy effectively inhibited the viability, proliferation, and tumorigenicity of HepG2, SGC7901, and SW620 cells without markedly damaging the oxygen-carrying ability or membrane integrity or increasing the oxidative stress of erythrocytes in vitro. These results demonstrated that 50 Gy irradiation in a standard 137Cs blood irradiator might be a safe and effective method of inactivating HepG2, SGC7901, and SW620 cells mixed with erythrocytes, which might help to safely allow IBS in cancer surgery.

  19. A novel bispecific antibody, S-Fab, induces potent cancer cell killing.

    Science.gov (United States)

    Li, Li; He, Ping; Zhou, Changhua; Jing, Li; Dong, Bin; Chen, Siqi; Zhang, Ning; Liu, Yawei; Miao, Ji; Wang, Zhong; Li, Qing

    2015-01-01

    Bispecific antibodies that engage immune cells to kill cancer cells have been actively studied in cancer immunotherapy. In this study, we present a novel bispecific format, S-Fab, fabricated by linking a single-domain anti-carcinoembryonic antigen VHH to a conventional anti-CD3 Fab. In contrast to most bispecific antibodies, the S-Fab bispecific antibody can be efficiently expressed and purified from bacteria. The purified S-Fab is stable in serum and is able to recruit T cells to drive potent cancer cell killing. In xenograft models, the S-Fab antibody suppresses tumor growth in the presence of human immune cells. Our study suggested that the bispecific S-Fab format can be applied to a wide range of immunotherapies.

  20. Ag nanoparticles sensitize IR-induced killing of cancer cells

    Institute of Scientific and Technical Information of China (English)

    Ruizhi Xu; Jun Ma; Xinchen Sun; Zhongping Chen; Xiaoli Jiang; Zhirui Guo; Lan Huang; Yang Li; Meng Wang; Changling Wang; Jiwei Liu; Xu Fan; Jiayu Gu; Xi Chen; Yu Zhang; Ning Gu

    2009-01-01

    @@ Dear Editor, Nanosized particulate systems combining better can-cer diagnosis with therapeutic effect are being designed based on the merging of nanotechnology with cellular and molecular techniques.

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

    Science.gov (United States)

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

    2016-01-01

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

  2. Vesicle-associated membrane protein 7 (VAMP-7) is essential for target cell killing in a natural killer cell line.

    Science.gov (United States)

    Marcet-Palacios, Marcelo; Odemuyiwa, Solomon O; Coughlin, Jason J; Garofoli, Daniella; Ewen, Catherine; Davidson, Courtney E; Ghaffari, Mazyar; Kane, Kevin P; Lacy, Paige; Logan, Michael R; Befus, A Dean; Bleackley, R Chris; Moqbel, Redwan

    2008-02-15

    Natural killer cells recognize and induce apoptosis in foreign, transformed or virus-infected cells through the release of perforin and granzymes from secretory lysosomes. Clinically, NK-cell mediated killing is a major limitation to successful allo- and xenotransplantation. The molecular mechanisms that regulate the fusion of granzyme B-containing secretory lysosomes to the plasma membrane in activated NK cells, prior to target cell killing, are not fully understood. Using the NK cell line YT-Indy as a model, we have investigated the expression of SNAP REceptors (SNAREs), both target (t-) and vesicular (v-) SNAREs, and their function in granzyme B-mediated target cell killing. Our data showed that YT-Indy cells express VAMP-7 and SNAP-23, but not VAMP-2. VAMP-7 was associated with granzyme B-containing lysosomal granules. Using VAMP-7 small interfering RNA (siRNA), we successfully knocked down the expression of VAMP-7 protein in YT-Indy to less than 10% of untreated cells in 24h. VAMP7-deficient YT-Indy cells activated via co-culture with Jurkat cells released <1ng/mL of granzyme B, compared to 1.5-2.5 microg/mL from controls. Using Jurkat cells as targets, we showed a 7-fold reduction in NK cell-mediated killing by VAMP-7 deficient YT-Indy cells. Our results show that VAMP-7 is a crucial component of granzyme B release and target cell killing in the NK cell line YT-Indy. Thus, targeting VAMP-7 expression specifically with siRNA, following transplantation, may be a viable strategy for preventing NK cell-mediated transplant rejection, in vivo.

  3. Managing Threat, Cost, and Incentive to Kill: The Short- and Long-Term Effects of Intervention in Mass Killings

    Science.gov (United States)

    Kathman, Jacob D.; Wood, Reed M.

    2011-01-01

    How do third-party interventions affect the severity of mass killings? The authors theorize that episodes of mass killing are the consequence of two factors: (1) the threat perceptions of the perpetrators and (2) the cost of implementing genocidal policies relative to other alternatives. To reduce genocidal hostilities, interveners must address…

  4. Cancer suicide gene therapy with TK.007: superior killing efficiency and bystander effect.

    Science.gov (United States)

    Preuss, Ellen; Muik, Alexander; Weber, Kristoffer; Otte, Jürgen; von Laer, Dorothee; Fehse, Boris

    2011-11-01

    Suicide gene therapy is a promising concept in oncology. We have recently introduced a novel suicide gene, TK.007, which was shown to excel established herpes simplex virus thymidine kinase (HSVtk) variants when used for donor-lymphocyte modification in adoptive immunotherapy models. Here, the potential of TK.007 in killing cancer cells was studied. Initially, we transduced tumour cell lines derived from different neoplasias (glioblastoma, melanoma, lung cancer, colon cancer) with lentiviral LeGO vectors encoding TK.007 or the splice-corrected (sc)HSVtk together with an eGFP/Neo-marker. Based on direct in vitro comparison, we found that TK.007 facilitates more efficient tumour cell killing at significantly lower ganciclovir doses in all tumour cell lines tested. Also, using different readout systems, we found a significantly stronger bystander effect of TK.007 as compared to scHSVtk. Importantly, in vitro data were confirmed in vivo using a subcutaneous G62 glioblastoma model in NOD/SCID mice. In mice transplanted with scHSVtk-positive tumours, treatment with low (10 mg/kg) or standard (50 mg/kg) ganciclovir doses resulted only in short-term growth inhibition or transient tumour remission, respectively. In striking contrast, in the TK.007 group, all animals achieved continuous complete remission after both standard and low-dose ganciclovir. Finally, a substantial bystander effect for TK.007 was also confirmed with the G62 model in vivo, where significantly prolonged survival for mice bearing tumours containing only 10% or 50% TK.007-expressing cells was observed. In summary, our data indicate strongly improved anti-tumour activity of TK.007 as compared to conventional HSVtk. We therefore suppose that TK.007 is an excellent candidate for cancer suicide gene therapy.

  5. Antimicrobial design of titanium surface that kill sessile bacteria but support stem cells adhesion

    Science.gov (United States)

    Zhu, Chen; Bao, Ni-Rong; Chen, Shuo; Zhao, Jian-Ning

    2016-12-01

    Implant-related bacterial infection is one of the most severe postoperative complications in orthopedic or dental surgery. In this context, from the perspective of surface modification, increasing efforts have been made to enhance the antibacterial capability of titanium surface. In this work, a hierarchical hybrid surface architecture was firstly constructed on titanium surface by two-step strategy of acid etching and H2O2 aging. Then silver nanoparticles were firmly immobilized on the hierarchical surface by ion implantation, showing no detectable release of silver ions from surface. The designed titanium surface showed good bioactivity. More importantly, this elaborately designed titanium surface can effectively inactivate the adherent S. aureus on surface by virtue of a contact-killing mode. Meanwhile, the designed titanium surface can significantly facilitate the initial adhesion and spreading behaviors of bone marrow mesenchymal stem cells (MSCs) on titanium. The results suggested that, the elaborately designed titanium surface might own a cell-favoring ability that can help mammalian cells win the initial adhesion race against bacteria. We hope the present study can provide a new insight for the better understanding and designing of antimicrobial titanium surface, and pave the way to satisfying clinical requirements.

  6. Killing multiple myeloma cells with the small molecule 3-bromopyruvate: implications for therapy.

    Science.gov (United States)

    Majkowska-Skrobek, Grażyna; Augustyniak, Daria; Lis, Paweł; Bartkowiak, Anna; Gonchar, Mykhailo; Ko, Young H; Pedersen, Peter L; Goffeau, Andre; Ułaszewski, Stanisław

    2014-07-01

    The small molecule 3-bromopyruvate (3-BP), which has emerged recently as the first member of a new class of potent anticancer agents, was tested for its capacity to kill multiple myeloma (MM) cancer cells. Human MM cells (RPMI 8226) begin to lose viability significantly within 8 h of incubation in the presence of 3-BP. The Km (0.3 mmol/l) for intracellular accumulation of 3-BP in MM cells is 24 times lower than that in control cells (7.2 mmol/l). Therefore, the uptake of 3-BP by MM cells is significantly higher than that by peripheral blood mononuclear cells. Further, the IC50 values for human MM cells and control peripheral blood mononuclear cells are 24 and 58 µmol/l, respectively. Therefore, specificity and selectivity of 3-BP toward MM cancer cells are evident on the basis of the above. In MM cells the transcription levels of the gene encoding the monocarboxylate transporter MCT1 is significantly amplified compared with control cells. The level of intracellular ATP in MM cells decreases by over 90% within 1 h after addition of 100 µmol/l 3-BP. The cytotoxicity of 3-BP, exemplified by a marked decrease in viability of MM cells, is potentiated by the inhibitor of glutathione synthesis buthionine sulfoximine. In addition, the lack of mutagenicity and its superior capacity relative to Glivec to kill MM cancer cells are presented in this study.

  7. Short Communication: Preferential Killing of HIV Latently Infected CD4+ T Cells by MALT1 Inhibitor

    Science.gov (United States)

    Li, Hongmei; He, Hui; Gong, Leyi; Fu, Mingui

    2016-01-01

    Abstract We report that the addition of an host paracaspase MALT1 inhibitor, MI-2, to HIV latently infected ACH-2, Jurkat E4, and J-LAT cells accelerated cell death in the presence of cell stimuli or the protein kinase C agonist, bryostatin 1. MI-2-mediated cell death correlated with the induction of the cellular RNase MCPIP1 and requires the presence of viral component(s). Altogether, the combination of MI-2 and bryostatin 1 displays selective killing of HIV latently infected CD4+ T cells. PMID:26728103

  8. Short Communication: Preferential Killing of HIV Latently Infected CD4(+) T Cells by MALT1 Inhibitor.

    Science.gov (United States)

    Li, Hongmei; He, Hui; Gong, Leyi; Fu, Mingui; Wang, Tony T

    2016-02-01

    We report that the addition of an host paracaspase MALT1 inhibitor, MI-2, to HIV latently infected ACH-2, Jurkat E4, and J-LAT cells accelerated cell death in the presence of cell stimuli or the protein kinase C agonist, bryostatin 1. MI-2-mediated cell death correlated with the induction of the cellular RNase MCPIP1 and requires the presence of viral component(s). Altogether, the combination of MI-2 and bryostatin 1 displays selective killing of HIV latently infected CD4(+) T cells.

  9. Kinase domain insert containing receptor promoter controlled suicide gene system selectively kills human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Wen-Yu Yang; Zong-Hai Huang; Li-Jun Lin; Zhou Li; Jing-Long Yu; Hui-Juan Song; Yong Qian; Xiao-Yan Che

    2006-01-01

    AIM: To study the selective killing of human umbilical vein endothelial cells (HUVECs) by a double suicide gene under the regulation of a kinase domain insert containing receptor (KDR) promoter and mediated by an adenoviral gene vector.METHODS: Human KDR promoter was cloned by polymerase chain reaction (PCR), and two recombinant adenoviral plasmids pAdKDR-CdglyTK, pAdCMV-CDglyTK were constructed according to a two-step transformation protocol. These two newly constructed plasmids were then transfected into 293 packaging cells to grow adenovirus, which were further multiplied and purified.HUVECs and LoVo cells were infected with either of the two resultant recombinant adenoviruses (AdKDR-CDglyTK and AdCMV-CDglyTK) respectively, and the infection rates were estimated by detection of green fluorescent protein (GFP) expression. Infected cells were cultured in culture media containing different concentrations of 5-fluorocytosine (5-FC) and ganciclovir (GCV), and the killing effects were measured.RESULTS: The two recombinant adenoviral plasmids pAdKDR-CdglyTK, pAdCMV-CDglyTK were successfully constructed and transfected into 293 cells. The resultant recombinant adenoviruses infected cells caused similar infection rates; and the infected cells exhibited different sensitivity to the prodrugs: HUVECs infected with AdCMV-CDglyTK and LoVo cells infected with AdCMVCDglyTK were highly sensitive to the prodrugs, and HUVECs infected with AdKDR-CDglyTK were similarly sensitive but significantly more sensitive than the LoVo cells infected with AdKDR-CdglyTK (P<0.001).CONCLJSION: Selective killing of HUVECs may be achieved by gene transfer of double suicide gene under the regulation of the KDR promoter. This finding may provide an optional way to target gene therapy of malignant tumors by abrogation of tumor blood vessels.

  10. Modulation of the Culture Supernatant of Decidual Cells with Exogenous Cytokines on Killing Activity of Natural Killer Cells in Early Pregnancy

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective To investigate the important function of cytokines in early pregnancy and to provide basic and experimental evidence for understanding the mechanism of their action. Methods Add interferon-γ (IFN-γ) , interleukin- 2(IL- 2) , interleukin- 6(IL-6) and epidermal growth factor (EGF) to the confluent culturing decidual cells with three different concentrations and harvest the culture supernatant after 12, 24 and 48 h separately. Observe the effect of the supernatant on killing activity of NK cells with radioimmunological assay of 51Cr immersion. Results The culture supernatant of decidual cells can promote the killing activity of NK cells in various degrees, and the effect is independent of the type, concentration and acting time of cytokines. Conclusion In normal pregnancy, decidual cytokine network is in a dynamic equilibri um. Exogenous cytokines would be harm to normal pregnancy by interfering the equi librium state, but the exact mechanism needs further study.

  11. Modulation of the Culture Supernatant of Decidual Cells with Exogenous Cytokines on Killing Activity of Natural Killer Cells in Early Pregnancy

    Institute of Scientific and Technical Information of China (English)

    胡冬梅; 王丽莉; 何援利

    2000-01-01

    Objective To investigate the important function of cytohines in early pregnancy and to provide basic and experimental evidence for understanding the mechanism of their action.Methods Add interferon-γ (IFN-γ) ,interleuhin-2(IL-2) , interleuhin-6(IL-6) andepidermal growth factor(EGF) to the confluent culturing decidual cells with three different concentrations and harvest the culture supernatant after 12, 24 and 48 h separately. Observe the effect of the supernatant on killing activity of NK cells with radioimmunological assay of 51Cr immersion.Results The culture supernatant of decidual cells can promote the killing activity of NK cells in various degrees, and the effect is independent of the type, concentration and acting time of cytokines.Conclusion In normal pregnancy, decidual cytokine network is in a dynamic equilibri-um. Exogenous cytokines would be harm to normal pregnancy by interfering the equi-librium state, but the exact mechanism needs further study.

  12. Real-time dynamic optical imaging of ACC-M tumor cells killed by HSV-tk/ACV system.

    Science.gov (United States)

    Xiong, Tao; Li, Yongjin; Li, Zhiyang; Xie, Xiangmo; Lu, Lisha

    2013-01-01

    HSV-tk/ACV induced and killed human adenoid cystic carcinoma cell (ACC-M) in vivo and in vitro, which were observed through optical imaging and green fluorescence protein (GFP) tagging technique. ACC-M was transfected with TK-GFP, and the single clone cell ACC-M-TK-GFP was selected by G418. With fluorescent stereomicroscope, whole-body fluorescent imaging system and fluorescent microscope, we could observe ACV treated ACC-M-TK-GFP cells in cell level and nude mice. The therapies of tumor were visualized clearly with optical imaging. This study proves that optical imaging is a very good approach for studying the effect of HSV-tk/ACV on the ACC-M tumor cells and decreasing the amount of vessel about tumors cell. Optical imaging will become a visual groundwork for monitoring tumor growth and evaluating in vivo curative effect of antitumor drugs.

  13. Assessment of attachment, ingestion, and killing of Escherichia coli by bovine polymorphonuclear cells with combined micromethods.

    Science.gov (United States)

    Rainard, P

    1985-11-01

    A set of microassays separately measuring attachment, ingestion, and overall killing of Escherichia coli by bovine granulocytes was devised and its analytical potential used to test the effect of drugs which block intracellular killing: sodium azide, phenylbutazone, chloroquine phosphate were all inactive, suggesting that O2-dependent systems were not the sole pathway involved in the killing of E.coli by granulocytes. The microtechniques were also used to investigate the opsonic requirements for phagocytosis of two E.coli strains. Absorption of normal bovine serum with the homologous and the heterologous strains showed that specific antibodies were necessary to induce attachment of bacteria to phagocytes. Once bound to granulocytes, the unencapsulated strain P4 was engulfed, whereas for the encapsulated strain B117, complement was required for the internalization step of phagocytosis. With immune serum the need for complement was not absolute.

  14. Combinatorial BTK and MALT1 inhibition augments killing of CD79 mutant diffuse large B cell lymphoma.

    Science.gov (United States)

    Nagel, Daniel; Bognar, Miriam; Eitelhuber, Andrea C; Kutzner, Kerstin; Vincendeau, Michelle; Krappmann, Daniel

    2015-12-08

    Survival of activated B cell-subtype (ABC) of diffuse large B cell lymphoma (DLBCL) is driven by chronic B cell receptor (BCR) signaling that activates the canonical NF-κB pathway. Inhibition of BTK by Ibrutinib has been shown to kill ABC DLBCL cells that carry activating mutations in the BCR adaptor CD79. However, mutations in BTK or in downstream components such as CARMA1/CARD11 can render lymphomas Ibrutinib resistant. Therefore, we assessed here the simultaneous inhibition of BTK and the protease MALT1 that acts downstream of CARMA1 and is essential for ABC DLBCL tumor growth. We show that in CD79 mutant cells BTK is a crucial upstream regulator of MALT1, but dispensable in CARMA1 mutant ABC DLBCL. Combined inhibition of BTK by Ibrutinib and MALT1 by S-Mepazine additively impaired MALT1 cleavage activity and expression of NF-κB pro-survival factors. Thereby, combinatorial Ibrutinib and S-Mepazine treatment enhanced killing of CD79 mutant ABC DLBCL cells. Moreover, while expression of oncogenic CARMA1 in CD79 mutant cells conferred Ibrutinib resistance, double mutant cells were still sensitive to MALT1 inhibition by S-Mepazine. Thus, based on the genetic background combinatorial BTK and MALT1 inhibition may improve effectiveness of therapeutic treatment and reduce the chances for the development of drug resistances.

  15. Valproic acid enhance the killing effect of cisplatin on HO8910 cells%丙戊酸钠协同顺铂对HO8910细胞杀伤作用机制研究

    Institute of Scientific and Technical Information of China (English)

    张丽娜; 马杰; 曹振平; 李彩荣; 冯定庆; 赵卫东

    2011-01-01

    Objective : To study the mechanism of which Valproic acid enhance the killing effect of cisplatin on human ovanan cancer cell line HO-8910. Methods: Pretreated with lμg/ml DDP and 3mmol/L VPA,the growth inhibition ratio of HO-8910 cells was detected with MTT. Morphological changes of cells in each group were observed with inverted microscope.Flow cytometry analyzed the cell cycle changes. The expression of Ac-H3 , p53 , p21 CyclinD1 gene was deteeed by semiquantitative RT- PCR and Western blot. Results : ( 1 ) VPA can inhibit HO8910 cell growth in a dose and time-dependent. The suppression ratio of treated with VPA combined DDP was higher than that of DDP treatment ( P<0. 05 ) . (2) Cell morphology changes can clearly be seen in the VPA group under light microscope : cell volume reduced , change to long spindle , membrane shrinkage , poor adherence. ( 3 ) VPA arrest ovarian cancer HO8910 cell cycle at G1 phase,but in VPA+DDP group G1/G2 phase cells increased as well as S phase decreased. (4)VPA and VPA combination with DDP can significantly enhance the acetylation of Histone H3 ,increase expression of p53 and p21 and reduce Cyclin Dl expression( P<0. 05 ).Conclusion : VPA can enhance the inhibitory effect of DDP on cell HO89IO;The mechanism of action may include increase in the level of histone acetylation and the expression of p53, p21 gene,decrease in the expression of Cyclin Dl and induction of cell cycle arrest.%目的:探讨丙戊酸钠(VPA)协同顺铂(DDP)对人卵巢癌HO8910细胞的杀伤作用及机制.方法:以1μg/ml DDP联合3mmol/L VPA处理细胞,MTT法检测细胞生长抑制率;光镜下观察药物作用下各组细胞形态学的改变;流式细胞术检测细胞周期阻滞情况;RT-PCR法检测p53、p21 mRNA水平表达的改变;Western blot法检测Ac-H3、p53、p21、Cyclin D1蛋白水平表达的改变.结果:(1)VPA能明显抑制HO8910细胞生长,且呈剂量和时间依赖性,VPA+DDP处理组抑制率高于DDP处理组,组

  16. Glycan elongation beyond the mucin associated Tn antigen protects tumor cells from immune-mediated killing.

    Directory of Open Access Journals (Sweden)

    Caroline B Madsen

    Full Text Available Membrane bound mucins are up-regulated and aberrantly glycosylated during malignant transformation in many cancer cells. This results in a negatively charged glycoprotein coat which may protect cancer cells from immune surveillance. However, only limited data have so far demonstrated the critical steps in glycan elongation that make aberrantly glycosylated mucins affect the interaction between cancer cells and cytotoxic effector cells of the immune system. Tn (GalNAc-Ser/Thr, STn (NeuAcα2-6GalNAc-Ser/Thr, T (Galβ1-3GalNAc-Ser/Thr, and ST (NeuAcα2-6Galβ1-3GalNAc-Ser/Thr antigens are recognized as cancer associated truncated glycans, and are expressed in many adenocarcinomas, e.g. breast- and pancreatic cancer cells. To investigate the role of the cancer associated glycan truncations in immune-mediated killing we created glyco-engineered breast- and pancreatic cancer cells expressing only the shortest possible mucin-like glycans (Tn and STn. Glyco-engineering was performed by zinc finger nuclease (ZFN knockout (KO of the Core 1 enzyme chaperone COSMC, thereby preventing glycan elongation beyond the initial GalNAc residue in O-linked glycans. We find that COSMC KO in the breast and pancreatic cancer cell lines T47D and Capan-1 increases sensitivity to both NK cell mediated antibody-dependent cellular-cytotoxicity (ADCC and cytotoxic T lymphocyte (CTL-mediated killing. In addition, we investigated the association between total cell surface expression of MUC1/MUC16 and NK or CTL mediated killing, and observed an inverse correlation between MUC16/MUC1 expression and the sensitivity to ADCC and CTL-mediated killing. Together, these data suggest that up-regulation of membrane bound mucins protects cells from immune mediated killing, and that particular glycosylation steps, as demonstrated for glycan elongation beyond Tn and STn, can be important for fine tuning of the immune escape mechanisms in cancer cells.

  17. Vitamin D and the human antimicrobial peptide LL-37 enhance group a streptococcus resistance to killing by human cells.

    Science.gov (United States)

    Love, John F; Tran-Winkler, Hien J; Wessels, Michael R

    2012-10-23

    The CsrRS two-component regulatory system of group A Streptococcus (GAS; Streptococcus pyogenes) responds to subinhibitory concentrations of the human antimicrobial peptide LL-37. LL-37 signaling through CsrRS results in upregulation of genes that direct synthesis of virulence factors, including the hyaluronic acid capsule and streptolysin O (SLO). Here, we demonstrate that a consequence of this response is augmented GAS resistance to killing by human oropharyngeal keratinocytes, neutrophils, and macrophages. LL-37-induced upregulation of SLO and hyaluronic acid capsule significantly reduced internalization of GAS by keratinocytes and phagocytic killing by neutrophils and macrophages. Because vitamin D induces LL-37 production by macrophages, we tested its effect on macrophage killing of GAS. In contrast to the reported enhancement of macrophage function in relation to other pathogens, treatment of macrophages with 1α,25-dihydroxy-vitamin D3 paradoxically reduced the ability of macrophages to control GAS infection. These observations demonstrate that LL-37 signals through CsrRS to induce a virulence phenotype in GAS characterized by heightened resistance to ingestion and killing by both epithelial cells and phagocytes. By inducing LL-37 production in macrophages, vitamin D may contribute to this paradoxical exacerbation of GAS infection. IMPORTANCE It remains poorly understood why group A Streptococcus (GAS) causes asymptomatic colonization or localized throat inflammation in most individuals but rarely progresses to invasive infection. The human antimicrobial peptide LL-37, which is produced as part of the innate immune response to GAS infection, signals through the GAS CsrRS two-component regulatory system to upregulate expression of multiple virulence factors. This study reports that two CsrRS-regulated GAS virulence factors-streptolysin O and the hyaluronic acid capsule-are critical in LL-37-induced resistance of GAS to killing by human throat epithelial cells

  18. Carbon-ion beam irradiation kills X-ray-resistant p53-null cancer cells by inducing mitotic catastrophe.

    Directory of Open Access Journals (Sweden)

    Napapat Amornwichet

    Full Text Available BACKGROUND AND PURPOSE: To understand the mechanisms involved in the strong killing effect of carbon-ion beam irradiation on cancer cells with TP53 tumor suppressor gene deficiencies. MATERIALS AND METHODS: DNA damage responses after carbon-ion beam or X-ray irradiation in isogenic HCT116 colorectal cancer cell lines with and without TP53 (p53+/+ and p53-/-, respectively were analyzed as follows: cell survival by clonogenic assay, cell death modes by morphologic observation of DAPI-stained nuclei, DNA double-strand breaks (DSBs by immunostaining of phosphorylated H2AX (γH2AX, and cell cycle by flow cytometry and immunostaining of Ser10-phosphorylated histone H3. RESULTS: The p53-/- cells were more resistant than the p53+/+ cells to X-ray irradiation, while the sensitivities of the p53+/+ and p53-/- cells to carbon-ion beam irradiation were comparable. X-ray and carbon-ion beam irradiations predominantly induced apoptosis of the p53+/+ cells but not the p53-/- cells. In the p53-/- cells, carbon-ion beam irradiation, but not X-ray irradiation, markedly induced mitotic catastrophe that was associated with premature mitotic entry with harboring long-retained DSBs at 24 h post-irradiation. CONCLUSIONS: Efficient induction of mitotic catastrophe in apoptosis-resistant p53-deficient cells implies a strong cancer cell-killing effect of carbon-ion beam irradiation that is independent of the p53 status, suggesting its biological advantage over X-ray treatment.

  19. Complete Cell Killing by Applying High Hydrostatic Pressure for Acellular Vascular Graft Preparation

    Directory of Open Access Journals (Sweden)

    Atsushi Mahara

    2014-01-01

    Full Text Available Pressure treatment has been developed in tissue engineering application. Although the tissue scaffold prepared by a ultrahydrostatic pressure treatment has been reported, an excessive pressure has a potential to disrupt a structure of extracellular matrix through protein denaturation. It is important to understand the suitable low-pressure condition and mechanisms for cell killing. In this study, cellular morphology, mitochondria activity, and membrane permeability of mammalian cells with various pressure treatments were investigated with in vitro models. When the cells were treated with a pressure of 100 MPa for 10 min, cell morphology and adherence were the same as an untreated cells. Dehydrogenase activity in mitochondria was almost the same as untreated cells. On the other hand, when the cells were treated with the pressure of more than 200 MPa, the cells did not adhere, and the dehydrogenase activity was completely suppressed. However, green fluorescence was observed in the live/dead staining images, and the cells were completely stained as red after above 500 MPa. That is, membrane permeability was disturbed with the pressure treatment of above 500 MPa. These results indicated that the pressure of 200 MPa for 10 min was enough to induce cell killing through inactivation of mitochondria activity.

  20. A Small-Molecule Inhibitor of BCL6 Kills DLBCL Cells In Vitro and In Vivo

    Energy Technology Data Exchange (ETDEWEB)

    Cerchietti, L.C.; Ghetu, A.F.; Zhu, X.; Da Silva, G.F.; Zhong, S.; Matthews, M.; Bunting, K.L.; Polo, J.M.; Fares, C.; Arrowsmith, C.H.; Yang, S.N.; Garcia, M.; Coop, A.; Mackerell, A.D.; Prive, G.G.; Melnick, A. (Cornell)

    2010-09-22

    The BCL6 transcriptional repressor is the most frequently involved oncogene in diffuse large B cell lymphoma (DLBCL). We combined computer-aided drug design with functional assays to identify low-molecular-weight compounds that bind to the corepressor binding groove of the BCL6 BTB domain. One such compound disrupted BCL6/corepressor complexes in vitro and in vivo, and was observed by X-ray crystallography and NMR to bind the critical site within the BTB groove. This compound could induce expression of BCL6 target genes and kill BCL6-positive DLBCL cell lines. In xenotransplantation experiments, the compound was nontoxic and potently suppressed DLBCL tumors in vivo. The compound also killed primary DLBCLs from human patients.

  1. Clinical-scale laser-based scanning and processing of live cells: selective photothermal killing of fluorescent tumor targets for autologous stem cell transplantation

    Science.gov (United States)

    Koller, Manfred R.; Hanania, Elie G.; Eisfeld, Timothy; O'Neal, Robert A.; Khovananth, Kevin M.; Palsson, Bernhard O.

    2001-04-01

    High-dose chemotherapy, followed by autologous hematopoietic stem cell (HSC) transplantation, is widely used for the treatment of cancer. However, contaminating tumor cells within HSC harvests continue to be of major concern since re-infused tumor cells have proven to contribute to disease relapse. Many tumor purging methods have been evaluated, but all leave detectable tumor cells in the transplant and result in significant loss of HSCs. These shortcomings cause engraftment delays and compromise the therapeutic value of purging. A novel approach integrating automated scanning cytometry, image analysis, and selective laser-induced killing of labeled cells within a cell mixture is described here. Non-Hodgkin's lymphoma (NHL) cells were spiked into cell mixtures, and fluorochrome-conjugated antibodies were used to label tumor cells within the mixture. Cells were then allowed to settle on a surface, and as the surface was scanned with a fluorescence excitation source, a laser pulse was fired at every detected tumor cell using high-speed beam steering mirrors. Tumor cells were selectively killed with little effect on adjacent non-target cells, demonstrating the feasibility of this automated cell processing approach. This technology has many potential research and clinical applications, one example of which is tumor cell purging for autologous HSC transplantation.

  2. Simultaneous Targeting of FcγRs and FcαRI Enhances Tumor Cell Killing

    NARCIS (Netherlands)

    Brandsma, Arianne M; Ten Broeke, Toine; Nederend, Maaike; Meulenbroek, Laura A P M; van Tetering, Geert; Meyer, Saskia; Jansen, J H Marco; Beltrán Buitrago, M Alejandra; Nagelkerke, Sietse Q; Németh, István; Ubink, Ruud; Rouwendal, Gerard; Lohse, Stefan; Valerius, Thomas; Leusen, Jeanette H W; Boross, Peter

    2015-01-01

    Efficacy of anticancer monoclonal antibodies (mAb) is limited by the exhaustion of effector mechanisms. IgG mAbs mediate cellular effector functions through FcγRs expressed on effector cells. IgA mAbs can also induce efficient tumor killing both in vitro and in vivo. IgA mAbs recruit FcαRI-expressin

  3. Killed but metabolically active Leishmania infantum as a novel whole-cell vaccine for visceral leishmaniasis.

    Science.gov (United States)

    Bruhn, Kevin W; Birnbaum, Ron; Haskell, Jacquelyn; Vanchinathan, Veena; Greger, Stephanie; Narayan, Rupa; Chang, Pei-Lin; Tran, Thu Anh; Hickerson, Suzanne M; Beverley, Stephen M; Wilson, Mary E; Craft, Noah

    2012-04-01

    There are currently no effective vaccines for visceral leishmaniasis, the second most deadly parasitic infection in the world. Here, we describe a novel whole-cell vaccine approach using Leishmania infantum chagasi promastigotes treated with the psoralen compound amotosalen (S-59) and low doses of UV A radiation. This treatment generates permanent, covalent DNA cross-links within parasites and results in Leishmania organisms termed killed but metabolically active (KBMA). In this report, we characterize the in vitro growth characteristics of both KBMA L. major and KBMA L. infantum chagasi. Concentrations of S-59 that generate optimally attenuated parasites were identified. Like live L. infantum chagasi, KBMA L. infantum chagasi parasites were able to initially enter liver cells in vivo after intravenous infection. However, whereas live L. infantum chagasi infection leads to hepatosplenomegaly in mice after 6 months, KBMA L. infantum chagasi parasites were undetectable in the organs of mice at this time point. In vitro, KBMA L. infantum chagasi retained the ability to enter macrophages and induce nitric oxide production. These characteristics of KBMA L. infantum chagasi correlated with the ability to prophylactically protect mice via subcutaneous vaccination at levels similar to vaccination with live, virulent organisms. Splenocytes from mice vaccinated with either live L. infantum chagasi or KBMA L. infantum chagasi displayed similar cytokine patterns in vitro. These results suggest that KBMA technology is a potentially safe and effective novel vaccine strategy against the intracellular protozoan L. infantum chagasi. This approach may represent a new method for whole-cell vaccination against other complex intracellular pathogens.

  4. The Effect of Bacteriophage Preparations on Intracellular Killing of Bacteria by Phagocytes.

    Science.gov (United States)

    Jończyk-Matysiak, Ewa; Łusiak-Szelachowska, Marzanna; Kłak, Marlena; Bubak, Barbara; Międzybrodzki, Ryszard; Weber-Dąbrowska, Beata; Żaczek, Maciej; Fortuna, Wojciech; Rogóż, Paweł; Letkiewicz, Sławomir; Szufnarowski, Krzysztof; Górski, Andrzej

    2015-01-01

    Intracellular killing of bacteria is one of the fundamental mechanisms against invading pathogens. Impaired intracellular killing of bacteria by phagocytes may be the reason of chronic infections and may be caused by antibiotics or substances that can be produced by some bacteria. Therefore, it was of great practical importance to examine whether phage preparations may influence the process of phagocyte intracellular killing of bacteria. It may be important especially in the case of patients qualified for experimental phage therapy (approximately half of the patients with chronic bacterial infections have their immunity impaired). Our analysis included 51 patients with chronic Gram-negative and Gram-positive bacterial infections treated with phage preparations at the Phage Therapy Unit in Wroclaw. The aim of the study was to investigate the effect of experimental phage therapy on intracellular killing of bacteria by patients' peripheral blood monocytes and polymorphonuclear neutrophils. We observed that phage therapy does not reduce patients' phagocytes' ability to kill bacteria, and it does not affect the activity of phagocytes in patients with initially reduced ability to kill bacteria intracellularly. Our results suggest that experimental phage therapy has no significant adverse effects on the bactericidal properties of phagocytes, which confirms the safety of the therapy.

  5. The Effect of Bacteriophage Preparations on Intracellular Killing of Bacteria by Phagocytes

    Directory of Open Access Journals (Sweden)

    Ewa Jończyk-Matysiak

    2015-01-01

    Full Text Available Intracellular killing of bacteria is one of the fundamental mechanisms against invading pathogens. Impaired intracellular killing of bacteria by phagocytes may be the reason of chronic infections and may be caused by antibiotics or substances that can be produced by some bacteria. Therefore, it was of great practical importance to examine whether phage preparations may influence the process of phagocyte intracellular killing of bacteria. It may be important especially in the case of patients qualified for experimental phage therapy (approximately half of the patients with chronic bacterial infections have their immunity impaired. Our analysis included 51 patients with chronic Gram-negative and Gram-positive bacterial infections treated with phage preparations at the Phage Therapy Unit in Wroclaw. The aim of the study was to investigate the effect of experimental phage therapy on intracellular killing of bacteria by patients’ peripheral blood monocytes and polymorphonuclear neutrophils. We observed that phage therapy does not reduce patients’ phagocytes’ ability to kill bacteria, and it does not affect the activity of phagocytes in patients with initially reduced ability to kill bacteria intracellularly. Our results suggest that experimental phage therapy has no significant adverse effects on the bactericidal properties of phagocytes, which confirms the safety of the therapy.

  6. IL-1α-induced microvascular endothelial cells promote neutrophil killing by increasing MMP-9 concentration and lysozyme activity.

    Science.gov (United States)

    Liu, Xiaoye; Dong, Hong; Wang, Mingming; Gao, Ying; Zhang, Tao; Hu, Ge; Duan, Huiqing; Mu, Xiang

    2016-02-01

    The recruitment of neutrophils by endothelial cells during infection has been extensively studied, but little is known about the regulation of neutrophils activity by endothelial cells. To examine the role of microvascular endothelial cells in neutrophil killing, we established a transmigration model using rat intestinal microvascular endothelial cells (RIMVECs) and measured the extracellular and intracellular killing of Escherichia coli, Lactobacillus acidophilus, and Staphylococcus aureus by transendothelial neutrophils. We observed that blood neutrophils engulfed bacteria but did not kill them, and lipopolysaccharide- or hemolysin-injured RIMVECs inhibited the extracellular and intracellular bactericidal activity of transendothelial neutrophils. In comparison, interleukin-1α-induced RIMVECs promoted the extracellular and intracellular killing activity of transendothelial neutrophils and significantly increased MMP-9 concentration and lysozyme activity in transendothelial neutrophils (p neutrophils and bacterial toxin damage of endothelial cells led to reduction in bactericidal activity of transendothelial neutrophils. These findings offered new insight into the role of endothelial cells in the bactericidal activity of neutrophils.

  7. Adenanthin targets peroxiredoxin I/II to kill hepatocellular carcinoma cells.

    Science.gov (United States)

    Hou, J-K; Huang, Y; He, W; Yan, Z-W; Fan, L; Liu, M-H; Xiao, W-L; Sun, H-D; Chen, G-Q

    2014-09-04

    Adenanthin, a natural diterpenoid isolated from the leaves of Isodon adenanthus, has recently been reported to induce leukemic cell differentiation by targeting peroxiredoxins (Prx) I and II. On the other hand, increasing lines of evidence propose that these Prx proteins would become potential targets to screen drugs for the prevention and treatment of solid tumors. Therefore, it is of significance to explore the potential activities of adenanthin on solid tumor cells. Here, we demonstrate that Prx I protein is essential for the survival of hepatocellular carcinoma (HCC) cells, and adenanthin can kill these malignant liver cells in vitro and xenografts. We also show that the cell death-inducing activity of adenanthin on HCC cells is mediated by the increased reactive oxygen species (ROS) levels. Furthermore, the silencing of Prx I or Prx II significantly enhances the cytotoxic activity of adenanthin on HCC, whereas the ectopic expression of Prx I and Prx II but not their mutants of adenanthin-bound cysteines can rescue adenanthin-induced cytotoxicity in Prxs-silenced HCC cells. Taken together, our results propose that adenanthin targets Prx I/II to kill HCC cells and its therapeutic significance warrants to be further explored in HCC patients.

  8. Targeted killing of a mammalian cell based upon its specialized metabolic state.

    Science.gov (United States)

    Alexander, Peter B; Wang, Jian; McKnight, Steven L

    2011-09-20

    Mouse ES cells use a mitochondrial threonine dehydrogenase (TDH) enzyme to catabolize threonine into glycine and acetyl-CoA. Measurements of mRNA abundance have given evidence that ES cells express upwards of 1,000-fold higher levels of TDH mRNA than any of seven other mouse tissues tested. When cell culture medium is deprived of threonine, ES cells rapidly discontinue DNA synthesis, arrest cell division, and eventually die. Such studies led to the conclusion that mouse ES cells exist in a threonine-dependent metabolic state. Proceeding with the assumption that the active TDH enzyme should be essential for the growth and viability of mouse ES cells, we performed a drug screen in search of specific inhibitors of the purified TDH enzyme. Such efforts led to the discovery of a class of quinazolinecarboxamide (Qc) compounds that inhibit the ability of the TDH enzyme to catabolize threonine into glycine and acetyl-CoA. Administration of Qc inhibitors of TDH to mouse ES cells impeded cell growth and resulted in the induction of autophagy. By contrast, the same chemicals failed to affect the growth of HeLa cells at concentrations 300-fold higher than that required to kill mouse ES cells. It was likewise observed that the Qc class of TDH inhibitors failed to affect the growth or viability of ES cell-derived embryoid body cells known to have extinguished TDH expression. These studies demonstrate how it is possible to kill a specific mammalian cell type on the basis of its specialized metabolic state.

  9. Cryptococcus Neoformans Modulates Extracellular Killing by Neutrophils

    OpenAIRE

    Qureshi, Asfia; Grey, Angus; Rose, Kristie L; Schey, Kevin L.; Del Poeta, Maurizio

    2011-01-01

    We recently established a key role for host sphingomyelin synthase (SMS) in regulating the killing activity of neutrophils against Cryptococcus neoformans. In this paper, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and natural killer (NK) cells (Tgε26 mice). To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in...

  10. Protection of hepatocytes from cytotoxic T cell mediated killing by interferon-alpha.

    Directory of Open Access Journals (Sweden)

    Christian B Willberg

    Full Text Available BACKGROUND: Cellular immunity plays a key role in determining the outcome of hepatitis C virus (HCV infection, although the majority of infections become persistent. The mechanisms behind persistence are still not clear; however, the primary site of infection, the liver, may be critical. We investigated the ability of CD8+ T-cells (CTL to recognise and kill hepatocytes under cytokine stimulation. METHODS/PRINCIPLE FINDINGS: Resting hepatocytes cell lines expressed low levels of MHC Class I, but remained susceptible to CTL cytotoxicity. IFN-alpha treatment, in vitro, markedly increased hepatocyte MHC Class I expression, however, reduced sensitivity to CTL cytotoxicity. IFN-alpha stimulated hepatocyte lines were still able to present antigen and induce IFN-gamma expression in interacting CTL. Resistance to killing was not due to the inhibition of the FASL/FAS- pathway, as stimulated hepatocytes were still susceptible to FAS-mediated apoptosis. In vitro stimulation with IFN-alpha, or the introduction of a subgenomic HCV replicon into the HepG2 line, upregulated the expression of the granzyme-B inhibitor-proteinase inhibitor 9 (PI-9. PI-9 expression was also observed in liver tissue biopsies from patients with chronic HCV infection. CONCLUSION/SIGNIFICANCE: IFN-alpha induces resistance in hepatocytes to perforin/granzyme mediate CTL killing pathways. One possible mechanism could be through the expression of the PI-9. Hindrance of CTL cytotoxicity could contribute to the chronicity of hepatic viral infections.

  11. Dynamic visualization the whole process of cytotoxic T lymphocytes killing the B16 tumor cells in vitro

    Science.gov (United States)

    Qi, Shuhong; Zhang, Zhihong

    2016-03-01

    Cytotoxic T lymphocytes (CTLs) played a key role in the immune system to destroy the tumor cells. Although some mechanisms of CTLs killing the tumor cells are revealed already, the dynamic information of CTLs interaction with tumor cells are still not known very clearly. Here we used confocal microscopy to visualize the whole process of CTLs killing the tumor cells in vitro. The imaging data showed that CTLs destroyed the target tumor cells rapidly and efficiently. Several CTLs surrounded one or some tumor cells and the average time for CTLs destroying one tumor cell is just a few minutes in vitro. The study displayed the temporal events of CTLs interacting with tumor cells at the beginning and finally killing them and directly presented the efficient tumor cell cytotoxicity of the CTLs. The results helped us to deeply understand the mechanism of the CTLs destroying the tumor cells and to develop the cancer immunotherapy.

  12. A model of radiation-induced cell killing: insights into mechanisms and applications for hadron therapy.

    Science.gov (United States)

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

    2013-09-01

    A mechanism-based, two-parameter biophysical model of cell killing was developed with the aim of elucidating the mechanisms underlying radiation-induced cell death and predicting cell killing by different radiation types, including protons and carbon ions at energies and doses of interest for cancer therapy. The model assumed that certain chromosome aberrations (dicentrics, rings and large deletions, called "lethal aberrations") lead to clonogenic inactivation, and that aberrations derive from μm-scale misrejoining of chromatin fragments, which in turn are produced by "dirty" double-strand breaks called "cluster lesions" (CLs). The average numbers of CLs per Gy per cell were left as a semi-free parameter and the threshold distance for chromatin-fragment rejoining was defined the second parameter. The model was "translated" into Monte Carlo code and provided simulated survival curves, which were compared with survival data on V79 cells exposed to protons, carbon ions and X rays. The agreement was good between simulations and survival data and supported the assumptions of the model at least for doses up to a few Gy. Dicentrics, rings and large deletions were found to be lethal not only for AG1522 cells exposed to X rays, as already reported by others, but also for V79 cells exposed to protons and carbon ions of different energies. Furthermore, the derived CL yields suggest that the critical DNA lesions leading to clonogenic inactivation are more complex than "clean" DSBs. After initial validation, the model was applied to characterize the particle and LET dependence of proton and carbon cell killing. Consistent with the proton data, the predicted fraction of inactivated cells after 2 Gy protons was 40-50% below 7.7 keV/μm, increased by a factor ∼1.6 between 7.7-30.5 keV/μm, and decreased by a factor ∼1.1 between 30.5-34.6 keV/μm. These LET values correspond to proton energies below a few MeV, which are always present in the distal region of hadron therapy

  13. Killing of targets by effector CD8 T cells in the mouse spleen follows the law of mass action

    Energy Technology Data Exchange (ETDEWEB)

    Ganusov, Vitaly V [Los Alamos National Laboratory

    2009-01-01

    In contrast with antibody-based vaccines, it has been difficult to measure the efficacy of T cell-based vaccines and to correlate the efficacy of CD8 T cell responses with protection again viral infections. In part, this difficulty is due to poor understanding of the in vivo efficacy of CD8 T cells produced by vaccination. Using a: recently developed experimental method of in vivo cytotoxicity we have investigated quantitative aspects of killing of peptide-pulsed targets by effector and memory CD8 T cells, specific to three epitopes of lymphocytic choriomeningitis virus (LCMV), in the mouse spleen. By analyzing data on killing of targets with varying number of epitope-specific effector and memory CD8 T cells, we find that killing of targets by effectors follows the law of mass-action, that is the death rate of peptide-pulsed targets is proportional to the frequency of CTLs in the spleen. In contrast, killing of targets by memory CD8 T cells does not follow the mass action law because the death rate of targets saturates at high frequencies of memory CD8 T cells. For both effector and memory cells, we also find little support for the killing term that includes the decrease of the death rate of targets with target cell density. Interestingly, our analysis suggests that at low CD8 T cell frequencies, memory CD8 T cells on the per capita basis are more efficient at killing peptide-pulsed targets than effectors, but at high frequencies, effectors are more efficient killers than memory T cells. Comparison of the estimated killing efficacy of effector T cells with the value that is predicted from theoretical physics and based on motility of T cells in lymphoid tissues, suggests that limiting step in the killing of peptide-pulsed targets is delivering the lethal hit and not finding the target. Our results thus form a basis for quantitative understanding of the process of killing of virus-infected cells by T cell responses in tissues and can be used to correlate the

  14. Identification and structural analysis of an L-asparaginase enzyme from guinea pig with putative tumor cell killing properties.

    Science.gov (United States)

    Schalk, Amanda M; Nguyen, Hien-Anh; Rigouin, Coraline; Lavie, Arnon

    2014-11-28

    The initial observation that guinea pig serum kills lymphoma cells marks the serendipitous discovery of a new class of anti-cancer agents. The serum cell killing factor was shown to be an enzyme with L-asparaginase (ASNase) activity. As a direct result of this observation, several bacterial L-asparaginases were developed and are currently approved by the Food and Drug Administration for the treatment of the subset of hematological malignancies that are dependent on the extracellular pool of the amino acid asparagine. As drugs, these enzymes act to hydrolyze asparagine to aspartate, thereby starving the cancer cells of this amino acid. Prior to the work presented here, the precise identity of this guinea pig enzyme has not been reported in the peer-reviewed literature. We discovered that the guinea pig enzyme annotated as H0W0T5_CAVPO, which we refer to as gpASNase1, has the required low Km property consistent with that possessed by the cell-killing guinea pig serum enzyme. Elucidation of the ligand-free and aspartate complex gpASNase1 crystal structures allows a direct comparison with the bacterial enzymes and serves to explain the lack of L-glutaminase activity in the guinea pig enzyme. The structures were also used to generate a homology model for the human homolog hASNase1 and to help explain its vastly different kinetic properties compared with gpASNase1, despite a 70% sequence identity. Given that the bacterial enzymes frequently present immunogenic and other toxic side effects, this work suggests that gpASNase1 could be a promising alternative to these bacterial enzymes.

  15. Identification and Structural Analysis of an l-Asparaginase Enzyme from Guinea Pig with Putative Tumor Cell Killing Properties*

    Science.gov (United States)

    Schalk, Amanda M.; Nguyen, Hien-Anh; Rigouin, Coraline; Lavie, Arnon

    2014-01-01

    The initial observation that guinea pig serum kills lymphoma cells marks the serendipitous discovery of a new class of anti-cancer agents. The serum cell killing factor was shown to be an enzyme with l-asparaginase (ASNase) activity. As a direct result of this observation, several bacterial l-asparaginases were developed and are currently approved by the Food and Drug Administration for the treatment of the subset of hematological malignancies that are dependent on the extracellular pool of the amino acid asparagine. As drugs, these enzymes act to hydrolyze asparagine to aspartate, thereby starving the cancer cells of this amino acid. Prior to the work presented here, the precise identity of this guinea pig enzyme has not been reported in the peer-reviewed literature. We discovered that the guinea pig enzyme annotated as H0W0T5_CAVPO, which we refer to as gpASNase1, has the required low Km property consistent with that possessed by the cell-killing guinea pig serum enzyme. Elucidation of the ligand-free and aspartate complex gpASNase1 crystal structures allows a direct comparison with the bacterial enzymes and serves to explain the lack of l-glutaminase activity in the guinea pig enzyme. The structures were also used to generate a homology model for the human homolog hASNase1 and to help explain its vastly different kinetic properties compared with gpASNase1, despite a 70% sequence identity. Given that the bacterial enzymes frequently present immunogenic and other toxic side effects, this work suggests that gpASNase1 could be a promising alternative to these bacterial enzymes. PMID:25320094

  16. 白细胞介素-2增强淋巴细胞对乳腺癌细胞的杀伤作用%Killing effect of interlukin-2 on breast cancer cells by promoting expansion of lymphocytes in vitro

    Institute of Scientific and Technical Information of China (English)

    熊一全; 黄韬; 赵向旺; 田元; 张景辉

    2015-01-01

    Objective To discuss the inhibitory effect of lymphocytes and interlukin-2 (IL-2)on hormone receptor positive (ER +) breast cancer cells MCF-7,and to investigate the mechanism by which IL-2 promotes the killing effects of lymphocytes.Methods Lymphocytes and IL-2 with different concentrations were co-cultured with MCF-7 cells respectively.Cell counting kit-8 (CCK-8) was used to analyze the the viability of cancer cells.Results In MCF-7 cells treated with 1 000 U IL-2,the effect of IL-2 promoting expansion of lymphocytes was 30.94%,and the killing effect of IL-2 on MCF-7 cells by lymphocytes was 91.24%.The killing effect of lymphocytes on MCF-7 cells was not enhanced by increasing the concentration of IL-2.Conclusion IL-2 prompted the tumor inhibitory effect on ER + breast cancer cells MCF-7%目的 观察淋巴细胞以及白细胞介素-2(IL-2)联合淋巴细胞对于激素受体阳性(ER+)的乳癌细胞株MCF-7的杀伤作用,探讨IL-2增强淋巴细胞的抑癌效应.方法 分别用不同浓度的淋巴细胞以及IL-2加淋巴细胞与乳腺癌细胞株MCF-7共培养,细胞计数试剂盒(CCK-8)法对比分析淋巴细胞对MCF-7细胞的杀伤效应的差异.结果 每孔1 000 U IL-2促进淋巴细胞增殖效应为30.94%;促进淋巴细胞杀伤MCF-7效应为91.24%,增加IL-2的浓度并未增加淋巴细胞杀伤MCF-7能力.结论 IL-2能促进淋巴细胞对MCF-7的杀伤作用.

  17. Killing Effect of Fusion Suicide Genes on Nasopharyngeal Carcinoma Cells in vivo%融合自杀基因对鼻咽癌细胞的体内杀伤作用

    Institute of Scientific and Technical Information of China (English)

    卿菁; 赵素萍; 蒋卫红; 章华

    2013-01-01

    [目的]研究融合自杀基因对鼻咽癌细胞的体内杀伤作用.[方法]培养鼻咽癌CNE-2细胞,建立裸鼠荷瘤模型.随机分为6组,每组5只,A组:脂质体包裹质粒瘤内注射瘤内组;B组:脂质体包裹质粒瘤内注射+肿瘤局部照射γ射线(总剂量9Gy)组;C组:脂质体包裹质粒瘤内注射+腹腔注射5氟胞嘧啶(5-Fc)组;D组:脂质体包裹质粒瘤内注射+肿瘤局部照射γ射线(总剂量9Gy)+腹腔注射5-Fc组;E组:肿瘤局部照射γ射线(总剂量9Gy)+腹腔注射5-Fc,以及空白对照组.记录肿瘤体积变化;计算各组肿瘤抑制率,绘制肿瘤生长曲线;取肿瘤组织进行病理观察;鉴定肿瘤组织中CD/UPRT.UL49基因.肿瘤体积变化的比较采用单向方差分析,组间多重比较采用LSD法.[结果]成功建立鼻咽癌裸鼠荷瘤模型,生长曲线显示:5组瘤体积差异有统计学意义(F=720.684,P<0.01),测序结果证实,除E组外,其余各处理组均检测到目的基因,且Western blot检测结果示:B和D组肿瘤组织内自杀基因的表达高于其余各组.病理切片证实肿块为低分化鳞癌,其中D组肿瘤组织在干预作用下出现明显坏死.[结论] E6.BARF1p.CD/UPRT.UL49自杀基因载体联合前药5-Fc在放射线的作用下,可明显抑制鼻咽癌移植瘤的生长,转染该融合自杀基因可增强放疗敏感性,为鼻咽癌的基因—放射治疗开辟了新思路.%[Purpose] To explore the killing effect of fusion suicide genes in human nasopharyngeal carcinoma (NPC) in vivo.[Methods] NPC CNE-2 cell line was hosted to establish mice model with tumor which were randomly divided into 6 groups,where there were 5 mice with different interventions.The volumes of tumor were measured and compared,the tumor growth inhibition rates were calculated.The tumor tissue was taken out for analysis by Hematoxylin-Eosin staining,and the existence of CD/UPRT.UL49 was detected in tumor tissue.Analysis of variance was used for the comparison of

  18. Effect of Legionella pneumophila sonicate on killing of Listeria monocytogenes by human polymorphonuclear neutrophils and monocytes

    DEFF Research Database (Denmark)

    Rechnitzer, C; Bangsborg, Jette Marie; Shand, G H

    1993-01-01

    polymorphonuclear neutrophils and monocytes. Preincubation of neutrophils with L. pneumophila sonicate did not affect phagocytosis of L. monocytogenes, whereas Listeria killing was significantly inhibited at sonicate concentrations of 1 and 2 mg/ml. The phenol phase of a phenol-water extraction, containing most...... of the lipopolysaccharide (LPS), had no inhibitory effect on the listericidal activity of neutrophils. Killing of Listeria by monocytes was inhibited in a similar manner. The inhibitory activity was mainly recovered in the sonicate fraction above 100 kDa, suggesting that components organized in larger molecular complexes...... are most likely to represent the inhibitory factors. The inhibitory activity of L. pneumophila sonic extract appears to be related to inhibition of killing mechanisms since uptake of Listeria was not affected by the sonicate. Our observations indicate that as Legionella infection progresses, bacterial...

  19. Influence of metronidazole and some electron acceptors on the chlorin e6 photosensitized killing of Ehrlich carcinoma cells

    Science.gov (United States)

    Chekulayev, V.; Shevchuk, Igor; Mihkelsoo, Virgo T.; Kallikorm, A. P.

    1992-06-01

    A decrease in the effectiveness of photosensitized killing of neoplasm cells was observed in the presence of chlorin-e6 at a reduced concentration of oxygen. But when metronidazole (MZ) was injected in vitro as well as in vivo, a significant increase in the photosensitized killing of Ehrlich carcinoma cells by chlorin-e6 was observed. Moreover, contrary to the hematoporphyrin derivative (HpD), MZ increases the effectiveness of photodynamic therapy (PDT) by using chlorin-e6 not only in the hypoxic but also in the aerobic conditions. The interaction between MZ and the excited photosensitizer may account for an increased phototoxicity of chlorin-e6. The formation of cytotoxic nitroimidazole radicals as a result of photochemical processes of type 1 is discussed. This property of the photosensitizer may be successfully used in working out a method of potentiating PDT in combination not only with nitroimidazoles, but also with other electron acceptor compounds (EACp), e.g., quinone antitumor antibiotics.

  20. 嗜酸乳杆菌活菌体和热灭活菌体对 IPEC-J2细胞的黏附和黏附拮抗效应%Adhesion and Adhesion Antagonistic Effect of Live and Heat-Killed Lactobacillus acidophilus on IPEC-J2 Cells

    Institute of Scientific and Technical Information of China (English)

    俞涵丽; 方华; 蔡旋; 徐建雄

    2014-01-01

    The adhesion and adhesion antagonistic effect of live and heat-killed Lactobacillus acidophilus on IPEC-J2 cells were studied in vitro. IPEC-J2 cells were treated with live and heat-killed Lactobacillus acidophi-lus,and then stained the cells and observed the adhesive property. Moreover,the inhibition ratio of live and heat-killed Lactobacillus acidophilus to inhibit the adhesion of Escherichia coli on IPEC-J2 cells through three treatments(competitive inhibition,exclusion inhibition and displacement inhibition)was assessed by eosin-methylene blue agar(EMB)culture medium. The results showed as follows:live and heat-killed Lactobacillus acidophilus all could adhere well to the IPEC-J2 cells. The number of heat-killed bacteria adhering to the per cell[(40±2)CFU]was higher than the number of live bacteria adhering to the per cell[(20±2)CFU]. The two forms of Lactobacillus acidophilus had a good ability to inhibit the Escherichia coli adhere to IPEC-J2 cells. In competitive and displacement inhibition experiments,live bacteria showed a significantly better inhibi-tion ratio than heat-killed bacteria(P<0.05),whereas,in exclusion inhibition experiment,heat-killed bacteria showed a significantly better inhibition ratio than live bacteria( P<0.05). In conclusion,live and heat-killed Lactobacillus acidophilus can adhere well to the IPEC-J2 cells,and are able to reduce the adhesion and inva-sion to IPEC-J2 cells by Escherichia coli. Heat-killed Lactobacillus acidophilus inhibits Escherichia coli adhe-sion mainly through competition for host cell-binding receptors. Apart from competition for host cell-binding re-ceptors,some antibacterial substances from live Lactobacillus acidophilus are able to inhibit the growth and re-production of Escherichia coli.%本试验旨在研究嗜酸乳杆菌活菌体和热灭活菌体在体外模拟条件下对 IPEC-J2细胞的黏附和黏附拮抗效应。用嗜酸乳杆菌活菌体和热灭活菌体处理 IPEC-J2细胞,染色后观察细

  1. Decitabine Treatment of Glioma-Initiating Cells Enhances Immune Recognition and Killing

    Science.gov (United States)

    Riccadonna, Cristina; Yacoub Maroun, Céline; Vuillefroy de Silly, Romain; Boehler, Margaux; Calvo Tardón, Marta; Jueliger, Simone; Taverna, Pietro; Barba, Leticia; Marinari, Eliana; Pellegatta, Serena; Bassoy, Esen Yonca; Martinvalet, Denis; Dietrich, Pierre-Yves; Walker, Paul R.

    2016-01-01

    Malignant gliomas are aggressive brain tumours with very poor prognosis. The majority of glioma cells are differentiated (glioma-differentiated cells: GDCs), whereas the smaller population (glioma-initiating cells, GICs) is undifferentiated and resistant to conventional therapies. Therefore, to better target this pool of heterogeneous cells, a combination of diverse therapeutic approaches is envisaged. Here we investigated whether the immunosensitising properties of the hypomethylating agent decitabine can be extended to GICs. Using the murine GL261 cell line, we demonstrate that decitabine augments the expression of the death receptor FAS both on GDCs and GICs. Interestingly, it had a higher impact on GICs and correlated with an enhanced sensitivity to FASL-mediated cell death. Moreover, the expression of other critical molecules involved in cognate recognition by cytotoxic T lymphocytes, MHCI and ICAM-1, was upregulated by decitabine treatment. Consequently, T-cell mediated killing of both GDCs and GICs was enhanced, as was T cell proliferation after reactivation. Overall, although GICs are described to resist classical therapies, our study shows that hypomethylating agents have the potential to enhance glioma cell recognition and subsequent destruction by immune cells, regardless of their differentiation status. These results support the development of combinatorial treatment modalities including epigenetic modulation together with immunotherapy in order to treat heterogenous malignancies such as glioblastoma. PMID:27579489

  2. A CSPG4-specific immunotoxin kills rhabdomyosarcoma cells and binds to primary tumor tissues.

    Science.gov (United States)

    Brehm, Hannes; Niesen, Judith; Mladenov, Radoslav; Stein, Christoph; Pardo, Alessa; Fey, Georg; Helfrich, Wijnand; Fischer, Rainer; Gattenlöhner, Stefan; Barth, Stefan

    2014-10-01

    The treatment of rhabdomyosarcoma (RMS) remains challenging, with metastatic and alveolar RMS offering a particularly poor prognosis. Therefore, the identification and evaluation of novel antigens, which are suitable targets for immunotherapy, is one attractive possibility to improve the treatment of this disease. Here we show that chondroitin sulfate proteoglycan 4 (CSPG4) is expressed on RMS cell lines and RMS patient material. We evaluated the immunotoxin (IT) αMCSP-ETA', which specifically recognizes CSPG4 on the RMS cell lines RD, FL-OH1, TE-671 and Rh30. It is internalized rapidly, induces apoptosis and thus kills RMS cells selectively. We also demonstrate the specific binding of this IT to RMS primary tumor material from three different patients.

  3. T-peptide Enhances the Killing Effects of Cisplatinum on Lung Cancer

    Directory of Open Access Journals (Sweden)

    Hongyi ZHANG

    2017-02-01

    Full Text Available Background and objective T peptide is extensively used in anti-tumor treatment. The aims of this study were to investigate whether T peptide enhances cisplatinum efficiency while reducing its side effects and to identify its effective mechanisms. Methods (1 Human macrophage U937 cells were treated with T peptide and/or cisplatinum. The levels of tumor necrosis factor-α (TNF-α and interferon-γ (IFN-γ of each group were detected by enzyme-linked immunosorbent assay (ELISA; (2 Xenograft mouse models of human lung cancer were treated with T peptide and/or cisplatinum once every five days for three times. Tumor volumes were measured during treatment; (3 The percentages of macrophages in the peripheral blood of the xenograft mouse models were measured by FACS. Results (1 Compared with other groups, the level of TNF-α was significantly higher in the human macrophage U937 cells that were treated with T peptide combined with cisplatinum. The levels of IFN-γ were significantly higher in human macrophage U937 cells that were treated with T peptide alone or T peptide combined with cisplatinum; (2 In the xenograft mouse models, T peptide combined with cisplatinum treatment significantly inhibited tumor growth without weight loss compared with the other groups; (3 The percentages of macrophages in the peripheral blood were significantly higher in the xenograft mouse models that were treated with T peptide combined with cisplatinum compared with in the other groups. Conclusion T peptide promotes macrophage proliferation and increases tumor cell killing factors (TNF-α, IFN-γ in vitro. Moreover, T peptide enhances the efficacy of cisplatin and reduces its toxicity in vivo.

  4. Neutrophil killing of human umbilical vein endothelial cells is oxygen radical-mediated and enhanced by TNF-. alpha

    Energy Technology Data Exchange (ETDEWEB)

    Dame, M.K.; Varani, J.; Weinberg, J.M.; Ward, P.A. (Univ. of Michigan, Ann Arbor (United States))

    1991-03-11

    Human umbilical vein endothelial cells are sensitive to killing by activated human neutrophils. Killing is inhibited in the presence of catalase and deferoxamine mesylate but not soybean trypsin inhibitor. Reagent hydrogen peroxide can substitute for activated neutrophils in producing endothelial cell injury. These data suggest that lethal injury is due to the production of oxygen radicals by activated neutrophils. In these respects, the human umbilical vein endothelial cells are similar to rat pulmonary artery endothelial cells in that pretreatment with TNF-{alpha} increases sensitivity to injury by activated neutrophils. In part, the increased endothelial cell sensitivity to killing by neutrophils may be due to up-regulation of surface adhesion molecules. However, it was observed that cells passaged more than two times in culture did not demonstrate increased killing after treatment with TNF-{alpha} while up-regulation of neutrophil adhesion could be detected through several additional passages. Although the human umbilical vein endothelial cells are qualitatively similar to rat pulmonary artery endothelial cells in their sensitivity to killing, they are quantitatively much more resistant. What accounts for the relative resistance of the human umbilical vein endothelial cells is not fully understood. In the rat pulmonary artery endothelial cells, killing is known to be dependent on an intraendothelial source of iron. Pre-treatment of the human umbilical vein endothelial cells with 8-hydroxyquinoline-bound iron increased their sensitivity to oxidant injury. These data suggest that the availability of iron within the human umbilical vein endothelial cells may be a limiting factor in sensitivity to oxygen radical-mediated injury.

  5. Anti-Sonic hedgehog blocking antibody enhances killing effect of PBMCs on cervical carcinoma HeLa cells%Sonic hedgehog阻断抗体增强外周血单个核细胞抗宫颈癌HeLa细胞的作用

    Institute of Scientific and Technical Information of China (English)

    孙艳; 黄莉霞; 黄文浩; 黄斌

    2011-01-01

    AIM: To investigate the effect of anti - Sonic hedgehog( Shh ) blocking antibody on the killing effect of peripheral blood mononuclear cells( PBMCs ) on cervical carcinoma HeLa cells. METHODS: The expression levels of Shh and Shh signaling molecules in HeLa cells were detected by immunocytochemistry and RT - PCR. PBMCs from health peoples were isolated by the method of Ficoll density gradient centrifugation, and then co - cultured with HeLa cells in vitro. The expression of CD3, CD69 and CD71 was assayed by flew cytometry. The concentrations of IFN - 7, IL - 10 and IL - 4 in culture supernatants were detected by ELISA. The killing effect of PBMCs on HeLa cells was observed under microscope. RESULTS: Shh and Shh signaling molecules were expressed in HeLa cells. The level of Shh expression didn t change significantly in the 6th passage of HeLa cells. CD3 + cells were increased in the co - culture system. The expression of CD69 and CD 71, and the secretion of IFN - 7 were increased, while the secretion of IL - 10 was decreased in the co - culture system treated with anti - Shh blocking antibody. Anti - Shh blocking antibody has no effect on the secretion of IL -4. The killing effect of PBMCs on HeLa cells was strengthened by anti - Shh blocking antibody. CONCLUSION : Anti - Shh blocking antibody promotes the activation of PBMCs and enhances the killing effect of PBMCs on cervical carcinoma HeLa cells.%目的:探讨Sonic hedgehog(Shh)阻断抗体对外周血单个核细胞(PBMCs)抗宫颈癌HeLa细胞作用的影响.方法:免疫细胞化学技术和RT-PCR法检测Shh及其信号分子在HeLa 细胞中的表达;Ficoll密度梯度离心法分离正常人PBMCs,并与HeLa细胞建立共培养体系;于共培养体系中加入Shh阻断抗体,流式细胞术检测CD3、CD69和CD71分子表达;ELISA法检测细胞因子IFN-γ、IL-10和IL-4分泌;显微镜观察PBMCs 对HeLa细胞的杀伤.结果:HeLa细胞表达Shh及其信号分子,HeLa细胞传代6次对Shh的表达水

  6. The engineered thymidylate kinase (TMPK)/AZT enzyme-prodrug axis offers efficient bystander cell killing for suicide gene therapy of cancer.

    Science.gov (United States)

    Sato, Takeya; Neschadim, Anton; Lavie, Arnon; Yanagisawa, Teruyuki; Medin, Jeffrey A

    2013-01-01

    We previously described a novel suicide (or 'cell fate control') gene therapy enzyme/prodrug system based on an engineered variant of human thymidylate kinase (TMPK) that potentiates azidothymidine (AZT) activation. Delivery of a suicide gene sequence into tumors by lentiviral transduction embodies a cancer gene therapy that could employ bystander cell killing as a mechanism driving significant tumor regression in vivo. Here we present evidence of a significant bystander cell killing in vitro and in vivo mediated by the TMPK/AZT suicide gene axis that is reliant on the formation of functional gap-junctional intercellular communications (GJICs). Potentiation of AZT activation by the engineered TMPK expressed in the human prostate cancer cell line, PC-3, resulted in effective bystander killing of PC-3 cells lacking TMPK expression--an effect that could be blocked by the GJIC inhibitor, carbenoxolone. Although GJICs are mainly formed by connexins, a new family of GJIC molecules designated pannexins has been recently identified. PC-3 cells expressed both connexin43 (Cx43) and Pannexin1 (Panx1), but Panx1 expression predominated at the plasma membrane, whereas Cx43 expression was primarily localized to the cytosol. The contribution of bystander effects to the reduction of solid tumor xenografts established by the PC-3 cell line was evaluated in an animal model. We demonstrate the contribution of bystander cell killing to tumor regression in a xenograft model relying on the delivery of expression of the TMPK suicide gene into tumors via direct intratumoral injection of recombinant therapeutic lentivirus. Taken together, our data underscore that the TMPK/AZT enzyme-prodrug axis can be effectively utilized in suicide gene therapy of solid tumors, wherein significant tumor regression can be achieved via bystander effects mediated by GJICs.

  7. The engineered thymidylate kinase (TMPK/AZT enzyme-prodrug axis offers efficient bystander cell killing for suicide gene therapy of cancer.

    Directory of Open Access Journals (Sweden)

    Takeya Sato

    Full Text Available We previously described a novel suicide (or 'cell fate control' gene therapy enzyme/prodrug system based on an engineered variant of human thymidylate kinase (TMPK that potentiates azidothymidine (AZT activation. Delivery of a suicide gene sequence into tumors by lentiviral transduction embodies a cancer gene therapy that could employ bystander cell killing as a mechanism driving significant tumor regression in vivo. Here we present evidence of a significant bystander cell killing in vitro and in vivo mediated by the TMPK/AZT suicide gene axis that is reliant on the formation of functional gap-junctional intercellular communications (GJICs. Potentiation of AZT activation by the engineered TMPK expressed in the human prostate cancer cell line, PC-3, resulted in effective bystander killing of PC-3 cells lacking TMPK expression--an effect that could be blocked by the GJIC inhibitor, carbenoxolone. Although GJICs are mainly formed by connexins, a new family of GJIC molecules designated pannexins has been recently identified. PC-3 cells expressed both connexin43 (Cx43 and Pannexin1 (Panx1, but Panx1 expression predominated at the plasma membrane, whereas Cx43 expression was primarily localized to the cytosol. The contribution of bystander effects to the reduction of solid tumor xenografts established by the PC-3 cell line was evaluated in an animal model. We demonstrate the contribution of bystander cell killing to tumor regression in a xenograft model relying on the delivery of expression of the TMPK suicide gene into tumors via direct intratumoral injection of recombinant therapeutic lentivirus. Taken together, our data underscore that the TMPK/AZT enzyme-prodrug axis can be effectively utilized in suicide gene therapy of solid tumors, wherein significant tumor regression can be achieved via bystander effects mediated by GJICs.

  8. TH17 cells promote microbial killing and innate immune sensing of DNA via interleukin 26

    KAUST Repository

    Meller, Stephan

    2015-07-13

    Interleukin 17-producing helper T cells (TH 17 cells) have a major role in protection against infections and in mediating autoimmune diseases, yet the mechanisms involved are incompletely understood. We found that interleukin 26 (IL-26), a human TH17 cell-derived cytokine, is a cationic amphipathic protein that kills extracellular bacteria via membrane-pore formation. Furthermore, TH17 cell-derived IL-26 formed complexes with bacterial DNA and self-DNA released by dying bacteria and host cells. The resulting IL-26-DNA complexes triggered the production of type I interferon by plasmacytoid dendritic cells via activation of Toll-like receptor 9, but independently of the IL-26 receptor. These findings provide insights into the potent antimicrobial and proinflammatory function of TH17 cells by showing that IL-26 is a natural human antimicrobial that promotes immune sensing of bacterial and host cell death. © 2015 Nature America, Inc.

  9. Induction of Foot-and-Mouth Disease Virus-Specific Cytotoxic T Cell Killing by Vaccination

    DEFF Research Database (Denmark)

    Patch, J.R.; Pedersen, Lasse Eggers; Toka, F.N.;

    2011-01-01

    Foot-and-mouth disease (FMD) continues to be a significant threat to the health and economic value of livestock species. This acute infection is caused by the highly contagious FMD virus (FMDV), which infects cloven-hoofed animals including large and small ruminants and swine. Current vaccine...... strategies are all directed toward the induction of neutralizing antibody responses. However, the role of cytotoxic T lymphocytes (CTLs) has not received a great deal of attention, in part because of the technical difficulties associated with establishing a reliable assay of cell killing for this highly...... cytopathic virus. Here, we have used recombinant human adenovirus vectors as a means of delivering FMDV antigens in a T cell-directed vaccine in pigs. We tested the hypothesis that impaired processing of the FMDV capsid would enhance cytolytic activity, presumably by targeting all proteins for degradation...

  10. In Vitro Killing Effect of Moxifloxacin on Clinical Isolates of Stenotrophomonas maltophilia Resistant to Trimethoprim-Sulfamethoxazole

    OpenAIRE

    Giamarellos-Bourboulis, Evangelos J.; Karnesis, Lazaros; Galani, Irene; Giamarellou, Helen

    2002-01-01

    The time-kill effect of moxifloxacin on 20 genetically distinct isolates of Stenotrophomonas maltophilia resistant to trimethoprim-sulfamethoxazole was studied. The majority (80%) were killed by a concentration equivalent to four times the MIC; the MIC induced a transient decrease in bacterial counts at 4 h, followed by regrowth. No effect was detected in four isolates. These results merit further clinical consideration.

  11. UV-inactivated HSV-1 potently activates NK cell killing of leukemic cells.

    Science.gov (United States)

    Samudio, Ismael; Rezvani, Katayoun; Shaim, Hila; Hofs, Elyse; Ngom, Mor; Bu, Luke; Liu, Guoyu; Lee, Jason T C; Imren, Suzan; Lam, Vivian; Poon, Grace F T; Ghaedi, Maryam; Takei, Fumio; Humphries, Keith; Jia, William; Krystal, Gerald

    2016-05-26

    Herein we demonstrate that oncolytic herpes simplex virus-1 (HSV-1) potently activates human peripheral blood mononuclear cells (PBMCs) to lyse leukemic cell lines and primary acute myeloid leukemia samples, but not healthy allogeneic lymphocytes. Intriguingly, we found that UV light-inactivated HSV-1 (UV-HSV-1) is equally effective in promoting PBMC cytolysis of leukemic cells and is 1000- to 10 000-fold more potent at stimulating innate antileukemic responses than UV-inactivated cytomegalovirus, vesicular stomatitis virus, reovirus, or adenovirus. Mechanistically, UV-HSV-1 stimulates PBMC cytolysis of leukemic cells, partly via Toll-like receptor-2/protein kinase C/nuclear factor-κB signaling, and potently stimulates expression of CD69, degranulation, migration, and cytokine production in natural killer (NK) cells, suggesting that surface components of UV-HSV-1 directly activate NK cells. Importantly, UV-HSV-1 synergizes with interleukin-15 (IL-15) and IL-2 in inducing activation and cytolytic activity of NK cells. Additionally, UV-HSV-1 stimulates glycolysis and fatty acid oxidation-dependent oxygen consumption in NK cells, but only glycolysis is required for their enhanced antileukemic activity. Last, we demonstrate that T cell-depleted human PBMCs exposed to UV-HSV-1 provide a survival benefit in a murine xenograft model of human acute myeloid leukemia (AML). Taken together, our results support the preclinical development of UV-HSV-1 as an adjuvant, alone or in combination with IL-15, for allogeneic donor mononuclear cell infusions to treat AML.

  12. 树突状细胞的诱导培养及对胃癌细胞株MKN45的体外杀伤效应的初步研究%Preliminary studying on dendritic cell culture and its killing effect on gastric carcinoma cell line MKN 45

    Institute of Scientific and Technical Information of China (English)

    孙梯业; 颜伟; 孙冬丽; 刘全达; 段伟宏; 周宁新

    2010-01-01

    目的 探讨树突状细胞(DC)体外培养的方法及CpG ODN1826刺激DC对胃癌细胞MKN45的杀伤效应.方法 分离正常人外周血DC,培养至第5天,实验分为4组,A组[粒细胞-巨噬细胞集落刺激因子(GM-CSF)+白细胞介素-4(IL-4)]、B组[GM-CSF+IL-4+肿瘤坏死因子-α(TNF-α)]、C组(nonCpG ODN)和D组(CpG ODN 1826).自外周血单核细胞(PBMC)诱导分离DC,流式细胞仪检测DC表面标志,MTT法测定其刺激对同种异体淋巴细胞增殖的影响及对胃癌细胞的杀伤效应.结果 体外培养至第10天,D组和B组均出现大量典型的DC形态.倒置显微镜下见细胞呈树突状、伪足状突起.流式细胞仪检测,D组显著高表达共刺激分子(CD40、CD1a、CD80、CD86)和MHC-Ⅱ类分子,均分别高于其他各组(P<0.05).在体外能强烈刺激同种异体混合淋巴细胞的增殖,显著增强DC杀伤胃癌细胞MKN45的活性.结论 该培养方法可获得较高纯度典型的DC,同时CpG ODN可显著诱导人外周血DC分化成熟,增强DC对胃癌细胞MKN45的杀伤作用.%Objective To explore the cultivated methods of dendritic cells (DC) and the killing effect of DC stimulated by CpG ODN1826 on gastritic cancer cells MKN45 in vitro. Methods DC was induced from peripheral blood monocytes stimulated by A group ( GM- CSF + IL-4 ), B group ( GM- CSF + IL-4 + TNF- α), C group(nonCpG ODN) and D group( CpG ODN 1826). The surface markers of DC was analyzed via flow cytometry, and the abilities to stimulate proliferation of allogenic lymphocyte by DC and antitumor experiment were detected by MTT assay. Results On day 10, a majority of cells showed typical morphology of DC in D group and B group with visible branching-like and pseudopod-like structures under microscope. The results of flow cytometry showed that there are significantly high expressed co-stimulated molecules such as CD40, CD1a,CD80, CD86 and MHC- Ⅱ in D group compared to other experimental groups ( P < 0.05 ), which

  13. Recombinant human tumor necrosis factor alpha does not potentiate cell killing after photodynamic therapy with a silicon phthalocyanine in A431 human epidermoid carcinoma cells.

    Science.gov (United States)

    Azizuddin, K; Kalka, K; Chiu, S M; Ahmad, N; Mukhtar, H; Separovic, D

    2001-02-01

    Photodynamic therapy (PDT) is a novel cancer treatment utilizing a photosensitizer, visible light and oxygen. PDT with the silicon phthalocyanine Pc 4, a new photosensitizer, is highly effective in cancer cell destruction and tumor ablation. The mechanisms underlying cancer cell killing by PDT are not fully understood. Tumor necrosis factor alpha (TNF) is a multifunctional cytokine that has been implicated in photocytotoxicity. We asked whether recombinant human TNF (rhTNF) affects Pc 4-PDT cytotoxicity in A431 human epidermoid carcinoma cells. Co-treatment of A431 cells with various doses of Pc 4-PDT and a sub-lethal rhTNF dose led to a sub-additive reduction in cell survival. In addition, in the presence of Pc 4-PDT or rhTNF, caspase-3 activity and apoptosis were induced. The combined treatment, however, did not potentiate either caspase-3 activity or apoptosis. Similar to previous findings we observed that Pc 4-PDT initiated a time-dependent extracellular TNF accumulation. The data suggest that: a) PDT and rhTNF induce cancer cell killing through different mechanisms; and b) Pc 4-PDT-induced TNF production is a stress response that may not directly affect photocytotoxicity.

  14. Pseudomonas exotoxin antisense RNA selectively kills hepatitis B virus infected cells

    Institute of Scientific and Technical Information of China (English)

    Peter Hafkemeyer; Ulrich Brinkmann; Elizabeth Brinkmann; Ira Pastan; Hubert E Blum; Thomas F Baumert

    2008-01-01

    AIM: To present an approach for selectively killing retrovirus-infected cells that combines the toxicity of Pseudomonas exotoxin (PE) and the presence of reverse transcriptase (RT) in infected cells. METHODS: PE antisense toxin RNA has palindromic stem loops at its 5' and 3' ends enabling self-primed generation of cDNA in the presence of RT. The RT activity expressed in retrovirus-infected cells converts "antisense-toxin-RNA" into a lethal toxin gene exclusively in these cells. RESULTS: Using cotransfection studies with Peexpressing RNAs and β-gal expressing reporter plasmids, we show that, in HepG2 and HepG2. 2. 15 hepatomacells as well as in duck hepatitis B virus (DHBV) infected cells, HBV or DHBV-polymerase reverse transcribe a lethal cDNA copy of an antisense toxin RNA, which is composed of sequences complementary to a PE gene and eukaryotic transcription and translation signals. CONCLUSION: This finding may have important implications as a novel therapeutic strategy aimed at the elimination of HBV infection.

  15. Anthrax lethal toxin-mediated killing of human and murine dendritic cells impairs the adaptive immune response.

    Directory of Open Access Journals (Sweden)

    Abdelkrim Alileche

    2005-10-01

    Full Text Available Many pathogens have acquired strategies to combat the immune response. Bacillus anthracis interferes with host defenses by releasing anthrax lethal toxin (LT, which inactivates mitogen-activated protein kinase pathways, rendering dendritic cells (DCs and T lymphocytes nonresponsive to immune stimulation. However, these cell types are considered resistant to killing by LT. Here we show that LT kills primary human DCs in vitro, and murine DCs in vitro and in vivo. Kinetics of LT-mediated killing of murine DCs, as well as cell death pathways induced, were dependent upon genetic background: LT triggered rapid necrosis in BALB/c-derived DCs, and slow apoptosis in C57BL/6-derived DCs. This is consistent with rapid and slow killing of LT-injected BALB/c and C57BL/6 mice, respectively. We present evidence that anthrax LT impairs adaptive immunity by specifically targeting DCs. This may represent an immune-evasion strategy of the bacterium, and contribute to anthrax disease progression. We also established that genetic background determines whether apoptosis or necrosis is induced by LT. Finally, killing of C57BL/6-derived DCs by LT mirrors that of human DCs, suggesting that C57BL/6 DCs represent a better model system for human anthrax than the prototypical BALB/c macrophages.

  16. Effect of 50% human serum on the killing activity of micafungin against eight Candida species using time-kill methodology.

    Science.gov (United States)

    Földi, Richárd; Szilágyi, Judit; Kardos, Gábor; Berényi, Réka; Kovács, Renátó; Majoros, László

    2012-08-01

    Micafungin activity was determined against 24 wild-type clinical isolates and 5 American Type Culture Collection strains belonging to 8 Candida species in RPMI-1640 with and without 50% serum using broth microdilution and time-kill methodology. MIC values increased from 4- to 128-folds in 50% serum for all Candida species. Micafungin was not fungicidal against C. albicans, C. tropicalis, and against 2 of 3 C. metapsilosis at ≥0.25, 1, and 1 μg/mL, respectively, after 48 h with 50% serum, showing good fungistatic activity. Fungicidal activity at ≥2, 4, and 32 μg/mL was noticed against C. glabrata, C. inconspicua, and C. krusei isolates, respectively. Micafungin at 8-32 μg/mL showed fungistatic activity against C. parapsilosis and C. orthopsilosis. Serum decreased the in vitro activity of micafungin. With serum binding of echinocandins taken into account, safely fungistatic or fungicidal concentrations seem to require elevated doses against some Candida species, including C. parapsilosis, C. orthopsilosis, and C. krusei.

  17. Long-Term Effects of Targeted Killings by Unmanned Aerial Vehicles

    Science.gov (United States)

    2015-04-03

    Approved for public release; distribution is unlimited Long-Term Effects of Targeted Killings by Unmanned Aerial Vehicles A Monograph by MAJ...by Unmanned Aerial Vehicles 5a. CONTRACT NUMBER 5b. GRANT NUMBER 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) MAJ Jonathan. C. Nadler 5d...manifested in two long-term wars within Afghanistan and Iraq, simultaneously incited growth of an Unmanned Aerial Vehicle (UAV) industry dominated

  18. A whole-genome RNAi screen uncovers a novel role for human potassium channels in cell killing by the parasite Entamoeba histolytica.

    Science.gov (United States)

    Marie, Chelsea; Verkerke, Hans P; Theodorescu, Dan; Petri, William A

    2015-09-08

    The parasite Entamoeba histolytica kills human cells resulting in ulceration, inflammation and invasion of the colonic epithelium. We used the cytotoxic properties of ameba to select a genome-wide RNAi library to reveal novel host factors that control susceptibility to amebic killing. We identified 281 candidate susceptibility genes and bioinformatics analyses revealed that ion transporters were significantly enriched among susceptibility genes. Potassium (K(+)) channels were the most common transporter identified. Their importance was further supported by colon biopsy of humans with amebiasis that demonstrated suppressed K(+) channel expression. Inhibition of human K(+) channels by genetic silencing, pharmacologic inhibitors and with excess K(+) protected diverse cell types from E. histolytica-induced death. Contact with E. histolytica parasites triggered K(+) channel activation and K(+) efflux by intestinal epithelial cells, which preceded cell killing. Specific inhibition of Ca(2+)-dependent K(+) channels was highly effective in preventing amebic cytotoxicity in intestinal epithelial cells and macrophages. Blockade of K(+) efflux also inhibited caspase-1 activation, IL-1β secretion and pyroptotic death in THP-1 macrophages. We concluded that K(+) channels are host mediators of amebic cytotoxicity in multiple cells types and of inflammasome activation in macrophages.

  19. Fenugreek, a naturally occurring edible spice, kills MCF-7 human breast cancer cells via an apoptotic pathway.

    Science.gov (United States)

    Khoja, Kholoud K; Shaf, Gowhar; Hasan, Tarique N; Syed, Naveed Ahmed; Al-Khalifa, Abdrohman S; Al-Assaf, Abdullah H; Alshatwi, Ali A

    2011-01-01

    There is growing use of anticancer complementary and alternative medicines worldwide. Trigonella foenum graecum (Fenugreek) is traditionally applied to treat disorders such as diabetes, high cholesterol, wounds, inflammation, and gastrointestinal ailments. Fenugreek is also reported to have anticancer properties due to its active beneficial chemical constituents. The mechanism of action of several anticancer drugs is based on their ability to induce apoptosis. The objective of the study was to characterize the downstream apoptotic genes targeted by FCE in MCF-7 human immortalized breast cells. FCE effectively killed MCF-7 cells through induction of apoptosis,confirmed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and RT-PCR assays. When cells were exposed to 50 μg/mL FCE for 24 hours, 23.2% apoptotic cells resulted, while a 48-hour exposure to 50 μg/mL caused 73.8% apoptosis. This was associated with increased expression of Caspase 3, 8, 9, p53, Fas, FADD, Bax and Bak in a time-and dose-dependent manner, as determined by real- time quantitative PCR. In summary, the induction of apoptosis by FCE is effected by its ability to increase the expression of pro-apoptotic genes and the spice holds promise for consideration in complementary therapy for breast cancer patients.

  20. A partner monoclonal antibody to Moab 730 kills 100% of DU145 and PC3 androgen-independent cancer cells

    Indian Academy of Sciences (India)

    Hemant Kumar Vyas; Rahul Pal; Nirmal K Lohiya; G P Talwar

    2009-12-01

    A number of therapeutic options are available for patients with prostate carcinoma till the time that the tumour is hormone dependent. However, no fully effective therapy is available for the treatment of androgen-independent prostate carcinomas. Antibodies directed at epitopes unique to or overexpressed on the cancer cells could be of therapeutic utility. A monoclonal antibody (Moab) 2C4 has been generated, which binds with cells of two androgenindependent prostate cancers, DU145 and PC3, and does not bind to peripheral blood leukocytes (PBLs) of healthy donors. This antibody, along with the previously developed Moab 730, kills 100% of both DU145 and PC3 cells in the presence of complement and does not have a deleterious effect on PBLs of healthy males. The anti-tumour action of the two antibodies prevents the establishment of DU145 cell tumour in nude mice in vivo. Moab 2C4 in combination with 730 has potential for use as therapy for androgen-independent cancers.

  1. Differential timing of antibody-mediated phagocytosis and cell-free killing of invasive African Salmonella allows immune evasion.

    Science.gov (United States)

    Siggins, Matthew K; O'Shaughnessy, Colette M; Pravin, John; Cunningham, Adam F; Henderson, Ian R; Drayson, Mark T; MacLennan, Calman A

    2014-04-01

    Nontyphoidal Salmonellae commonly cause fatal bacteraemia in African children lacking anti-Salmonella antibodies. These are facultative intracellular bacteria capable of cell-free and intracellular survival within macrophages. To better understand the relationship between extracellular and intracellular infection in blood and general mechanisms of Ab-related protection against Salmonella, we used human blood and sera to measure kinetics of Ab and complement deposition, serum-mediated bactericidal killing and phagocytosis of invasive African Salmonella enterica serovar Typhimurium D23580. Binding of antibodies peaked by 30 s, but C3 deposition lagged behind, peaking after 2-4 min. C5b-9 deposition was undetectable until between 2 and 6 min and peaked after 10 min, after which time an increase in serum-mediated killing occurred. In contrast, intracellular, opsonized Salmonellae were readily detectable within 5 min. By 10 min, around half of monocytes and most neutrophils contained bacteria. The same kinetics of serum-mediated killing and phagocytosis were observed with S. enterica Typhimurium laboratory strain SL1344, and the S. enterica Enteritidis African invasive isolate D24954 and laboratory strain PT4. The differential kinetics between cell-free killing and phagocytosis of invasive nontyphoidal Salmonella allows these bacteria to escape the blood and establish intracellular infection before they are killed by the membrane attack complex.

  2. PD-L1 Expression on Retrovirus-Infected Cells Mediates Immune Escape from CD8+ T Cell Killing.

    Directory of Open Access Journals (Sweden)

    Ilseyar Akhmetzyanova

    2015-10-01

    Full Text Available Cytotoxic CD8+ T Lymphocytes (CTL efficiently control acute virus infections but can become exhausted when a chronic infection develops. Signaling of the inhibitory receptor PD-1 is an important mechanism for the development of virus-specific CD8+ T cell dysfunction. However, it has recently been shown that during the initial phase of infection virus-specific CD8+ T cells express high levels of PD-1, but are fully competent in producing cytokines and killing virus-infected target cells. To better understand the role of the PD-1 signaling pathway in CD8+ T cell cytotoxicity during acute viral infections we analyzed the expression of the ligand on retrovirus-infected cells targeted by CTLs. We observed increased levels of PD-L1 expression after infection of cells with the murine Friend retrovirus (FV or with HIV. In FV infected mice, virus-specific CTLs efficiently eliminated infected target cells that expressed low levels of PD-L1 or that were deficient for PD-L1 but the population of PD-L1high cells escaped elimination and formed a reservoir for chronic FV replication. Infected cells with high PD-L1 expression mediated a negative feedback on CD8+ T cells and inhibited their expansion and cytotoxic functions. These findings provide evidence for a novel immune escape mechanism during acute retroviral infection based on PD-L1 expression levels on virus infected target cells.

  3. Enhanced effect of CD8++ T cells activated by tumor lysate -pulsed DCs on killing autologous tumor cells%通过肿瘤致敏的DCs活化的CD8+T细胞可有效地杀死肿瘤细胞

    Institute of Scientific and Technical Information of China (English)

    唐小龙; 江振友; 蔡淑玉

    2008-01-01

    AIM:To evaluate the ability of dendritic cells (DCs) loaded with tumor lysate to initiate cell mediated immune responses by stimulating naive T cells, and the efficiency of activated T cells to kill autologous tumor cells in vitro. METHODS: The peripheral blood lymphocytes and monocytes were obtained from the advanced renal cell carcinoma patient by eonglutination method. The immature dendritic cells were generated in the presence of interleukin -4(IL-4) and granulocyte/macrophage colony-stimulating factor (GM-CSF) from monocytes of healthy individuals.These cells were pulsed with tumor lysate or not. Induction of tumor-specific cytotoxic T lymphocytes(CTLs) response by mature dendritic cells (mDCs) was evaluated by the CD95(Fas) expression assay through FCM and the cytotoxic assay a gninst autolognns human tumor cells. RESULTS: Human immature dendritic cells and T cells obtained from healthy donors were stimulated with tumor- pulsed dendritic cells. The immature dendritic cells were applied to the cytotoxicity assay a gainst target autologons tumor cells. The CD95 (Fas) expression, IFN-γ, and TNF -α secreted by the CTLs in tumor lysate-plused DC group were higher than those of other groups. The capacity of the CTLs to kill autolognns tumor cells was significantly different(P<0. 05). Antigen-specific DCs vaccine can induce T cells activation and proliferation, thus we can obtain higher proportion of tumor specific cytotoxic T cells(CTLs), and enhance the CTLs to secret IFN-γ and TNF-α. CONCLUSION: Our results indicate that monocyte-derived human dendritic cells pulsed with tumor lysate could in duce the specific antitumor effect against autologons tumors. This in vitro model offers a new and simple approach to the development of DC + CTL - based immunotherapy.%目的:探索肿瘤裂解物负载的DCs诱导活化的初始T细胞介导细胞免疫及活化的T细胞杀死肿瘤细胞的能力.方法:应用黏附法分离外周血中的淋巴细胞

  4. 腺病毒介导的KDR-HSV-tk系统对血管内皮细胞的杀伤作用及旁观者效应%The killing and bystander effect of KDR-HSV-tk system mediated by adenovirus on vascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    朱欠元; 曾友根; 张愈延; 李宝金

    2015-01-01

    目的:研究腺病毒介导的KDR-HSV-tk系统对血管内皮细胞的杀伤作用及旁杀效应。方法体外构建受 KDR启动子和巨细胞病毒( CMV)启动子调控并可表达HSV-tk基因的AdKDR-HSV-tk和AdCMV-HSV-tk,经293细胞包装、扩增后,分别感染表达KDR的人脐静脉血管内皮细胞(HUVEC)和不表达KDR的人鼻咽癌细胞CNE-2,建立HUVEC/tk及CNE-2/tk,观察丙氧鸟苷(GCV)处理后 HUVEC 杀伤率及旁观者效应。结果受感染的HUVEC和CNE-2细胞中均有绿色荧光蛋白的表达。在感染复数( MOI)为100的条件下,GCV浓度由0增至50 mg/L 时含 AdKDR-HSV-tk的HUVEC和CNE-2细胞存活率从100%分别降至(23.1±3.9)%和(69.8±3.1)%(P<0.01);感染AdCMV-HSV-tk的HUVEC和 CNE-2细胞存活率从100%分别降至(13.9±2.9)%和(15.7±5.2)%(P<0.05);HUVEC/tk细胞对GCV的敏感性明显高于亲代HUVEC细胞, HUVEC+HUVEC/tk混合细胞的生存率随HUVEC/tk比例的增加而下降,说明AdKDR-HSV-tk/GCV系统不但能杀伤转染tk基因的HUVEC,也能杀伤未导入转染tk基因的HUVEC细胞,存在明显的旁观者效应。结论腺病毒介导的KDR-HSV-tk/GCV系统对血管内皮细胞具有特异的杀伤作用及旁观者效应,为抗肿瘤血管靶向治疗奠定了基础。%Objective To study the killing and bystander effect of KDR-HSV-tk system study of adenovirus mediated vascular endo-thelial cells.Methods AdKDR-HSV-tk and AdCMV-HSV-tk controlled by KDR promoter and cytomegalovirus (CMV) promoter were con-structed in vitro,and infected human umbilical vein endothelial cells (HUVEC) expressing KDR and human nasopharyngeal carcinoma cell line CNE-2 without KDR to establish HUVEC/tk and CNE-2/tk respectively .The killing rate and bystander effect were observed after ganci-clovir(GCV) treatment.Results The infected HUVEC and CNE-2 cells were

  5. PNP-TK融合自杀基因及PNP单自杀基因对肝癌细胞杀伤效率%Cytotoxic killing effects of a PNP-TK chimeric suicide gene system and a single PNP suicide gene system on hepatoma cells

    Institute of Scientific and Technical Information of China (English)

    周俊立; 蔡晓坤; 易学锋; 王德全; 姚振江; 周卫平

    2009-01-01

    目的 分析PNP-TK融合自杀基因系统及PNP单自杀基因系统对肝癌细胞的杀伤作用及二者对肝癌细胞潜在的杀伤机制. 方法构建融合基因表达载体pcDNA3.0/PNP-TK,经酶切、PCR及测序鉴定重组体,G418筛选获得稳定转染了pcDNA3.0/PNP-TK的HepG2细胞克隆.RT-PCR和Western Blotting检测融合基因在HepG2细胞中的表达.台盼兰排斥法检测细胞的生长曲线,MTT法检测细胞对相应前药的敏感性及分别在一种和两种前药作用下所导致的旁观者效应.结果 融合基因片段PNP-TK正确插入了pcDNA3.0载体中,pcDNA3.0/PNP-TK在肝癌细胞株HepG2中实现了表达.抗性细胞克隆对相应的前药十分敏感.在两种前药的联合作用下,pcDNA3.0/PNP-TK所导致的旁观者效应明显强于只给予MeP-dR一种前药以及pcDNA3.0/PNP单基因系统所致的旁观者效应.结论 具有双自杀基因功能的表达载体pcDNA3.0/PNP-TK对肝癌细胞的杀伤效果优于PNP单自杀基因系统.%Objective To analyze the killing effects of a PNP-TK chimeras gene system on the basis of PNP suicide gene and a single PNP suicide gene system on hepatoma cells and explore the potential mechanism of their killing effects. Methods A vector harboring a chimeric gene,pcDNA3.0/PNP-TK was constructed. The recombinant was identified by recombinant enzyme, PCR and sequencing. Then it was transfected into HepG2 cells by liposome-mediated method. The G-418 resistant cell clone with the stable transfection of pcDNA3.0/PNP-TK was developed. The expression of PNP-TK gene was detected by RT-PCR and Western blotting method. The cellular growth curves were determined by trypan blue exclusion. The sensitivity of cell clones to their respective prodrugs and the bystander effects they resulted in were also assayed by MTT method. Results The PNP-TK fusion gene was inserted into pcDNA3.0 correctly,and the stable expression of PNP-TK gene in HepG2 cells was positive. Two clones stably expressing

  6. T Cell Receptor Vβ Staining Identifies the Malignant Clone in Adult T cell Leukemia and Reveals Killing of Leukemia Cells by Autologous CD8+ T cells.

    Science.gov (United States)

    Rowan, Aileen G; Witkover, Aviva; Melamed, Anat; Tanaka, Yuetsu; Cook, Lucy B M; Fields, Paul; Taylor, Graham P; Bangham, Charles R M

    2016-11-01

    There is growing evidence that CD8+ cytotoxic T lymphocyte (CTL) responses can contribute to long-term remission of many malignancies. The etiological agent of adult T-cell leukemia/lymphoma (ATL), human T lymphotropic virus type-1 (HTLV-1), contains highly immunogenic CTL epitopes, but ATL patients typically have low frequencies of cytokine-producing HTLV-1-specific CD8+ cells in the circulation. It remains unclear whether patients with ATL possess CTLs that can kill the malignant HTLV-1 infected clone. Here we used flow cytometric staining of TCRVβ and cell adhesion molecule-1 (CADM1) to identify monoclonal populations of HTLV-1-infected T cells in the peripheral blood of patients with ATL. Thus, we quantified the rate of CD8+-mediated killing of the putative malignant clone in ex vivo blood samples. We observed that CD8+ cells from ATL patients were unable to lyse autologous ATL clones when tested directly ex vivo. However, short in vitro culture restored the ability of CD8+ cells to kill ex vivo ATL clones in some donors. The capacity of CD8+ cells to lyse HTLV-1 infected cells which expressed the viral sense strand gene products was significantly enhanced after in vitro culture, and donors with an ATL clone that expressed the HTLV-1 Tax gene were most likely to make a detectable lytic CD8+ response to the ATL cells. We conclude that some patients with ATL possess functional tumour-specific CTLs which could be exploited to contribute to control of the disease.

  7. Analyses of CD20 Monoclonal Antibody–Mediated Tumor Cell Killing Mechanisms: Rational Design of Dosing Strategies

    Science.gov (United States)

    Lindorfer, Margaret A.

    2014-01-01

    Since approval of rituximab for treatment of B cell non-Hodgkin lymphoma, development of monoclonal antibodies (mAbs) for cancer treatment and elucidation of their cytotoxic mechanisms have been subject to intense investigations. Compelling evidence indicates that rituximab and another CD20 mAb, ofatumumab, must use the body’s cellular and humoral immune effector functions to kill malignant cells. Other U.S. Food and Drug Administration–approved mAbs, including obinutuzumab, cetuximab, and trastuzumab, require, in part, these effector mechanisms to eliminate tumor cells. Although gram quantities of mAbs can be administered to patients, our investigations of CD20 mAb-based therapies for chronic lymphocytic leukemia (CLL), including correlative measurements in clinical trials and studies with primary cells and cell lines, indicate that effector mechanisms necessary for mAb activity can be saturated or exhausted if tumor burdens are high, thus substantially compromising the efficacy of high-dose mAb therapy. Under these conditions, another reaction (trogocytosis) predominates in which bound CD20 mAb and CD20 are removed from targeted cells by effector cells that express Fcγ receptors, thereby allowing malignant cells to escape unharmed and continue to promote disease pathology. To address this problem, we propose that a low-dose strategy, based on administering 30–50 mg of CD20 mAb three times per week, may be far more effective for CLL than standard dosing because it will minimize effector function saturation and reduce trogocytosis. This approach may have general applicability to other mAbs that use immune effector functions, and could be formulated into a subcutaneous treatment strategy that would be more accessible and possibly more efficacious for patients. PMID:24944188

  8. Cryptococcus neoformans modulates extracellular killing by neutrophils

    Directory of Open Access Journals (Sweden)

    Asfia eQureshi

    2011-09-01

    Full Text Available We recently established a key role for host sphingomyelin synthase (SMS in the regulation of the killing activity of neutrophils against Cryptococcus neoformans. In this work, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and NK cells (Tgε26 mice. To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in vitro and in vivo. We show that unlike C. albicans, neither the presence nor the capsule size of C. neoformans cells have any effect on neutrophil viability. Interestingly, melanized C. neoformans cells totally abrogated the killing activity of neutrophils. Next, we monitored how exposure of neutrophils to C. neoformans cells would interfere with any further killing activity of the medium and found that pre-incubation with live but not heat-killed fungal cells significantly inhibits further killing activity of the medium. We next studied whether activation of SMS at the site of C. neoformans infection is dependent on T and NK cells. Using matrix-assisted laser desorption-ionization (MALDI tissue imaging in infected lung we found that similarly to previous observations in the isogenic wild type CBA/J mice, SM 16:0 levels are significantly elevated at the site of infection in mice lacking T and NK cells but only at early time points. This study highlights that C. neoformans may negatively regulate the killing activity of neutrophils and that SMS activation in neutrophils appears to be partially independent of T and/or NK cells.

  9. Cryptococcus neoformans modulates extracellular killing by neutrophils.

    Science.gov (United States)

    Qureshi, Asfia; Grey, Angus; Rose, Kristie L; Schey, Kevin L; Del Poeta, Maurizio

    2011-01-01

    We recently established a key role for host sphingomyelin synthase (SMS) in regulating the killing activity of neutrophils against Cryptococcus neoformans. In this paper, we studied the effect of C. neoformans on the killing activity of neutrophils and whether SMS would still be a player against C. neoformans in immunocompromised mice lacking T and natural killer (NK) cells (Tgε26 mice). To this end, we analyzed whether C. neoformans would have any effect on neutrophil survival and killing in vitro and in vivo. We show that unlike Candida albicans, neither the presence nor the capsule size of C. neoformans cells have any effect on neutrophil viability. Interestingly, melanized C. neoformans cells totally abrogated the killing activity of neutrophils. We monitored how exposure of neutrophils to C. neoformans cells would interfere with any further killing activity of the conditioned medium and found that pre-incubation with live but not "heat-killed" fungal cells significantly inhibits further killing activity of the medium. We then studied whether activation of SMS at the site of C. neoformans infection is dependent on T and NK cells. Using matrix-assisted laser desorption-ionization tissue imaging in infected lung we found that similar to previous observations in the isogenic wild-type CBA/J mice, SM 16:0 levels are significantly elevated at the site of infection in mice lacking T and NK cells, but only at early time points. This study highlights that C. neoformans may negatively regulate the killing activity of neutrophils and that SMS activation in neutrophils appears to be partially independent of T and/or NK cells.

  10. Comparative analysis of cell killing and autosomal mutation in mouse kidney epithelium exposed to 1 GeV protons in vitro or in vivo.

    Science.gov (United States)

    Kronenberg, Amy; Gauny, Stacey; Kwoh, Ely; Grossi, Gianfranco; Dan, Cristian; Grygoryev, Dmytro; Lasarev, Michael; Turker, Mitchell S

    2013-05-01

    Human exposure to high-energy protons occurs in space flight scenarios or, where necessary, during radiotherapy for cancer or benign conditions. However, few studies have assessed the mutagenic effectiveness of high-energy protons, which may contribute to cancer risk. Mutations cause cancer and most cancer-associated mutations occur at autosomal loci. This study addresses the cytotoxic and mutagenic effects of 1 GeV protons in mouse kidney epithelium. Mutant fractions were measured for an endogenous autosomal locus (Aprt) that detects all types of mutagenic events. Results for kidneys irradiated in vivo are compared with the results for kidney cells from the same strain exposed in vitro. The results demonstrate dose-dependent cell killing in vitro and for cells explanted 3-4 months postirradiation in vivo. Incubation in vivo for longer periods (8-9 months) further attenuates proton-induced cell killing. Protons are mutagenic to cells in vitro and for in vivo irradiated kidneys. The dose-response for Aprt mutation is curvilinear after in vitro or in vivo exposure, bending upward at the higher doses. While the absolute mutant fractions are higher in vivo, the fold-increase over background is similar for both in vitro and in situ exposures. Results are also presented for a limited study on the effect of dose fractionation on the induction of Aprt mutations in kidney epithelial cells. Dose-fractionation reduces the fraction of proton-induced Aprt mutants in vitro and in vivo and also results in less cell killing. Taken together, the mutation burden in the epithelium is slightly reduced by dose-fractionation. Autosomal mutations accumulated during clinical exposure to high-energy protons may contribute to the risk of treatment-associated neoplasms, thereby highlighting the need for rigorous treatment planning to reduce the dose to normal tissues. For low dose exposures that occur during most space flight scenarios, the mutagenic effects of protons appear to be modest.

  11. Comparative analysis of cell killing and autosomal mutation in mouse kidney epithelium exposed to 1 GeV/nucleon iron ions in vitro or in situ.

    Science.gov (United States)

    Kronenberg, Amy; Gauny, Stacey; Kwoh, Ely; Connolly, Lanelle; Dan, Cristian; Lasarev, Michael; Turker, Mitchell S

    2009-11-01

    Astronauts receive exposures to high-energy heavy ions from galactic cosmic radiation. Although high-energy heavy ions are mutagenic and carcinogenic, their mutagenic potency in epithelial cells, where most human cancers develop, is poorly understood. Mutations are a critical component of human cancer, and mutations involving autosomal loci predominate. This study addresses the cytotoxic and mutagenic effects of 1 GeV/nucleon iron ions in mouse kidney epithelium. Mutant fractions were measured for an endogenous autosomal locus (Aprt) that detects all types of mutagenic events contributing to human cancer. Results for kidneys irradiated in situ are compared with results for kidney cells from the same strain exposed in vitro. The results demonstrate dose-dependent cell killing in vitro and for cells explanted 3-4 months postirradiation in situ, but in situ exposures were less likely to result in cell death than in vitro exposures. Prolonged incubation in situ (8-9 months) further attenuated cell killing at lower doses. Iron ions were mutagenic to cells in vitro and for irradiated kidneys. No sparing was seen for mutant frequency with a long incubation period in situ. In addition, the degree of mutation induction (relative increase over background) was similar for cells exposed in vitro or in situ. We speculate that the latent effects of iron-ion exposure contribute to the maintenance of an elevated mutation burden in an epithelial tissue.

  12. In Vitro Killing Effect of Moxifloxacin on Clinical Isolates of Stenotrophomonas maltophilia Resistant to Trimethoprim-Sulfamethoxazole

    Science.gov (United States)

    Giamarellos-Bourboulis, Evangelos J.; Karnesis, Lazaros; Galani, Irene; Giamarellou, Helen

    2002-01-01

    The time-kill effect of moxifloxacin on 20 genetically distinct isolates of Stenotrophomonas maltophilia resistant to trimethoprim-sulfamethoxazole was studied. The majority (80%) were killed by a concentration equivalent to four times the MIC; the MIC induced a transient decrease in bacterial counts at 4 h, followed by regrowth. No effect was detected in four isolates. These results merit further clinical consideration. PMID:12435710

  13. Cytolethal distending toxin B as a cell-killing component of tumor-targeted anthrax toxin fusion proteins.

    Science.gov (United States)

    Bachran, C; Hasikova, R; Leysath, C E; Sastalla, I; Zhang, Y; Fattah, R J; Liu, S; Leppla, S H

    2014-01-16

    Cytolethal distending toxin (Cdt) is produced by Gram-negative bacteria of several species. It is composed of three subunits, CdtA, CdtB, and CdtC, with CdtB being the catalytic subunit. We fused CdtB from Haemophilus ducreyi to the N-terminal 255 amino acids of Bacillus anthracis toxin lethal factor (LFn) to design a novel, potentially potent antitumor drug. As a result of this fusion, CdtB was transported into the cytosol of targeted cells via the efficient delivery mechanism of anthrax toxin. The fusion protein efficiently killed various human tumor cell lines by first inducing a complete cell cycle arrest in the G2/M phase, followed by induction of apoptosis. The fusion protein showed very low toxicity in mouse experiments and impressive antitumor effects in a Lewis Lung carcinoma model, with a 90% cure rate. This study demonstrates that efficient drug delivery by a modified anthrax toxin system combined with the enzymatic activity of CdtB has great potential as anticancer treatment and should be considered for the development of novel anticancer drugs.

  14. Nuclear estrogen receptor targeted photodynamic therapy: selective uptake and killing of MCF-7 breast cancer cells by a C17alpha-alkynylestradiol-porphyrin conjugate.

    Science.gov (United States)

    Swamy, Narasimha; Purohit, Ajay; Fernandez-Gacio, Ana; Jones, Graham B; Ray, Rahul

    2006-10-15

    We hypothesized that over-expression of estrogen receptor (ER) in hormone-sensitive breast cancer could be harnessed synergistically with the tumor-migrating effect of porphyrins to selectively deliver estrogen-porphyrin conjugates into breast tumor cells, and preferentially kill the tumor cells upon exposure to red light. In the present work we synthesized four (4) conjugates of C17-alpha-alkynylestradiol and chlorin e6-dimethyl ester with varying tether lengths, and showed that all these conjugates specifically bound to recombinant ER alpha. In a cellular uptake assay with ER-positive MCF-7 and ER-negative MDA-MB 231 human breast cancer cell-lines, we observed that one such conjugate (E17-POR, XIV) was selectively taken up in a dose-dependent and saturable manner by MCF-7 cells, but not by MDA-MB 231 cells. Furthermore, MCF-7 cells, but not MDA-MB 231 cells, were selectively and efficiently killed by exposure to red light after incubation with E17-POR. Therefore, the combination approach, including drug and process modalities has the potential to be applied clinically for hormone-sensitive cancers in organs where ER is significantly expressed. This could potentially be carried out either as monotherapy involving a photo-induced selective destruction of tumor cells and/or adjuvant therapy in post-surgical treatment for the destruction of residual cancer cells in tissues surrounding the tumor.

  15. Enhanced killing of SCC17B human head and neck squamous cell carcinoma cells after photodynamic therapy plus fenretinide via the de novo sphingolipid biosynthesis pathway and apoptosis.

    Science.gov (United States)

    Boppana, Nithin B; Stochaj, Ursula; Kodiha, Mohamed; Bielawska, Alicja; Bielawski, Jacek; Pierce, Jason S; Korbelik, Mladen; Separovic, Duska

    2015-05-01

    Because photodynamic therapy (PDT) alone is not always effective as an anticancer treatment, PDT is combined with other anticancer agents for improved efficacy. The clinically-relevant fenretinide [N-(4-hydroxyphenyl) retinamide; 4HPR], was combined with the silicon phthalocyanine photosensitizer Pc4-mediated PDT to test for their potential to enhance killing of SCC17B cells, a clinically-relevant model of human head and neck squamous cell carcinoma. Because each of these treatments induces apoptosis and regulates the de novo sphingolipid (SL) biosynthesis pathway, the role of ceramide synthase, the pathway-associated enzyme, in PDT+4HPR-induced apoptotic cell death was determined using the ceramide synthase inhibitor fumonisin B1 (FB). PDT+4HPR enhanced loss of clonogenicity. zVAD-fmk, a pan-caspase inhibitor, and FB, protected cells from death post-PDT+4HPR. In contrast, the anti-apoptotic protein Bcl2 inhibitor ABT199 enhanced cell killing after PDT+4HPR. Combining PDT with 4HPR led to FB-sensitive, enhanced Bax associated with mitochondria and cytochrome c redistribution. Mass spectrometry data showed that the accumulation of C16-dihydroceramide, a precursor of ceramide in the de novo SL biosynthesis pathway, was enhanced after PDT+4HPR. Using quantitative confocal microscopy, we found that PDT+4HPR enhanced dihydroceramide/ceramide accumulation in the ER, which was inhibited by FB. The results suggest that SCC17B cells are sensitized to PDT by 4HPR via the de novo SL biosynthesis pathway and apoptosis, and imply potential clinical relevance of the combination for cancer treatment.

  16. Zika Kills Vital Nervous System Cells in Adult Mice, Study Finds

    Science.gov (United States)

    ... effects of the Zika-related changes in neural stem cell numbers in mice. However, research with animals often doesn't translate to similar findings in humans. SOURCE: Cell Stem Cell , news release, Aug. 18, 2016 HealthDay Copyright ( ...

  17. Integrating Enzymatic Self-Assembly and Mitochondria Targeting for Selectively Killing Cancer Cells without Acquired Drug Resistance.

    Science.gov (United States)

    Wang, Huaimin; Feng, Zhaoqianqi; Wang, Youzhi; Zhou, Rong; Yang, Zhimou; Xu, Bing

    2016-12-14

    Targeting organelles by modulating the redox potential of mitochondria is a promising approach to kill cancer cells that minimizes acquired drug resistance. However, it lacks selectivity because mitochondria perform essential functions for (almost) all cells. We show that enzyme-instructed self-assembly (EISA), a bioinspired molecular process, selectively generates the assemblies of redox modulators (e.g., triphenyl phosphinium (TPP)) in the pericellular space of cancer cells for uptake, which allows selectively targeting the mitochondria of cancer cells. The attachment of TPP to a pair of enantiomeric, phosphorylated tetrapeptides produces the precursors (L-1P or D-1P) that form oligomers. Upon dephosphorylation catalyzed by ectophosphatases (e.g., alkaline phosphatase (ALP)) overexpressed on cancer cells (e.g., Saos2), the oligomers self-assemble to form nanoscale assemblies only on the surface of the cancer cells. The cancer cells thus uptake these assemblies of TPP via endocytosis, mainly via a caveolae/raft-dependent pathway. Inside the cells, the assemblies of TPP-peptide conjugates escape from the lysosome, induce dysfunction of mitochondria to release cytochrome c, and result in cell death, while the controls (i.e., omitting TPP motif, inhibiting ALP, or removing phosphate trigger) hardly kill the Saos2 cells. Most importantly, the repeated stimulation of the cancers by the precursors, unexpectedly, sensitizes the cancer cells to the precursors. As the first example of the integration of subcellular targeting with cell targeting, this study validates the spatial control of the assemblies of nonspecific cytotoxic agents by EISA as a promising molecular process for selectively killing cancer cells without inducing acquired drug resistance.

  18. EGL-1 BH3 mutants reveal the importance of protein levels and target affinity for cell-killing potency.

    Science.gov (United States)

    Lee, E F; Chen, L; Yang, H; Colman, P M; Huang, D C S; Fairlie, W D

    2008-10-01

    Studies of the cell death pathway in the nematode Caenorhabditis elegans provided the first evidence of the evolutionary conservation of apoptosis signalling. Here we show that the worm Bcl-2 homology domain-3 (BH3)-only protein EGL-1 binds mammalian pro-survival proteins very poorly, but can be converted into a high-affinity ligand for Bcl-2 and Bcl-x(L) by subtle mutation of the cysteine residue at position 62 within the BH3 domain. A 100-fold increase in affinity was observed following a single atom change (cysteine to serine substitution), and a further 10-fold increase by replacement with glycine. The low affinity of wild-type EGL-1 for mammalian pro-survival proteins and its poor expression correlates with its weak killing activity in mammalian cells whereas the high-affinity C62G mutant is a very potent killer of cells lacking Mcl-1. Cell killing by the C62S mutant with intermediate affinity only occurs when this EGL-1 BH3 domain is placed in a more stable context, namely that of Bim(S), which allows higher expression, though the kinetics of cell death now vary depending on whether Mcl-1 is neutralized by Noxa or genetically deleted. These results demonstrate how levels of BH3-only proteins, target affinity and the spectrum of neutralization of pro-survival proteins all contribute to killing activity.

  19. The Killing

    DEFF Research Database (Denmark)

    Agger, Gunhild

    2013-01-01

    This article tracks the uncanny locations of The Killing (2007–2012), relating them to place, space and atmosphere, putting bits and pieces from the topographic puzzle together with cues from the symbolic space in order to see how they fit into the overall pattern of Nordic Noir. In The Killing......, the abstract level of space and atmosphere meets the concrete level of place, both influencing the notion of location. This meeting, I suggest, has contributed towards the simultaneous domestic and international appeal of The Killing....

  20. Killing effect of CIKs on pancreatic cancer cells enhanced by DCs loaded with K-ras mutant peptide%K-ras突变多肽负载的DC细胞增强CIK细胞对胰腺癌细胞的杀伤作用

    Institute of Scientific and Technical Information of China (English)

    窦春鹏; 李奎武; 谭广

    2015-01-01

    Objective:To observe the killing effect of the cytokine induced killer cells (CIKs) atfer co-culture with dendritic cells (DCs) harboring K-ras (12-Val) mutant peptide on pancreatic cancer PANC-1 cells. Methods:DCs and CIKs were induced and enriched from peripheral blood of healthy donors, respectively. DCs were loaded with the K-ras mutant epitope peptide (K-ras-DCs), and CIKs were co-cultured with un-loaded DCs or K-ras-DCs to obtain the DC-CIKs and K-ras-DC-CIKs, respectively. hTe proliferative activities between CIKs and K-ras-DC-CIKs were compared, the difference in immunophenotype between DCs and K-ras-DCs as well as between CIKs and K-ras-DC-CIKs were analyzed, the IFN-γand IL-12 levels in the culture supernatants from CIKs, DC-CIKs and K-ras-DC-CIKs were measured, and the killing abilities of CIKs, DC-CIKs and K-ras-DC-CIKs on PANC-1 cells in vitro were determined. Results:The proliferative ability of K-ras-DC-CIKs was significantly greater than that of the untreated CIKs (P Conclusion:K-ras mutant peptide can promote DCs maturation, and DCs harboring K-ras mutant peptide can increase the proliferation of CIKs and killing effect on pancreatic cancer cells.%目的:观察经K-ras(12-Val)突变多肽负载的树突状细胞(DC)与细胞因子诱导的杀伤细胞(CIK)共培养以后对胰腺癌PANC-1细胞的杀伤作用。  方法:取健康人外周血体外诱导分别扩增出DC和CIK;用K-ras突变体抗原表位肽负载DC(K-ras-DC),将单纯DC或K-ras-DC与CIK共培养,获得DC-CIK或K-ras-DC-CIK。比较CIK与K-ras-DC-CIK的增殖活性;分别分析DC与K-ras-DC以及CIK与K-ras-DC-CIK的免疫表型差异;检测CIK、DC-CIK、K-ras-DC-CIK上清液中IFN-γ、IL-12的水平;检测K-ras-DC-CIK、DC-CIK、CIK对PANC-1细胞的体外杀伤力。  结果:K-ras-DC-CIK的增殖能力明显强于单纯CIK(P  结论:K-ras突变多肽负载后能促进DC的成熟,负载K-ras突变多肽后的DC能增加CIK

  1. The Killing Effect on EJ Bladder Cancer Cells by Combination of Radiation-responsive Promoter Conjoined CD Gene and Ionizing Radiation of 125 I%辐射敏感性启动子调控CD基因表达联合125I照射对膀肤癌EJ细胞的杀伤作用

    Institute of Scientific and Technical Information of China (English)

    李玲; 张春丽; 李囡; 康磊; 卢霞; 张丽; 闫平; 王荣福

    2011-01-01

    以脂质体介导的辐射敏感性基因联合胞嘧啶脱氨酶(cytosine deantinase,CD)基因转染膀胱癌EJ细胞,研究放射性核素125 I照射后5-氟胞嘧啶(5-fluorocytosine,5-FC)对转染膀胧癌EJ细胞的杀伤作用.人工合成辐射敏感性启动子E8,将启动子克隆至质粒pCD2的CD基因上游,构建以E8为启动子、CD基因为目的基因的新质粒,并采用DNA测序法测定E8和CD基因的序列;脂质体Lipofectamine2000介导pE8-CD转染膀胧癌EJ细胞,用[3]I照射(吸收剂量为2舜)后,蛋白质免疫印迹分析(Western blot)测定CD蛋白表达;在转染EJ细胞中分别加人不同剂量125 I勺和5-FC,四哇盐比色法(MTT法)测定各组细胞存活率,并以未经125 I勺照射组、未加5-FC组和5-氟尿啼吮(5-FU)组(阳性对照组)进行对照.DNA测序显示构建的pE8-CD质粒含E8启动子及CD基因序列;Westem blot可检测到CD基因表达;125 I加5-FC组细胞存活率明显低于未经125 I照射组及未加5-FC组,与5-FU组相近.这表明放射性核素与基因治疗联合对肿瘤细胞具有协同杀伤作用.%To investigate the killing effect on EJ human bladder cancer cells by combination of radiation-responsive gene promoter conjoined CD/S-FC system and ionizing radiation with radionuclide 125 I, plasmid vector containing synthetic gene promoter E8 responsive to ionizing radiation(IR)and CD gene in downstream was constructed, analyzed by DNA sequencing, and transiently transfected into EJ bladder cancer cells by liposome-mediated method. Expression of downstream CD gene was detected via ionizing radiation of radionuclide 1311 at 2Gy by Western Blot. The killing effect induced by different doses of 125I and 5-FC in plasmid-transfected EJ cells was investigated by MTT colorimetric assay. DNA sequencing demonstrated that the plasmid vector contained E8 promoter and CD gene sequences , Western Blot detected the protein CD, and MTT colorimetric assay showed that the death rate of plasmid

  2. Cell killing, nuclear damage and apoptosis in Chinese hamster V79 cells after irradiation with heavy-ion beams of (16)O, (12)C and (7)Li.

    Science.gov (United States)

    Pathak, Rupak; Dey, Subrata Kumar; Sarma, Asiti; Khuda-Bukhsh, Anisur Rahman

    2007-08-15

    Chinese hamster V79 cells were exposed to high LET (linear energy transfer) (16)O-beam (625keV/mum) radiation in the dose range of 0-9.83Gy. Cell survival, micronuclei (MN), chromosomal aberrations (CA) and induction of apoptosis were studied as a follow up of our earlier study on high LET radiations ((7)Li-beam of 60keV/mum and (12)C-beam of 295keV/mum) as well as (60)Co gamma-rays. Dose dependent decline in surviving fraction was noticed along with the increase of MN frequency, CA frequency as well as percentage of apoptosis as detected by nuclear fragmentation assay. The relative intensity of DNA ladder, which is a useful marker for the determination of the extent of apoptosis induction, was also increased in a dose dependent manner. Additionally, expression of tyrosine kinase lck-1 gene, which plays an important role in response to ionizing radiation induced apoptosis, was increased with the increase of radiation doses and also with incubation time. The present study showed that all the high LET radiations were generally more effective in cell killing and inflicting other cytogenetic damages than that of low LET gamma-rays. The dose response curves revealed that (7)Li-beam was most effective in cell killing as well as inducing other nuclear damages followed by (12)C, (16)O and (60)Co gamma-rays, in that order. The result of this study may have some application in biological dosimetry for assessment of genotoxicity in heavy ion exposed subjects and in determining suitable doses for radiotherapy in cancer patients where various species of heavy ions are now being generally used.

  3. High vancomycin MICs within the susceptible range in Staphylococcus aureus bacteraemia isolates are associated with increased cell wall thickness and reduced intracellular killing by human phagocytes.

    Science.gov (United States)

    Falcón, Rocío; Martínez, Alba; Albert, Eliseo; Madrid, Silvia; Oltra, Rosa; Giménez, Estela; Soriano, Mario; Vinuesa, Víctor; Gozalbo, Daniel; Gil, María Luisa; Navarro, David

    2016-05-01

    Vancomycin minimum inhibitory concentrations (MICs) at the upper end of the susceptible range for Staphylococcus aureus have been associated with poor clinical outcomes of bloodstream infections. We tested the hypothesis that high vancomycin MICs in S. aureus bacteraemia isolates are associated with increased cell wall thickness and suboptimal bacterial internalisation or lysis by human phagocytes. In total, 95 isolates were evaluated. Original vancomycin MICs were determined by Etest. The susceptibility of S. aureus isolates to killing by phagocytes was assessed in a human whole blood assay. Internalisation of bacterial cells by phagocytes was investigated by flow cytometry. Cell wall thickness was evaluated by transmission electron microscopy. Genotypic analysis of S. aureus isolates was performed using a DNA microarray system. Vancomycin MICs were significantly higher (P=0.006) in isolates that were killed suboptimally (killing index 70%) and tended to correlate inversely (P=0.08) with the killing indices. Isolates in both killing groups were internalised by human neutrophils and monocytes with comparable efficiency. The cell wall was significantly thicker (P=0.03) in isolates in the low killing group. No genotypic differences were found between the isolates in both killing groups. In summary, high vancomycin MICs in S. aureus bacteraemia isolates were associated with increased cell wall thickness and reduced intracellular killing by phagocytes.

  4. Oncolytic adenovirus SG600-IL24 selectively kills hepatocellular carcinoma cell lines

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    AIM: To investigate the effect of oncolytic adenovirus SG600-IL24 and replication-incompetent adenovirus Ad.IL-24 on hepatocellular carcinoma (HCC) cell lines and normal liver cell line. METHODS: HCC cell lines (HepG2, Hep3B and MHCC97L) and normal liver cell line (L02) with a different p53 status were infected with SG600-IL24 and Ad.IL-24, respectively. Melanoma differentiation-associated (MDA)-7/interleukin (IL)-24 mRNA and protein expressions in infected cells were detected by reverse transcription-polym...

  5. Selective killing of cancer cells by iron oxide nanoparticles mediated through reactive oxygen species via p53 pathway

    Energy Technology Data Exchange (ETDEWEB)

    Ahamed, Maqusood, E-mail: maqusood@gmail.com; Alhadlaq, Hisham A.; Khan, M. A. Majeed [King Saud University, King Abdullah Institute for Nanotechnology (Saudi Arabia); Akhtar, Mohd. Javed [University of Lucknow, Department of Zoology (India)

    2013-01-15

    Iron oxide (Fe{sub 3}O{sub 4}) nanoparticles (NPs) are increasingly recognized for their utility in biomedical applications. However, little is known about the anticancer activity of Fe{sub 3}O{sub 4} NPs. This study was designed to investigate whether Fe{sub 3}O{sub 4} NPs induced toxicity in a cell-specific manner and determine the possible mechanisms of toxicity caused by Fe{sub 3}O{sub 4} NPs in cancer cells. Fe{sub 3}O{sub 4} NPs used in this study were synthesized by green method using {alpha}-d-glucose as a reducing agent. Prepared Fe{sub 3}O{sub 4} NPs were spherical in shape with a smooth surface, were fairly distributed, and had an average diameter of 23 nm. Cytotoxicity of Fe{sub 3}O{sub 4} NPs was examined against two types of cancer cells (human hepatocellular carcinoma HepG2 and human lung adenocarcinoma A549) and two normal cells (human lung fibroblast IMR-90 and rat hepatocytes). Fe{sub 3}O{sub 4} NPs exerted distinct effects on cell viability via killing of cancer cells while posing no toxicity on normal cells. Fe{sub 3}O{sub 4} NPs were found to induce depletion of glutathione and induction of reactive oxygen species (ROS) in both types of cancer cells (HepG2 and A549). Further, co-exposure of ascorbic acid significantly attenuated the Fe{sub 3}O{sub 4} NPs-induced oxidative stress. The mRNA levels of tumor suppressor gene p53 and apoptotic genes (caspase-3 and caspase-9) were up-regulated in both types of cancer cells due to Fe{sub 3}O{sub 4} NPs exposure. Protein level of p53, along with the higher activity of caspase-3 and caspase-9 enzymes, was also up-regulated by Fe{sub 3}O{sub 4} NPs. Taken together, our data demonstrated that Fe{sub 3}O{sub 4} NPs selectively induced apoptosis in cancer cells (HepG2 and A549) through up-regulation of p53 that might be mediated by ROS through which most of the anticancer drugs trigger apoptosis. The present study warrants further investigation on anticancer activity of Fe{sub 3}O{sub 4} NPs in relevant

  6. Selective killing of cancer cells by iron oxide nanoparticles mediated through reactive oxygen species via p53 pathway

    Science.gov (United States)

    Ahamed, Maqusood; Alhadlaq, Hisham A.; Khan, M. A. Majeed; Akhtar, Mohd. Javed

    2013-01-01

    Iron oxide (Fe3O4) nanoparticles (NPs) are increasingly recognized for their utility in biomedical applications. However, little is known about the anticancer activity of Fe3O4 NPs. This study was designed to investigate whether Fe3O4 NPs induced toxicity in a cell-specific manner and determine the possible mechanisms of toxicity caused by Fe3O4 NPs in cancer cells. Fe3O4 NPs used in this study were synthesized by green method using α- d-glucose as a reducing agent. Prepared Fe3O4 NPs were spherical in shape with a smooth surface, were fairly distributed, and had an average diameter of 23 nm. Cytotoxicity of Fe3O4 NPs was examined against two types of cancer cells (human hepatocellular carcinoma HepG2 and human lung adenocarcinoma A549) and two normal cells (human lung fibroblast IMR-90 and rat hepatocytes). Fe3O4 NPs exerted distinct effects on cell viability via killing of cancer cells while posing no toxicity on normal cells. Fe3O4 NPs were found to induce depletion of glutathione and induction of reactive oxygen species (ROS) in both types of cancer cells (HepG2 and A549). Further, co-exposure of ascorbic acid significantly attenuated the Fe3O4 NPs-induced oxidative stress. The mRNA levels of tumor suppressor gene p53 and apoptotic genes (caspase-3 and caspase-9) were up-regulated in both types of cancer cells due to Fe3O4 NPs exposure. Protein level of p53, along with the higher activity of caspase-3 and caspase-9 enzymes, was also up-regulated by Fe3O4 NPs. Taken together, our data demonstrated that Fe3O4 NPs selectively induced apoptosis in cancer cells (HepG2 and A549) through up-regulation of p53 that might be mediated by ROS through which most of the anticancer drugs trigger apoptosis. The present study warrants further investigation on anticancer activity of Fe3O4 NPs in relevant animal models.

  7. Mathematical analysis and simulations involving chemotherapy and surgery on large human tumours under a suitable cell-kill functional response.

    Science.gov (United States)

    Rodrigues, Diego Samuel; de Arruda Mancera, Paulo Fernando

    2013-02-01

    Dosage and frequency of treatment schedules are important for successful chemotherapy. However, in this work we argue that cell-kill response and tumoral growth should not be seen as separate and therefore are essential in a mathematical cancer model. This paper presents a mathematical model for sequencing of cancer chemotherapy and surgery. Our purpose is to investigate treatments for large human tumours considering a suitable cell-kill dynamics. We use some biological and pharmacological data in a numerical approach, where drug administration occurs in cycles (periodic infusion) and surgery is performed instantaneously. Moreover, we also present an analysis of stability for a chemotherapeutic model with continuous drug administration. According to Norton and Simon [22], our results indicate that chemotherapy is less efficient in treating tumours that have reached a plateau level of growing and that a combination with surgical treatment can provide better outcomes.

  8. Effect of Refractory on Nonmetallic Inclusions in Al-Killed Steel

    Science.gov (United States)

    Deng, Zhiyin; Zhu, Miaoyong; Sichen, Du

    2016-10-01

    To understand the effect of refractory on the evolution of inclusions in the secondary steelmaking, steel samples were taken at different stages of the process train. Laboratory experiments were carried out using different refractories (alumina, spinel, and MgO). In the laboratory study, the types of inclusions considered were alumina, spinel, and calcium aluminate. The focus was given to Al-killed steel. The results showed that alumina refractory and spinel refractory had little effect on all the three types of inclusions, while the effect of MgO refractory depended on the activity of dissolved oxygen in liquid steel. With lower oxygen activity, alumina inclusions could transform into spinel inclusions. No evident change could be found for spinel and calcium aluminate inclusions. When the oxygen activity was high enough, spinel inclusions could not be generated from alumina inclusions. The laboratory results helped in understanding the evolution of the inclusions in the industrial process.

  9. Hypothermia postpones DNA damage repair in irradiated cells and protects against cell killing

    Energy Technology Data Exchange (ETDEWEB)

    Baird, Brandon J.; Dickey, Jennifer S.; Nakamura, Asako J.; Redon, Christophe E.; Parekh, Palak [Laboratory of Molecular Pharmacology, CCR, NCI, Bethesda, MD 20892 (United States); Griko, Yuri V. [Radiation and Space Biotechnology Branch, NASA Ames Research Center, Moffett Field, CA 94035 (United States); Aziz, Khaled; Georgakilas, Alexandros G. [Biology Department, East Carolina University, Greenville, NC 27858 (United States); Bonner, William M. [Laboratory of Molecular Pharmacology, CCR, NCI, Bethesda, MD 20892 (United States); Martin, Olga A., E-mail: sedelnio@mail.nih.gov [Laboratory of Molecular Pharmacology, CCR, NCI, Bethesda, MD 20892 (United States)

    2011-06-03

    Hibernation is an established strategy used by some homeothermic organisms to survive cold environments. In true hibernation, the core body temperature of an animal may drop to below 0 {sup o}C and metabolic activity almost cease. The phenomenon of hibernation in humans is receiving renewed interest since several cases of victims exhibiting core body temperatures as low as 13.7 {sup o}C have been revived with minimal lasting deficits. In addition, local cooling during radiotherapy has resulted in normal tissue protection. The experiments described in this paper were prompted by the results of a very limited pilot study, which showed a suppressed DNA repair response of mouse lymphocytes collected from animals subjected to 7-Gy total body irradiation under hypothermic (13 {sup o}C) conditions, compared to normothermic controls. Here we report that human BJ-hTERT cells exhibited a pronounced radioprotective effect on clonogenic survival when cooled to 13 {sup o}C during and 12 h after irradiation. Mild hypothermia at 20 and 30 {sup o}C also resulted in some radioprotection. The neutral comet assay revealed an apparent lack on double strand break (DSB) rejoining at 13 {sup o}C. Extension of the mouse lymphocyte study to ex vivo-irradiated human lymphocytes confirmed lower levels of induced phosphorylated H2AX ({gamma}-H2AX) and persistence of the lesions at hypothermia compared to the normal temperature. Parallel studies of radiation-induced oxidatively clustered DNA lesions (OCDLs) revealed partial repair at 13 {sup o}C compared to the rapid repair at 37 {sup o}C. For both {gamma}-H2AX foci and OCDLs, the return of lymphocytes to 37 {sup o}C resulted in the resumption of normal repair kinetics. These results, as well as observations made by others and reviewed in this study, have implications for understanding the radiobiology and protective mechanisms underlying hypothermia and potential opportunities for exploitation in terms of protecting normal tissues against

  10. Intracellular growth of Mycobacterium tuberculosis after macrophage cell death leads to serial killing of host cells

    Science.gov (United States)

    Mahamed, Deeqa; Boulle, Mikael; Ganga, Yashica; Mc Arthur, Chanelle; Skroch, Steven; Oom, Lance; Catinas, Oana; Pillay, Kelly; Naicker, Myshnee; Rampersad, Sanisha; Mathonsi, Colisile; Hunter, Jessica; Sreejit, Gopalkrishna; Pym, Alexander S; Lustig, Gila; Sigal, Alex

    2017-01-01

    A hallmark of pulmonary tuberculosis is the formation of macrophage-rich granulomas. These may restrict Mycobacterium tuberculosis (Mtb) growth, or progress to central necrosis and cavitation, facilitating pathogen growth. To determine factors leading to Mtb proliferation and host cell death, we used live cell imaging to track Mtb infection outcomes in individual primary human macrophages. Internalization of Mtb aggregates caused macrophage death, and phagocytosis of large aggregates was more cytotoxic than multiple small aggregates containing similar numbers of bacilli. Macrophage death did not result in clearance of Mtb. Rather, it led to accelerated intracellular Mtb growth regardless of prior activation or macrophage type. In contrast, bacillary replication was controlled in live phagocytes. Mtb grew as a clump in dead cells, and macrophages which internalized dead infected cells were very likely to die themselves, leading to a cell death cascade. This demonstrates how pathogen virulence can be achieved through numbers and aggregation states. DOI: http://dx.doi.org/10.7554/eLife.22028.001 PMID:28130921

  11. Human papillomavirus E6E7-mediated adenovirus cell killing: selectivity of mutant adenovirus replication in organotypic cultures of human keratinocytes.

    Science.gov (United States)

    Balagué, C; Noya, F; Alemany, R; Chow, L T; Curiel, D T

    2001-08-01

    Replication-competent adenoviruses are being investigated as potential anticancer agents. Exclusive virus replication in cancer cells has been proposed as a safety trait to be considered in the design of oncolytic adenoviruses. From this perspective, we have investigated several adenovirus mutants for their potential to conditionally replicate and promote the killing of cells expressing human papillomavirus (HPV) E6 and E7 oncoproteins, which are present in a high percentage of anogenital cancers. For this purpose, we have employed an organotypic model of human stratified squamous epithelium derived from primary keratinocytes that have been engineered to express HPV-18 oncoproteins stably. We show that, whereas wild-type adenovirus promotes a widespread cytopathic effect in all infected cells, E1A- and E1A/E1B-deleted adenoviruses cause no deleterious effect regardless of the coexpression of HPV18 E6E7. An adenovirus deleted in the CR2 domain of E1A, necessary for binding to the pRB family of pocket proteins, shows no selectivity of replication as it efficiently kills all normal and E6E7-expressing keratinocytes. Finally, an adenovirus mutant deleted in the CR1 and CR2 domains of E1A exhibits preferential replication and cell killing in HPV E6E7-expressing cultures. We conclude that the organotypic keratinocyte culture represents a distinct model to evaluate adenovirus selectivity and that, based on this model, further modifications of the adenovirus genome are required to restrict adenovirus replication to tumor cells.

  12. Investigating CTL mediated killing with a 3D cellular automaton.

    Directory of Open Access Journals (Sweden)

    Frederik Graw

    2009-08-01

    Full Text Available Cytotoxic T lymphocytes (CTLs are important immune effectors against intra-cellular pathogens. These cells search for infected cells and kill them. Recently developed experimental methods in combination with mathematical models allow for the quantification of the efficacy of CTL killing in vivo and, hence, for the estimation of parameters that characterize the effect of CTL killing on the target cell populations. It is not known how these population-level parameters relate to single-cell properties. To address this question, we developed a three-dimensional cellular automaton model of the region of the spleen where CTL killing takes place. The cellular automaton model describes the movement of different cell populations and their interactions. Cell movement patterns in our cellular automaton model agree with observations from two-photon microscopy. We find that, despite the strong spatial nature of the kinetics in our cellular automaton model, the killing of target cells by CTLs can be described by a term which is linear in the target cell frequency and saturates with respect to the CTL levels. Further, we find that the parameters describing CTL killing on the population level are most strongly impacted by the time a CTL needs to kill a target cell. This suggests that the killing of target cells, rather than their localization, is the limiting step in CTL killing dynamics given reasonable frequencies of CTL. Our analysis identifies additional experimental directions which are of particular importance to interpret estimates of killing rates and could advance our quantitative understanding of CTL killing.

  13. ABT-737, a Bcl-2 Selective Inhibitor, and Chloroquine Synergistically Kill Renal Cancer Cells.

    Science.gov (United States)

    Yin, Pei; Jia, Jinpeng; Li, Jijun; Song, Yan; Zhang, Yiyan; Chen, Fengkun

    2016-01-01

    Renal cell carcinoma (RCC) is the most common malignancy in the kidney in the world, and the 5-year overall survival for patients remains poor due to the lack of effective treatment strategies. Although ABT-737, as a Bcl-2/Bcl-xL inhibitor, has recently emerged as a novel cancer therapeutic reagent, apoptosis induced by ABT-737 is often blocked in several types of cancer cells. This study investigated whether the combination of the small-molecule BH3 mimetic ABT-737 and the lysosome inhibitor chloroquine was an effective strategy for treating renal cancer cells. We found that the combination of ABT-737 and chloroquine synergistically decreased cell viability when compared to treatment with either single reagent. Cell apoptosis induced by a combined treatment was markedly inhibited by the caspase inhibitors z-DEVD-FMK and z-VAD-FMK. It was also inhibited by cathepsin inhibitor E-64 and CTSI (cathepsin inhibitor), which suggested that apoptosis was dependent on the cascade of caspase activation and cathepsins released from lysosomes. Furthermore, we found that ABT-737 could increase the cell level of ROS, which triggers cathepsin-mediated cell death and augments the role of chloroquine in cell death. So the combination of ABT-737 and chloroquine was an effective strategy for the treatment of renal cancer cells, and this combined strategy may widen the therapeutic window of ABT-737 and chloroquine as well as enhance the clinical efficacy of synergistic drug combinations.

  14. Cisplatin and 5-fluorouracil Enhance Expression of NKG2D Ligands and CIK Cells Killing Effect of Human Esophagus Carcinoma Cell EC9706%顺铂和氟尿嘧啶增强食管癌细胞NKG2D配体的表达及CIK细胞的杀伤活性

    Institute of Scientific and Technical Information of China (English)

    李瑞君; 梅家转; 禹萌; 刘桂举; 冯睿婷; 张晓娟

    2012-01-01

    20:1. Results DDP, 5-Fu could decrease the proliferation and survival rate of EC9706 cells,the IC50. was 5 jug/ml and 10 μg/ml, respectively,which increased mRNA expressions of signal pathway molecules involved in DNA damage and repair system. MICA, MICB, ULBP2, ULBP3 on EG9706 cells were over expressed after 72 h cultured with 1/2 IC50 DDP, while expression of MICA, UL.BP2, UI.BP3 were higher after treated by 1/2 IC50 5-Fu. Cytotoxichy of CIK cells against EC9706 cells cultured by 1/2 IC50 DDP, 1/2 IC50 5-Fu were (37. OS ±0.62)%.(59. 33 ± 2. 10)% , (52. 44 ± 0. 97)%,respectively. Cytotoxicity of CIK cells against EC9706 cells cultured by either 1/2 IC50 DDP or 1/2 IC50 5-Fu was significantly enhanced. Conclusion The results indicate that DDP or 5-Fu can enhance the susceptibility of EC9706 cells to CIK cells-mediated killing effect by upregulating the expressions of NKG2D ligands through activating signal pathway molecules involved in DNA damage and repair system.

  15. Inductive heating kills cells that contribute to plaque: a proof-of-concept

    Directory of Open Access Journals (Sweden)

    Angelo Gaitas

    2015-04-01

    Full Text Available Inducing cell death by heating targeted particles shows promise in cancer treatment. Here, we aim to demonstrate the feasibility of extending the use of this technique to treat and remove vascular deposits and thrombosis. We used induction heating of macrophages, which are key contributors to atherosclerosis and have demonstrated clear feasibility for heating and destroying these cells using ferromagnetic and pure iron particles. Specifically, iron particles achieved maximum temperatures of 51 ± 0.5 °C and spherical particles achieved a maximum temperature of 43.9 ± 0.2 °C (N = 6 after 30 min of inductive heating. Two days of subsequent observation demonstrated that inductive heating led to a significant reduction in cell number. Prior to induction heating, cell density was 105,000 ± 20,820 cells/ml (N = 3. This number was reduced to 6,666 ± 4,410 cells/ml for the spherical particles and 16,666 ± 9,280 cells/ml for the iron particles 24 h after inductive heating. Though cell density increased on the second day following inductive heating, the growth was minimal. Cells grew to 26,667 ± 6,670 cells/ml and 30,000 ± 15,280 cells/ml respectively. Compared to cell cultures with iron and spherical particles that were not subjected to induction heating, we observed a 97% reduction in cell count for the spherical particles and a 91% reduction for the iron particles after the first 24 h. After 48 h we observed a 95% reduction in cell growth for both spherical and iron particles. Induction heating of microparticles was thus highly effective in reducing the macrophage population and preventing their growth. These results demonstrate the feasibility of targeting cells involved in atherosclerosis and warrant further research into potential clinical applications.

  16. Inductive heating kills cells that contribute to plaque: a proof-of-concept.

    Science.gov (United States)

    Gaitas, Angelo; Kim, Gwangseong

    2015-01-01

    Inducing cell death by heating targeted particles shows promise in cancer treatment. Here, we aim to demonstrate the feasibility of extending the use of this technique to treat and remove vascular deposits and thrombosis. We used induction heating of macrophages, which are key contributors to atherosclerosis and have demonstrated clear feasibility for heating and destroying these cells using ferromagnetic and pure iron particles. Specifically, iron particles achieved maximum temperatures of 51 ± 0.5 °C and spherical particles achieved a maximum temperature of 43.9 ± 0.2 °C (N = 6) after 30 min of inductive heating. Two days of subsequent observation demonstrated that inductive heating led to a significant reduction in cell number. Prior to induction heating, cell density was 105,000 ± 20,820 cells/ml (N = 3). This number was reduced to 6,666 ± 4,410 cells/ml for the spherical particles and 16,666 ± 9,280 cells/ml for the iron particles 24 h after inductive heating. Though cell density increased on the second day following inductive heating, the growth was minimal. Cells grew to 26,667 ± 6,670 cells/ml and 30,000 ± 15,280 cells/ml respectively. Compared to cell cultures with iron and spherical particles that were not subjected to induction heating, we observed a 97% reduction in cell count for the spherical particles and a 91% reduction for the iron particles after the first 24 h. After 48 h we observed a 95% reduction in cell growth for both spherical and iron particles. Induction heating of microparticles was thus highly effective in reducing the macrophage population and preventing their growth. These results demonstrate the feasibility of targeting cells involved in atherosclerosis and warrant further research into potential clinical applications.

  17. Enhanced Expression of T-Cell Immunoglobulin and Mucin Domain Protein 3 in Endothelial Cells Facilitates Intracellular Killing of Rickettsia heilongjiangensis.

    Science.gov (United States)

    Yang, Xiaomei; Jiao, Jun; Han, Gencheng; Gong, Wenping; Wang, Pengcheng; Xiong, Xiaolu; Wen, Bohai

    2016-01-01

    Rickettsia heilongjiangensis is the pathogen of Far eastern spotted fever, and T-cell immunoglobulin and mucin domain protein 3 (Tim-3) is expressed in human vascular endothelial cells, the major target cells of rickettsiae. In the present study, we investigated the effects of altered Tim-3 expression in vivo in mice and in vitro in human endothelial cells, on day 3 after R. heilongjiangensis infection. Compared with corresponding controls, rickettsial burdens both in vivo and in vitro were significantly higher with blocked Tim-3 signaling or silenced Tim-3 and significantly lower with overexpressed Tim-3. Additionally, the expression of inducible nitric oxide synthase and interferon γ in endothelial cells with blocked Tim-3 signaling or silenced Tim-3 was significantly lower, while the expression of inducible nitric oxide synthase, interferon γ, and tumor necrosis factor α in transgenic mice with Tim-3 overexpression was significantly higher. These results reveal that enhanced Tim-3 expression facilitates intracellular rickettsial killing in a nitric oxide-dependent manner in endothelial cells during the early phase of rickettsial infection.

  18. Cell killing and chromatid damage in primary human bronchial epithelial cells irradiated with accelerated 56Fe ions

    Science.gov (United States)

    Suzuki, M.; Piao, C.; Hall, E. J.; Hei, T. K.

    2001-01-01

    We examined cell killing and chromatid damage in primary human bronchial epithelial cells irradiated with high-energy 56Fe ions. Cells were irradiated with graded doses of 56Fe ions (1 GeV/nucleon) accelerated with the Alternating Gradient Synchrotron at Brookhaven National Laboratory. The survival curves for cells plated 1 h after irradiation (immediate plating) showed little or no shoulder. However, the survival curves for cells plated 24 h after irradiation (delayed plating) had a small initial shoulder. The RBE for 56Fe ions compared to 137Cs gamma rays was 1.99 for immediate plating and 2.73 for delayed plating at the D10. The repair ratio (delayed plating/immediate plating) was 1.67 for 137Cs gamma rays and 1.22 for 56Fe ions. The dose-response curves for initially measured and residual chromatid fragments detected by the Calyculin A-mediated premature chromosome condensation technique showed a linear response. The results indicated that the induction frequency for initially measured fragments was the same for 137Cs gamma rays and 56Fe ions. On the other hand, approximately 85% of the fragments induced by 137Cs gamma rays had rejoined after 24 h of postirradiation incubation; the corresponding amount for 56Fe ions was 37%. Furthermore, the frequency of chromatid exchanges induced by gamma rays measured 24 h after irradiation was higher than that induced by 56Fe ions. No difference in the amount of chromatid damage induced by the two types of radiations was detected when assayed 1 h after irradiation. The results suggest that high-energy 56Fe ions induce a higher frequency of complex, unrepairable damage at both the cellular and chromosomal levels than 137Cs gamma rays in the target cells for radiation-induced lung cancers.

  19. Design, synthesis, and in vitro and in vivo biological studies of a 3'-deoxythymidine conjugate that potentially kills cancer cells selectively.

    Directory of Open Access Journals (Sweden)

    Qiong Wei

    Full Text Available Thymidine kinases (TKs have been considered one of the potential targets for anticancer therapeutic because of their elevated expressions in cancer cells. However, nucleobase analogs targeting TKs have shown poor selective cytotoxicity in cancer cells despite effective antiviral activity. 3'-Deoxythymidine phenylquinoxaline conjugate (dT-QX was designed as a novel nucleobase analog to target TKs in cancer cells and block cell replication via conjugated DNA intercalating quinoxaline moiety. In vitro cell screening showed that dT-QX selectively kills a variety of cancer cells including liver carcinoma, breast adenocarcinoma and brain glioma cells; whereas it had a low cytotoxicity in normal cells such as normal human liver cells. The anticancer activity of dT-QX was attributed to its selective inhibition of DNA synthesis resulting in extensive mitochondrial superoxide stress in cancer cells. We demonstrate that covalent linkage with 3'-deoxythymidine uniquely directed cytotoxic phenylquinoxaline moiety more toward cancer cells than normal cells. Preliminary mouse study with subcutaneous liver tumor model showed that dT-QX effectively inhibited the growth of tumors. dT-QX is the first molecule of its kind with highly amendable constituents that exhibits this selective cytotoxicity in cancer cells.

  20. Afa/Dr diffusely adhering Escherichia coli strain C1845 induces neutrophil extracellular traps that kill bacteria and damage human enterocyte-like cells.

    Science.gov (United States)

    Marin-Esteban, Viviana; Turbica, Isabelle; Dufour, Guillaume; Semiramoth, Nicolas; Gleizes, Aude; Gorges, Roseline; Beau, Isabelle; Servin, Alain L; Lievin-Le Moal, Vanessa; Sandré, Catherine; Chollet-Martin, Sylvie

    2012-05-01

    We recently documented the neutrophil response to enterovirulent diffusely adherent Escherichia coli expressing Afa/Dr fimbriae (Afa/Dr DAEC), using the human myeloid cell line PLB-985 differentiated into fully mature neutrophils. Upon activation, particularly during infections, neutrophils release neutrophil extracellular traps (NETs), composed of a nuclear DNA backbone associated with antimicrobial peptides, histones, and proteases, which entrap and kill pathogens. Here, using fluorescence microscopy and field emission scanning electron microscopy, we observed NET production by PLB-985 cells infected with the Afa/Dr wild-type (WT) E. coli strain C1845. We found that these NETs were able to capture, immobilize, and kill WT C1845 bacteria. We also developed a coculture model of human enterocyte-like Caco-2/TC7 cells and PLB-985 cells previously treated with WT C1845 and found, for the first time, that the F-actin cytoskeleton of enterocyte-like cells is damaged in the presence of bacterium-induced NETs and that this deleterious effect is prevented by inhibition of protease release. These findings provide new insights into the neutrophil response to bacterial infection via the production of bactericidal NETs and suggest that NETs may damage the intestinal epithelium, particularly in situations such as inflammatory bowel diseases.

  1. Development of a novel in-vivo drug/in-vitro light system to investigate mechanisms of cell killing with photodynamic therapy

    Science.gov (United States)

    Hampton, James A.; Selman, Steven H.

    1991-06-01

    Over the last decade considerable interest has developed in the use of exogenously administered chromophores in combination with visible light for the treatment of human tumors. Whether cells are killed directly, or indirectly as a result of disruption of the tissue, microvasculature is unknown. The authors have developed methods to assess in short term culture the effects of PDT on precision cut tissue slices. The use of these tissue slices provide an important link between in vivo studies and in vitro isolated cultured cells for the following reasons: 1) slices contain all of the normal cells in their proper in vivo architectural arrangement; 2) since slices can be obtained relatively easily and in a very short period of time (a few minutes), animals can be treated with compounds in vivo, the tissue can be removed, sliced and mechanistic studies performed in vitro (without the several hours delay required to produce cultured cells); 3) in vitro comparisons between species, including human, can be readily made; and 4) mechanisms of PDT-induced cell killing can be studied in the absence of a functioning vascular system. Using this in vivo drug/in vitro light system, the results presented will detail the findings using normal rat liver and a transplantable rat tumor model.

  2. ROS accumulation by PEITC selectively kills ovarian cancer cells via UPR-mediated apoptosis

    Directory of Open Access Journals (Sweden)

    Yoon-hee eHong

    2015-07-01

    Full Text Available Unfolded protein response (UPR is crucial for both survival and death of mammalian cells, which is regulated by reactive oxygen species (ROS and nutrient depletion. In this study, we demonstrated the effect of ROS-accumulation, induced by β-phenethyl isothiocyanate (PEITC, on UPR mediated apoptosis in ovarian cancer cells. We used ovarian cancer cell lines, PA-1 and SKOV-3, with different p53 status (wild- and null- type, respectively. PEITC caused increased ROS-accumulation and inhibited proliferation selectively in ovarian cancer cells, and glutathione (GSH depletion in SKOV-3. However, PEITC did not cause any effect in normal ovarian epithelial cells and peripheral blood mononuclear cells. After 48 h of PEITC treatment (5 µM, apoptotic cell death was shown to increase significantly in the ovarian cancer cells and not in the normal cells. The key regulator of UPR-mediated apoptosis, CHOP/GADD153 and ER resident chaperone BiP/GRP78 were parallely up-regulated with activation of two major sensors of the UPR (PERK and ATF-6 in PA-1; PERK, and IRE1α in SKOV-3 in response to ROS accumulation induced by PEITC (5 µM. ROS scavenger, N-acetyl-cysteine (NAC, attenuated the effect of PEITC on UPR signatures (P-PERK, IRE1α, CHOP/GADD153, and BiP/GRP78, suggesting the involvement of ROS in UPR-mediated apoptosis. Altogether, PEITC induces UPR-mediated apoptosis in ovarian cancer cells via accumulation of ROS in a cancer-specific manner.

  3. Effect of Temper Rolling on Tensile Properties of Low-Si AI-Killed Sheet Steel

    Institute of Scientific and Technical Information of China (English)

    MA Qing-long; WANG Dong-cheng; LIU Hong-min; LU Hai-ming

    2009-01-01

    The tensile properties of steel after temper rolling are affected by the reduction;low-Si Al-killed sheet steel was taken to study the effect of temper rolling on the tensile properties.The results indicate that the yield strength first decreases with the increase of reduction,and then increases.The relationship between the yield strength and the reduction can be expressed using quadratic function.The tensile strength increases with the increase of the reduction,while the total elongation decreases with the increase of the reduction,and the relationship between them and the reduction can be expressed using power function.Under the same condition,the results also indicate that the yield strength and tensile strength of steels across the rolling direction are all larger than those along the rolling direction; there is no obvious distinction between the total elongation along the rolling direction and that across the rolling direction.

  4. Nanostructured Surfaces to Target and Kill Circulating Tumor Cells While Repelling Leukocytes

    Directory of Open Access Journals (Sweden)

    Michael J. Mitchell

    2012-01-01

    Full Text Available Hematogenous metastasis, the process of cancer cell migration from a primary to distal location via the bloodstream, typically leads to a poor patient prognosis. Selectin proteins hold promise in delivering drug-containing nanocarriers to circulating tumor cells (CTCs in the bloodstream, due to their rapid, force-dependent binding kinetics. However, it is challenging to deliver such nanocarriers while avoiding toxic effects on healthy blood cells, as many possess ligands that adhesively interact with selectins. Herein, we describe a nanostructured surface to capture flowing cancer cells, while preventing human neutrophil adhesion. Microtube surfaces with immobilized halloysite nanotubes (HNTs and E-selectin functionalized liposomal doxorubicin (ES-PEG L-DXR significantly increased the number of breast adenocarcinoma MCF7 cells captured from flow, yet also significantly reduced the number of captured neutrophils. Neutrophils firmly adhered and projected pseudopods on surfaces coated only with liposomes, while neutrophils adherent to HNT-liposome surfaces maintained a round morphology. Perfusion of both MCF7 cells and neutrophils resulted in primarily cancer cell adhesion to the HNT-liposome surface, and induced significant cancer cell death. This work demonstrates that nanostructured surfaces consisting of HNTs and ES-PEG L-DXR can increase CTC recruitment for chemotherapeutic delivery, while also preventing healthy cell adhesion and uptake of therapeutic intended for CTCs.

  5. Irradiation-induced up-regulation of HLA-E on macrovascular endothelial cells confers protection against killing by activated natural killer cells.

    Directory of Open Access Journals (Sweden)

    Isabelle Riederer

    Full Text Available BACKGROUND: Apart from the platelet/endothelial cell adhesion molecule 1 (PECAM-1, CD31, endoglin (CD105 and a positive factor VIII-related antigen staining, human primary and immortalized macro- and microvascular endothelial cells (ECs differ in their cell surface expression of activating and inhibitory ligands for natural killer (NK cells. Here we comparatively study the effects of irradiation on the phenotype of ECs and their interaction with resting and activated NK cells. METHODOLOGY/PRINCIPAL FINDINGS: Primary macrovascular human umbilical vein endothelial cells (HUVECs only express UL16 binding protein 2 (ULBP2 and the major histocompatibility complex (MHC class I chain-related protein MIC-A (MIC-A as activating signals for NK cells, whereas the corresponding immortalized EA.hy926 EC cell line additionally present ULBP3, membrane heat shock protein 70 (Hsp70, intercellular adhesion molecule ICAM-1 (CD54 and HLA-E. Apart from MIC-B, the immortalized human microvascular endothelial cell line HMEC, resembles the phenotype of EA.hy926. Surprisingly, primary HUVECs are more sensitive to Hsp70 peptide (TKD plus IL-2 (TKD/IL-2-activated NK cells than their immortalized EC counterpatrs. This finding is most likely due to the absence of the inhibitory ligand HLA-E, since the activating ligands are shared among the ECs. The co-culture of HUVECs with activated NK cells induces ICAM-1 (CD54 and HLA-E expression on the former which drops to the initial low levels (below 5% when NK cells are removed. Sublethal irradiation of HUVECs induces similar but less pronounced effects on HUVECs. Along with these findings, irradiation also induces HLA-E expression on macrovascular ECs and this correlates with an increased resistance to killing by activated NK cells. Irradiation had no effect on HLA-E expression on microvascular ECs and the sensitivity of these cells to NK cells remained unaffected. CONCLUSION/SIGNIFICANCE: These data emphasize that an irradiation

  6. Bispecific Antibodies that Mediate Killing of Cells Infected with Human Immunodeficiency Virus of Any Strain

    Science.gov (United States)

    Berg, Jorg; Lotscher, Erika; Steimer, Kathelyn S.; Capon, Daniel J.; Baenziger, Jurg; Jack, Hans-Martin; Wabl, Matthias

    1991-06-01

    Although AIDS patients lose human immunodeficiency virus (HIV)-specific cytotoxic T cells, their remaining CD8-positive T lymphocytes maintain cytotoxic function. To exploit this fact we have constructed bispecific antibodies that direct cytotoxic T lymphocytes of any specificity to cells that express gp120 of HIV. These bispecific antibodies comprise one heavy/light chain pair from an antibody to CD3, linked to a heavy chain whose variable region has been replaced with sequences from CD4 plus a second light chain. CD3 is part of the antigen receptor on T cells and is responsible for signal transduction. In the presence of these bispecific antibodies, T cells of irrelevant specificity effectively lyse HIV-infected cells in vitro.

  7. How effective is road mitigation at reducing road-kill? A meta-analysis

    NARCIS (Netherlands)

    Rytwinski, Trina; Soanes, Kylie; Jaeger, Jochen A.G.; Fahrig, Lenore; Findlay, C.S.; Houlahan, Jeff; Ree, van der Rodney; Grift, van der Edgar A.

    2016-01-01

    Road traffic kills hundreds of millions of animals every year, posing a critical threat to the
    populations of many species. To address this problem there are more than forty types of
    road mitigation measures available that aim to reduce wildlife mortality on roads (road-kill).
    For road p

  8. Adenovirus vector expressing mda-7 selectively kills hepatocellular carcinoma cell line Hep3B

    Institute of Scientific and Technical Information of China (English)

    Xin-Bo Xue; Kun Chen; Cong-Jun Wang; Jian-Wei Zheng; Yuan Yu; Zhi-Hai Peng; Zai-De Wu

    2008-01-01

    BACKGROUND: Melanoma differentiation associated gene-7 (mda-7) is a novel tumor suppressor gene, which has suppressor activity in a broad spectrum of human cancer cells both in vitro and in vivo through activation of various intracellular signaling pathways. In this study, we investigated the potential effect of mda-7 on human hepatocellular carcinoma (HCC) in vitro. METHODS: Cells from the human HCC cell line Hep3B and the human liver cell line L-02 were assigned to three groups. One was cultured in Dulbecco's modiifed Eagle's medium without serum (control). The others were transfected with adenovirus expressing the mda-7 gene (Ad.mda-7) or adenovirus vector serving as negative control (Ad.vec). The expression of MDA-7 and Bcl-2 proteins in Hep3B and L-02 cells was conifrmed by the reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay. The methyl thiazolyl tetrazolium colorimetric assay and lfow cytometry were used to assess tumor cell proliferation and the cell cycle. Hoechst and Annexin-V/propidium iodide staining were used to study mda-7 gene expression in Hep3B and L-02 cells. The expression of MDA-7, Bcl-2 and Bax proteins were detected by Western blotting. RESULTS: The mda-7 gene was expressed in Hep3B and L-02 cells. The protein concentrations of MDA-7 in supernatants were 790 and 810 pg/ml, respectively. mda-7 induced Hep3B growth suppression and apoptosis, compared with Ad.mda-7 and control (P CONCLUSIONS: mda-7 selectively induces growth inhibi-tion and apoptosis in the HCC cell line Hep3B but not in the normal liver cell line L-02 via downregulating the anti-apoptosis protein Bcl-2. It could be an ideal gene for gene therapy in HCC.

  9. N′1,N′3-Dimethyl-N′1,N′3-bis(phenylcarbonothioyl Propanedihydrazide (Elesclomol Selectively Kills Cisplatin Resistant Lung Cancer Cells through Reactive Oxygen Species (ROS

    Directory of Open Access Journals (Sweden)

    Niramol Savaraj

    2009-12-01

    Full Text Available Cisplatin is an important chemotherapeutic agent in lung cancer treatment. The mechanism of drug resistance to cisplatin is complex and historically has been difficult to overcome. We report here that cisplatin resistant lung cancer cell lines possess high basal levels of reactive oxygen species (ROS when compared to normal cells and their parental cell counterparts. These resistant cells also have low thioredoxin (TRX levels which may be one of the contributory factors to high ROS. N′1,N′3-dimethyl-N′1,N'3-bis(phenylcarbonothioyl propanedihydrazide (elesclomol, an agent known to increase ROS is selectively toxic to cisplatin-resistant cells, while sparing normal cells and the parental counterpart. The cytotoxic effect of elesclomol in resistant cells is accompanied by further decreases in TRX and glutathione (GSH antioxidant systems, while opposite results were found in parental cells. The ID50 of elesclomol in cisplatin-resistant cells ranged from 5–10 nM, which is well within clinically achievable ranges. N-Acetylcysteine (NAC, which is known to neutralize ROS, can abolish the cytotoxic effect of elesclomol, suggesting that the cytotoxic effect results from increased ROS. Overall, our data suggest that elesclomol selectively kills cisplatin-resistant tumor cells through increased ROS. This agent may hold potential to overcome cisplatin resistance and should be further explored to treat patients who have failed cisplatin therapy.

  10. 不同剂量γ射线辐照对小鼠红内期疟原虫杀伤作用的研究%The exposure dose-effectiveness of γirradiation on killing the plasmedium in mice red blood cells

    Institute of Scientific and Technical Information of China (English)

    刘忠湘; 李英辉; 赵亚; 雷俊川; 薛采芳; 夏爱军; 张献清

    2009-01-01

    Objective To study the exposure dose-effectiveness of γ irradiation on killing the plasmodium in the mice RBC, for the further exploration on the method that could kill the plasmodium in RBC without affecting the activity and function of normal RBC. Methods After infection with Plamodium yoelii (P. y), blood was collected from mice and ex-posed to γ irradiation (radiated group). An unirradiated group served as control. In the irradiated group, P. y infected blood was divided into three aliquots, each aliquot was irradiated one time by γ radiation using Gammacell 1000 Elite blood radiation apparatus. The dosage of each aliquot was 25, 35 and 45Gy. After irradiation, the blood samples were stored at 4℃. Then mice were inoculated with these irradiated blood stored for 1,3 or 5 days after irradiation, or with unirradiated blood. Two days later, the blood samples were taken from inoculated mice and were examined under microscope and plas-modium infection rates were calculated. Results The mice in the control group had parasitemia much earlier than those in irradiated group (1——2 days), and the plasmedium infection rate in the control group was significantly higher than that in thc irradiated group(3.7% vs 0. 07%). With increasing dosage of irradiation, the survived plasmodium in blood de-creased, and survival of mice increased(8——12 days). After 45 Gy irradiation and 5 day storage at 4℃, there were no plasmodium found in the red blood cell of inoculated mice. In the control group, blood testing result was positive, and all the mice died. Conclusion Plasmodium in mice RBC can be killed effectively when blood is exposed to 45Gy irradiation and stored at 4℃ for 5 days.%目的 探寻不影响正常红细胞活性及功能且能够有效杀伤血液中疟原虫的γ射线辐照方法.方法 感染约氏疟原虫(Plasmodium yoclii,P.y)种鼠血液,设实验组(辐照组,n=5)和对照组(未辐照,n=5),辐照组采用Gammacdl 1000 Elite血液

  11. Accelerated killing of cancer cells using a multifunctional single-walled carbon nanotube-based system for targeted drug delivery in combination with photothermal therapy

    Directory of Open Access Journals (Sweden)

    Jeyamohan P

    2013-07-01

    Full Text Available Prashanti Jeyamohan, Takashi Hasumura, Yutaka Nagaoka, Yasuhiko Yoshida, Toru Maekawa, D Sakthi Kumar Bio-Nano Electronics Research Centre, Graduate School of Interdisciplinary New Science, Toyo University, Kawagoe, Japan Abstract: The photothermal effect of single-walled carbon nanotubes (SWCNTs in combination with the anticancer drug doxorubicin (DOX for targeting and accelerated destruction of breast cancer cells is demonstrated in this paper. A targeted drug-delivery system was developed for selective killing of breast cancer cells with polyethylene glycol biofunctionalized and DOX-loaded SWCNTs conjugated with folic acid. In our work, in vitro drug-release studies showed that the drug (DOX binds at physiological pH (pH 7.4 and is released only at a lower pH, ie, lysosomal pH (pH 4.0, which is the characteristic pH of the tumor environment. A sustained release of DOX from the SWCNTs was observed for a period of 3 days. SWCNTs have strong optical absorbance in the near-infrared (NIR region. In this special spectral window, biological systems are highly transparent. Our study reports that under laser irradiation at 800 nm, SWCNTs exhibited strong light–heat transfer characteristics. These optical properties of SWCNTs open the way for selective photothermal ablation in cancer therapy. It was also observed that internalization and uptake of folate-conjugated NTs into cancer cells was achieved by a receptor-mediated endocytosis mechanism. Results of the in vitro experiments show that laser was effective in destroying the cancer cells, while sparing the normal cells. When the above laser effect was combined with DOX-conjugated SWCNTs, we found enhanced and accelerated killing of breast cancer cells. Thus, this nanodrug-delivery system, consisting of laser, drug, and SWCNTs, looks to be a promising selective modality with high treatment efficacy and low side effects for cancer therapy. Keywords: cancer, nanotherapy, SWCNTs, targeted drug delivery

  12. Dendritic cells engineered to express defined allo-HLA peptide complexes induce antigen-specific cytotoxic T cells efficiently killing tumour cells

    DEFF Research Database (Denmark)

    Stronen, E; Abrahamsen, I W; Gaudernack, G;

    2009-01-01

    presented by a non-self human leucocyte antigen (HLA) molecule and transferred to cancer patients expressing that HLA molecule. Obtaining allo-restricted CTL of high-avidity and low cross-reactivity has, however, proven difficult. Here, we show that dendritic cells transfected with mRNA encoding HLA-A*0201...... and efficiently killed HLA-A*0201(+) melanoma cells, whilst sparing HLA-A*0201(+) B-cells. Allo-restricted CTL specific for peptides from the leukaemia-associated antigens CD33 and CD19 were obtained with comparable efficiency. Collectively, the results show that dendritic cells engineered to express defined allo......Most tumour-associated antigens (TAA) are non-mutated self-antigens. The peripheral T cell repertoire is devoid of high-avidity TAA-specific cytotoxic T lymphocytes (CTL) due to self-tolerance. As tolerance is major histocompatibility complex-restricted, T cells may be immunized against TAA...

  13. Ag nanoparticles generated using bio-reduction and -coating cause microbial killing without cell lysis.

    Science.gov (United States)

    Gade, Aniket; Adams, Joshua; Britt, David W; Shen, Fen-Ann; McLean, Joan E; Jacobson, Astrid; Kim, Young-Cheol; Anderson, Anne J

    2016-04-01

    Cost-effective "green" methods of producing Ag nanoparticles (NPs) are being examined because of the potential of these NPs as antimicrobials. Ag NPs were generated from Ag ions using extracellular metabolites from a soil-borne Pythium species. The NPs were variable in size, but had one dimension less than 50 nm and were biocoated; aggregation and coating changed with acetone precipitation. They had dose-dependent lethal effects on a soil pseudomonad, Pseudomonas chlororaphis O6, and were about 30-fold more effective than Ag(+) ions. A role of reactive oxygen species in cell death was demonstrated by use of fluorescent dyes responsive to superoxide anion and peroxide accumulation. Also mutants of the pseudomonad, defective in enzymes that protect against oxidative stress, were more sensitive than the wild type strain; mutant sensitivity differed between exposure to Ag NPs and Ag(+) ions demonstrating a nano-effect. Imaging of bacterial cells treated with the biocoated Ag NPs revealed no cell lysis, but there were changes in surface properties and cell height. These findings support that biocoating the NPs results in limited Ag release and yet they retained potent antimicrobial activity.

  14. NY-ESO-1 致敏树突状细胞诱导的细胞毒性 T 淋巴细胞对视网膜母细胞瘤细胞的特异性杀伤作用%The killing effect of NY-ESO-1-sensitzed dendritic cells-induced cellular toxic lymphocyte on human retinoblastoma cell

    Institute of Scientific and Technical Information of China (English)

    周学智; 周希瑗

    2012-01-01

    (54.2%),CD83(39.7%)and CD86 (94.8%).The CTL that was induced by DCs-sensitized with NY-ESO-1 specifically killed HXO-RB44 cells.CTL induced by the sensitized DCs had a stronger cytotoxic effect against HXO-RB44 cells compared with un-sensitized DCs and CTL un-induced with DCs,as shown by MTT asssay(P<0.05).The anti-tumor activity was highest when the ratio of effector to target was 75∶1(P<0.05).Conclusions DCs transfected by the recombinant plasmid pDC316/NY-ESO-1 can induce the proliferation of allogenic CTLs,which showed a specific anti-tumor effect against HXO-RB44 cells.These results present a new type of immunotherapy for the treatment of RB.%背景 目前用细胞免疫的方法治疗视网膜母细胞瘤(RB)成为新的研究热点,肿瘤-睾丸抗原是最具免疫原性的人类肿瘤抗原之一,已用于全身一些肿瘤的免疫治疗,但其对RB治疗作用的研究报道较少.目的 探讨肿瘤-睾丸抗原的New York-esophageal-1(NY-ESO-1)致敏树突状细胞(DCs)诱导特异性细胞毒性T淋巴细胞(CTL)对RB细胞株HXO-RB44的杀伤作用.方法 用聚合酶链反应(PCR)技术对NY-ESO-1质粒的目的基因片段进行扩增,使用SalI限制酶和EcoRI限制酶进行双酶切,切胶回收DNA片段.将其连接入pDC316质粒,构建pDC316/NY-ESO-1真核表达载体并再进行酶切鉴定.免疫荧光及Western blot 法检测NY-ESO-1蛋白在HXO-RB44细胞株中的表达.Ficoll法分离血获得单个核细胞,通过RPMI 1640重悬,调整细胞密度为1×106个/ml,通过重组人集落刺激因子(rhGM-CSF)和重组人白细胞介素4(rhIL-4)诱导培养外周血来源的DCs,通过重组质粒转染DCs,以脂多糖诱导DCs成熟,DCs与淋巴细胞数量比为1∶25的比例混合培养HXO-RB44细胞,通过MTT法检测CTL的增生情况.在培养液中同时加入CTL细胞,MTT法检测HXO-RB44细胞的活力.结果 重组质粒pDC316/NY-ESO-1的目的片段序列与NY-ESO-1基因序列完全相同,酶切

  15. Bacterial-killing effect of atmospheric pressure non-equilibrium plasma jet and oral mucosa response.

    Science.gov (United States)

    Liu, Dexi; Xiong, Zilan; Du, Tianfeng; Zhou, Xincai; Cao, Yingguang; Lu, Xinpei

    2011-12-01

    Recently, plasma sterilization has attracted increasing attention in dental community for the atmospheric pressure non-equilibrium plasma jet (APNPs), which is driven by a kilohertz pulsed DC power, may be applied to the dental and oral diseases. However, it is still in doubt whether APNPs can effectively kill pathogenic bacteria in the oral cavity and produce no harmful effects on normal oral tissues, especially on normal mucosa. The aim of this study was to evaluate the bacterial-killing effect of APNPs in the biofilms containing a single breed of bacteria (Porphyromonas gingivalis, P.g.), and the pathological changes of the oral mucosa after treatment by APNPs. P.g. was incubated to form the biofilms in vitro, and the samples were divided into three groups randomly: group A (blank control); group B in which the biofilms were treated by APNPs (the setting of the equipment: 10 kHz, 1600 ns and 8 kV); group C in which the biofilms were exposed only to a gas jet without ignition of the plasma. Each group had three samples and each sample was processed for up to 5 min. The biofilms were then fluorescently stained, observed and photographed under a laser scanning confocal microscope. In the animal experiment, six male Japanese white rabbits were divided into two groups randomly (n=3 in each group) in terms of the different post-treatment time (1-day group and 5-day group). The buccal mucosa of the left side and the mucosa of the ventral surface of the tongue were treated by APNPs for 10 min in the same way as the bacterial biofilm experiment in each rabbit, and the corresponding mucosa of the other sides served as normal control. The clinical manifestations of the oral mucosa were observed and recorded every day. The rabbits were sacrificed one or five day(s) after APNPs treatment. The oral mucosa were harvested and prepared to haematoxylin and eosin-stained sections. Clinical observation and histopathological scores were used to assess mucosal changes. The results

  16. Construction of recombinant adenovirus carrying HSV-TK controlled by hMAM enhancer and promoter and its targeted killing effect on human breast cancer cells%hMAM-EP调控HSV-TK腺病毒载体的构建及其对乳腺癌细胞的靶向杀伤

    Institute of Scientific and Technical Information of China (English)

    于建刚; 吴金香; 庞春秀; 林德馨

    2012-01-01

    breast cancer. Methods: Two recombinant plasmid vectors, hMAM-EP-EGFP and hMAM-EP-TK, were constructed, which respectively carried reporter gene EGFP and suicide gene HSV-TK at the downstream of hMAM-EP. Recombinant adenovirus vectors Ad-EP-EGFP and Ad-EP-TK were obtained after the target genes from the recombinant plasmids were transferred into adenovirus skeleton cosmid pAxcwit2; recombinant adenovirus vectors Ad-EP-EGFP and Ad-EP-TK were then transfected into breast cancer T-47D cells, ZR-75-30 cells and nasopharyngeal cancer 5-8F cells. The expression of EGFP was observed under a fluorescence microscope. Recombinant adenovirus AdEP-TK-infected T-47D cells were cultured with 1 , 10, 20 and 50μg/ml prodrug GCV to observe specific cell-killing effect on breast cancer cells. Results: The recombinant plasmid vectors Ad-EP-EGFP and Ad-EP-TK controlled by hMAM-EP were successfully constructed. EGFP could be observed in human breast cancer T-47D cells infected with Ad-EP-EGFP recombinant adenovirus, and could not be detected in ZR-75-30 and 5-8F cells. Compared with un-infected and Ad-EP-EGFP-infected groups, the survival rate of T-47D cells in Ad-EP-EGFP-infection combined with GCV (50 jig/ml) group was significantly decreased ( [35. 69 ±0. O7]% vs [91. 74 ±0.02]% , [87.69 ±0. 11 ] % , P<0.05). With an increase in mass concentration of GCV, the survival rate decreased. Cell survival rates were (94. 34 ± 0. 04)% , (86. 26 ±0.02)%, (66.51 ±0.09)% and (35.69 ±0.07)% when T47D cells were infected with hMAM-EP-TK in a MOI of 100 and cultured with 1, 10, 20, and 50 |xg/ml GCV. HSV-TK suicide gene controlled by hMAM-EP is specifically expressed in breast cancer T-47D cells, and T-47D cells can be killed by Ad-EP-TK combined with GCV.

  17. Cell killing by simian virus 40: impairment of membrane formation and function.

    Science.gov (United States)

    Norkin, L C

    1977-03-01

    Simian virus 40 infection of the CV-1 line of green monkey kidney cells results in the release of mitochondrial malic dehydrogenase as early as 24 h. Released malic dehydrogenase is detected in the cytoplasm prior to its appearance in the overlay medium. Infected cells lose the ability to consume oxygen between 48 and 56 h, and damage to the elctron transport system is indicated. Nevertheless, cellular ATP levels remain high as late as 72 h. Infection leads to a stimulation of membrane phospholipid synthesis, which reaches a peak at about 32 h. This is followed by a severe decline in new membrane synthesis, which correlates in time with the release of cytoplasmic lactic dehydrogenase into the overlay media. Lactic dehydrogenase release precedes the accumulation of trypan blue-stainable cells by about 6 h. Infection had no effect on the turnover of prelabeled membrane phospholipids. An early simian virus 40 mutant, tsA58, and a late mutant, tsB11, are both less effective than wild-type virus at causing reduced levels of phospholipid synthesis, enzyme release, and the accumulation of trypan blue-stainable cells. Another late mutant, tsB8, is similar to wild-type virus in these respects. At 64 h, there is no detectable cell-associated lactic dehydrogenase and nearly all the cells are trypan blue stainable. Nevertheless, at concentrations of deoxyglucose in the medium below the transport Km, deoxyglucose uptake was similar in infected and control cultures. With higher concentrations of deoxyglucose in the medium, uptake by the infected cultures exceeded that by the control cultures.

  18. 低温等离子体对不同恶性肿瘤细胞的杀伤作用及其机制研究%Killing effect and its mechanism of low-temperature plasma on different human cancer cell lines

    Institute of Scientific and Technical Information of China (English)

    王利利; 秦颂兵; 徐晓婷; 胡超; 钱丹琪; 叶超; 周菊英

    2016-01-01

    目的:探讨低温等离子体( LTP )对人肝癌细胞株 HepG2、非小细胞肺癌细胞株A549和人子宫颈癌细胞株HeLa的杀伤作用及其可能作用机制。方法采用四甲基偶氮唑蓝法测定LTP对人肝癌细胞株 HepG2、非小细胞肺癌细胞株A549和人子宫颈癌细胞株HeLa的增殖抑制作用,透射电子显微镜下观察LTP对HepG2细胞、A549细胞和HeLa细胞的超微结构改变的作用,Muse细胞分析仪检测细胞凋亡,Western blot检测凋亡相关蛋白的变化。结果 HepG2细胞、HeLa细胞和A549细胞存活率为50%时,LTP 的照射时间分别为107、121和127 s。 LTP 对HepG2细胞、A549细胞和HeLa细胞的超微结构均有不同程度的破坏,表现为细胞核和细胞器碎裂。 HepG2细胞、A549细胞和HeLa细胞经LTP处理(处理时间为细胞存活率为50%时照射时间的1/6)后24 h,与对照组比较,凋亡率增加( P<0.05)。 Western blot检测结果显示,抗凋亡蛋白Bcl⁃2蛋白表达下调,促凋亡蛋白Bax蛋白的表达增高,DNA修复蛋白X 线修复交错互补基因1蛋白表达下调。结论 LTP 对HepG2细胞株、A549细胞株和HeLa细胞株均有明显的杀伤作用,可能与诱导细胞凋亡和抑制DNA修复有关。%Objective To investigate the killing effect of low⁃temperature plasma ( LTP ) on HepG2, A549 and HeLa cell lines and explore its possible mechanism. Methods The inhibitory effect of LTP on the proliferation of HepG2, A549 and HeLa cells was determined by MTT assay. Transmission electron microscopy was used to observe the ultrastructural changes of HepG2, A549 and HeLa cells treated with LTP. Cell apoptosis was detected by Muse cytometry. Western blot was used to detect the expression of apoptosis⁃related proteins. Results The survival rates of LTP⁃irradiated HepG2 cells (irradiated for 107 s), HeLa cells ( irradiated for 121 s ) and A549 cells ( irradiated for 127 s ) were 50%. LTP destroyed

  19. Surface chemistry influences cancer killing effect of TiO2 nanoparticles.

    Science.gov (United States)

    Thevenot, Paul; Cho, Jai; Wavhal, Dattatray; Timmons, Richard B; Tang, Liping

    2008-09-01

    Photocatalyzed titanium dioxide (TiO2) nanoparticles have been shown to eradicate cancer cells. However, the required in situ introduction of ultraviolet light limits the use of such a therapy in humans. In the present study the nonphotocatalytic anticancer effect of surface-functionalized TiO2 was examined. Nanoparticles bearing -OH, -NH(2), or -COOH surface groups were tested for their effect on in vitro survival of several cancer and control cell lines. The cells tested included B16F10 melanoma, Lewis lung carcinoma, JHU prostate cancer cells, and 3T3 fibroblasts. Cell viability was observed to depend on particle concentrations, cell types, and surface chemistry. Specifically, -NH(2) and -OH groups showed significantly higher toxicity than -COOH. Microscopic and spectrophotometric studies revealed nanoparticle-mediated cell membrane disruption leading to cell death. The results suggest that functionalized TiO2, and presumably other nanoparticles, can be surface-engineered for targeted cancer therapy.

  20. Melatonin Protects Human Cells from Clustered DNA Damages, Killing and Acquisition of Soft Agar Growth Induced by X-rays or 970 MeV/n Fe ions

    Energy Technology Data Exchange (ETDEWEB)

    Das, B.; Sutherland, B.; Bennett, P. V.; Cutter, N. C.; Sutherland, J. C.

    2011-06-01

    We tested the ability of melatonin (N-acetyl-5 methoxytryptamine), a highly effective radical scavenger and human hormone, to protect DNA in solution and in human cells against induction of complex DNA clusters and biological damage induced by low or high linear energy transfer radiation (100 kVp X-rays, 970 MeV/nucleon Fe ions). Plasmid DNA in solution was treated with increasing concentrations of melatonin (0.0-3.5 mM) and were irradiated with X-rays. Human cells (28SC monocytes) were also irradiated with X-rays and Fe ions with and without 2 mM melatonin. Agarose plugs containing genomic DNA were subjected to Contour Clamped Homogeneous Electrophoretic Field (CHEF) followed by imaging and clustered DNA damages were measured by using Number Average length analysis. Transformation experiments on human primary fibroblast cells using soft agar colony assay were carried out which were irradiated with Fe ions with or without 2 mM melatonin. In plasmid DNA in solution, melatonin reduced the induction of single- and double-strand breaks. Pretreatment of human 28SC cells for 24 h before irradiation with 2 mM melatonin reduced the level of X-ray induced double-strand breaks by {approx}50%, of abasic clustered damages about 40%, and of Fe ion-induced double-strand breaks (41% reduction) and abasic clusters (34% reduction). It decreased transformation to soft agar growth of human primary cells by a factor of 10, but reduced killing by Fe ions only by 20-40%. Melatonin's effective reduction of radiation-induced critical DNA damages, cell killing, and striking decrease of transformation suggest that it is an excellent candidate as a countermeasure against radiation exposure, including radiation exposure to astronaut crews in space travel.

  1. Engineered metal nanoparticles in the sub-nanomolar levels kill cancer cells

    Directory of Open Access Journals (Sweden)

    Vodyanoy V

    2016-04-01

    Full Text Available Vitaly Vodyanoy,1 Yasmine Daniels,2 Oleg Pustovyy,1 William A MacCrehan,2 Shin Muramoto,2 Gheorghe Stan21Department of Anatomy, Physiology and Pharmacology, Auburn University College of Veterinary Medicine, Auburn, AL, 2Material Measurement Laboratory, National Institute of Standards and Technology, Gaithersburg, MA, USA Background: Small metal nanoparticles obtained from animal blood were observed to be toxic to cultured cancer cells, whereas noncancerous cells were much less affected. In this work, engineered zinc and copper metal nanoparticles were produced from bulk metal rods by an underwater high-voltage discharge method. The metal nanoparticles were characterized by atomic force microscopy and X-ray photoelectron spectroscopy. The metal nanoparticles, with estimated diameters of 1 nm–2 nm, were determined to be more than 85% nonoxidized. A cell viability assay and high-resolution light microscopy showed that exposure of RG2, cultured rat brain glioma cancer cells, to the zinc and copper nanoparticles resulted in cell morphological changes, including decreased cell adherence, shrinking/rounding, nuclear condensation, and budding from cell bodies. The metal-induced cell injuries were similar to the effects of staurosporine, an active apoptotic reagent. The viability experiments conducted for zinc and copper yielded values of dissociation constants of 0.22±0.08 nmol/L (standard error [SE] and 0.12±0.02 nmol/L (SE, respectively. The noncancerous astrocytes were not affected at the same conditions. Because metal nanoparticles were lethal to the cancer cells at sub-nanomolar concentrations, they are potentially important as nanomedicine.Purpose: Lethal concentrations of synthetic metal nanoparticles reported in the literature are a few orders of magnitude higher than the natural, blood-isolated metal nanoparticles; therefore, in this work, engineered metal nanoparticles were examined to mimic the properties of endogenous metal

  2. HAMLET kills tumor cells by an apoptosis-like mechanism--cellular, molecular, and therapeutic aspects.

    Science.gov (United States)

    Svanborg, Catharina; Agerstam, Helena; Aronson, Annika; Bjerkvig, Rolf; Düringer, Caroline; Fischer, Walter; Gustafsson, Lotta; Hallgren, Oskar; Leijonhuvud, Irene; Linse, Sara; Mossberg, Ann-Kristin; Nilsson, Hanna; Pettersson, Jenny; Svensson, Malin

    2003-01-01

    HAMLET (human alpha-lactalbumin made lethal to tumor cells) is a protein-lipid complex that induces apoptosis-like death in tumor cells, but leaves fully differentiated cells unaffected. This review summarizes the information on the in vivo effects of HAMLET in patients and tumor models on the tumor cell biology, and on the molecular characteristics of the complex. HAMLET limits the progression of human glioblastomas in a xenograft model and removes skin papillomas in patients. This broad anti-tumor activity includes >40 different lymphomas and carcinomas and apoptosis is independent of p53 or bcl-2. In tumor cells HAMLET enters the cytoplasm, translocates to the perinuclear area, and enters the nuclei where it accumulates. HAMLET binds strongly to histones and disrupts the chromatin organization. In the cytoplasm, HAMLET targets ribosomes and activates caspases. The formation of HAMLET relies on the propensity of alpha-lactalbumin to alter its conformation when the strongly bound Ca2+ ion is released and the protein adopts the apo-conformation that exposes a new fatty acid binding site. Oleic acid (C18:1,9 cis) fits this site with high specificity, and stabilizes the altered protein conformation. The results illustrate how protein folding variants may be beneficial, and how their formation in peripheral tissues may depend on the folding change and the availability of the lipid cofactor. One example is the acid pH in the stomach of the breast-fed child that promotes the formation of HAMLET. This mechanism may contribute to the protective effect of breastfeeding against childhood tumors. We propose that HAMLET should be explored as a novel approach to tumor therapy.

  3. Cocoa Extract Indicated Has Activity on Selectively Killing Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ariza Budi Tunjung Sari

    2015-09-01

    Full Text Available Effect of the cocoa crude extract on mortality of breast cancer cell lines i.e. MCF-7, T47D and normal cell (Vero, was observed. Crude cocoa extract prepared from a freshly dried cocoa bean that was containing 14% catechin and 0.6% caffeine. Catechin and caffeine content were modulated to 2-folds (28% catechin or 1.2% caffeine and 3-folds (42% catechin or 1.8% caffeine by adding pure compounds. Extracts were dissolved in dimethylsulfoxide (DMSO at concentrations ranging from 200 to 1600 μg/ml. The positive control was doxorubicin (0.5-16 μg/ml in DMSO. Cell lines (MCF-7, T47D, and Vero were incubated in test sample for 24h at 37°, prior to 3-(4,4-dimetylthiazole-2-yl-2,5-diphenyltetrazolium bromide (MTT assay. The absorbance of each well was measured at 550 nm, and lethal concentration (LC50 was calculated. The cocoa extract induced mortality of breast cancer cell lines but not in Vero cells. The effect on MCF-7 was greater than on T47D, given the LC50 was 1236 μg/ml (MCF-7 and 1893 μg/ml (T47D. Cytotoxic potential of cocoa extract was much lower than doxorubicin whose LC50 was 0,777 μg/ml (MCF-7 and 0,082 μg/ml (T47D. Increasing catechin content to 2-folds did not significantly affect LC50 value, but 3-folds catechin content reduced LC50 to 1021 μg/ml. Meanwhile increasing caffeine content to 2-folds significantly reduced LC50 to 750 μg/ml, however, 3-fold content resulted in slightly higher LC50 at 780 μg/ml. This indicates that cocoa extract have anti-cancer potential, and purification may improve this property.

  4. Effects of Rhodomyrtus tomentosa Extract on Killing Activity of Human Neutrophils and Membrane Integrity of Enterohaemorrhagic Escherichia coli O157:H7.

    Science.gov (United States)

    Hmoteh, Jutharat; Syed Musthafa, Khadar; Pomwised, Rattanaruji; Voravuthikunchai, Supayang Piyawan

    2016-05-27

    Enterohaemorrhagic Escherichia coli (E. coli) O157:H7 is one of the most virulent causative agents of foodborne disease. Use of antibiotics for the treatment against E. coli O157:H7 infection leads to hemolytic uremic syndrome. The present study evaluated the potential of ethanolic leaf extract of a medicinal plant, Rhodomyrtus tomentosa in enhancing the killing activity of human neutrophils against E. coli O157:H7. In addition, the effects of the extract on membrane permeability of the organisms were studied. In the killing assay, percentage survival of the bacterial cells after being exposed to human neutrophils in the presence of various concentrations of the extract were determined. At 45 min, percentage survival of E. coli O157:H7 and E. coli ATCC 25922 after treated with neutrophils in the presence of the extract at 125-250 µg/mL was 58.48%-50.28% and 69.13%-35.35%, respectively. Furthermore, upon treatment with R. tomentosa at 250 µg/mL uptake of crystal violet by E. coli O157:H7 and E. coli ATCC 25922 was increased to 40.07% and 36.16%, respectively. Therefore, it is suggested that the extract exhibited dual effects as immunostimulant and membrane permeabilizing agent perhaps resulted in enhancing the killing activity of neutrophils against the organisms.

  5. Effects of Rhodomyrtus tomentosa Extract on Killing Activity of Human Neutrophils and Membrane Integrity of Enterohaemorrhagic Escherichia coli O157:H7

    Directory of Open Access Journals (Sweden)

    Jutharat Hmoteh

    2016-05-01

    Full Text Available Enterohaemorrhagic Escherichia coli (E. coli O157:H7 is one of the most virulent causative agents of foodborne disease. Use of antibiotics for the treatment against E. coli O157:H7 infection leads to hemolytic uremic syndrome. The present study evaluated the potential of ethanolic leaf extract of a medicinal plant, Rhodomyrtus tomentosa in enhancing the killing activity of human neutrophils against E. coli O157:H7. In addition, the effects of the extract on membrane permeability of the organisms were studied. In the killing assay, percentage survival of the bacterial cells after being exposed to human neutrophils in the presence of various concentrations of the extract were determined. At 45 min, percentage survival of E. coli O157:H7 and E. coli ATCC 25922 after treated with neutrophils in the presence of the extract at 125–250 µg/mL was 58.48%–50.28% and 69.13%–35.35%, respectively. Furthermore, upon treatment with R. tomentosa at 250 µg/mL uptake of crystal violet by E. coli O157:H7 and E. coli ATCC 25922 was increased to 40.07% and 36.16%, respectively. Therefore, it is suggested that the extract exhibited dual effects as immunostimulant and membrane permeabilizing agent perhaps resulted in enhancing the killing activity of neutrophils against the organisms.

  6. 外磁场协同卟啉-葡聚糖磁性纳米微粒的光动力学对人膀胱癌细胞的杀伤作用%Synergetic killing effects of external magnetic fields combined with porphyrin-dextran magnetic nanoparticles on the human bladder cancer cells

    Institute of Scientific and Technical Information of China (English)

    罗道升; 米其武; 孟祥军; 高勇; 戴宇平; 邓春华

    2012-01-01

    目的:探讨外磁场协同卟啉-葡聚糖磁性纳米微粒(porphyrin-dextran magnetic nanoparticles,PDMN)的光动力学作用对人膀胱癌细胞杀伤作用的影响.方法:用化学共沉淀和氧化还原法制备PDMN,并检测其理化性质.采用四甲基偶氮唑蓝(methyl thiazolyl tetrazolium,MTT)法、流式细胞术分别观察脉冲电磁场协同PDMN的光动力学作用对人膀胱癌BIU-87细胞的杀伤作用.结果:PDMN的直径、比饱和磁化强度(saturation magnetization,Ms)分别为10 ~ 15 nm、0.20 emu/g,有效粒径为94.8nm.PDMN光动力学作用可显著抑制BIU-87细胞增值,抑制率达(17.61±2.73)%,并诱导BIU-87细胞发生明显的凋亡,凋亡率为(24.53 +5.74)%.脉冲外电磁场(5 mT)也可抑制BIU-87细胞增殖和诱导发生凋亡,且协同PDMN的光动力学作用对BIU-87细胞的生长抑制和诱导肿瘤细胞凋亡的效应均显著增强,抑制率和凋亡率分别为(28.11±4.25)%、(24.53±5.74)%,明显高于其他各组(P<0.01).结论:化学修饰后的PDMN的光动力学作用能有效地抑制人膀胱癌BIU-87细胞的增殖并诱导其凋亡,与恒定外磁场协同作用对BIU-87细胞的杀伤作用更强.%Objective: To study the synergetic killing effects of external magnetic fields combined with the photodynamic action of porphyrin-dextran iron oxide magnetic nanoparticles ( PDMN) on human bladder cancer cells in vitro. Methods: The PDMN were produced by using the chemical co-precipitation and redox process and the physicochemical properties were characterized. Methyl thiazolyl tetrazolium ( MTT) and flow cytometry were used to determine the effects of photodynamic therapy of PDMN combined with external pulsed electromagnetic fields (5 mT) on killing human bladder cancer BIU-87 cells respectively. Results: The diameters of PDMN were 10 - 15 nm and the saturation magnetization was 0. 20 emu/g. Effective diameter of PDMN was 94. 8 nm. PDMN could remarkably inhibit the proliferation and

  7. Resveratrol and arsenic trioxide act synergistically to kill tumor cells in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Xiao-Yan Zhao

    Full Text Available BACKGROUND AND AIMS: Arsenic trioxide (As2O3, which used as an effective agent in the treatment of leukaemia and other solid tumors, is largely limited by its toxicity. QT prolongation, torsades de pointes and sudden heart death have been implicated in the cardiotoxicity of As2O3. The present study was designed to explore whether the combination of As2O3 and resveratrol could generate a more powerful anti-cancer effect both in vitro and in vivo. MATERIALS AND METHODS: MTT assay was performed to assess the proliferation of Hela, MCF-7 and NB4 cells. Isobolographic analysis was used to evaluate combination index values from cell viability data. The apoptosis and the cellular reactive oxygen species (ROS level were assessed by fluorescent microscopy and flow cytometry separately in vitro. The effect of As2O3, alone and in combination with resveratrol on Hela tumor growth in an orthotopic nude mouse model was also investigated. The tumor volume and the immunohistochemical analysis of CD31, CD34 and VEGF were determined. RESULTS: Resveratrol dramatically enhanced the anti-cancer effect induced by As2O3 in vitro. In addition, isobolographic analysis further demonstrated that As2O3 and resveratrol generated a synergistic action. More apoptosis and ROS generation were observed in the combination treatment group. Similar synergistic effects were found in nude mice in vivo. The combination of As2O3 and resveratrol dramatically suppressed both tumor growth and angiogenesis in nude mice. CONCLUSIONS: Combining As2O3 with resveratrol would be a novel strategy to treat cancer in clinical practice.

  8. D-polyglutamine amyloid recruits L-polyglutamine monomers and kills cells

    Science.gov (United States)

    Kar, Karunakar; Arduini, Irene; Drombosky, Kenneth W.; van der Wel, Patrick C. A.; Wetzel, Ronald

    2014-01-01

    Polyglutamine (polyQ) amyloid fibrils are observed in disease tissue and have been implicated as toxic agents responsible for neurodegeneration in expanded CAG repeat diseases like Huntington’s disease (HD). Despite intensive efforts, the mechanism of amyloid toxicity remains unknown. As a novel approach to probing polyQ toxicity, we investigate here how some cellular and physical properties of polyQ amyloid vary with the chirality of the glutamine residues in the polyQ. We challenged PC12 cells with small amyloid fibrils composed of either L- or D-polyQ peptides and found that D-fibrils are as cytotoxic as L-fibrils. We also found using fluorescence microscopy that both aggregates effectively seed the aggregation of cell-produced L-polyQ proteins, suggesting a surprising lack of stereochemical restriction in seeded elongation of polyQ amyloid. To investigate this effect further, we studied chemically synthesized D- and L-polyQ in vitro. We found that, as expected, D-polyQ monomers are not recognized by proteins that recognize L-polyQ monomers. However, amyloid fibrils prepared from D-polyQ peptides can efficiently seed the aggregation of L-polyQ monomers in vitro, and vice versa. This result is consistent with our cell results on polyQ recruitment, but is inconsistent with previous literature reports on the chiral specificity of amyloid seeding. This chiral cross-seeding can be rationalized by a model for seeded elongation featuring a “rippled β-sheet” interface between seed fibril and docked monomers of opposite chirality. The lack of chiral discrimination in polyQ amyloid cytotoxicity is consistent with several toxicity mechanisms, including recruitment of cellular polyQ proteins. PMID:24291210

  9. Introducing the RadBioStat Educational Software: Computer-Assisted Teaching of the Random Nature of Cell Killing

    Directory of Open Access Journals (Sweden)

    Safari A

    2014-06-01

    Full Text Available The interaction of radiation with cells and tissues has a random nature. Therefore, understanding the random nature of cell killing that is determined by Poisson distribution statistics is an essential point in education of radiation biology. RadBioStat is a newly developed educational MATLAB-based software designed for computer-assisted learning of the target theory in radiation biology. Although its potential applications is developing rapidly, currently RadBioStat software can be a useful tool in computerassisted education of radiobiological models such as single target single hit, multiple target single hit and multiple target multiple hit. Scholars’ feedback is valuable to the producers of this software and help them continuously improve this product, add new features and increase its desirability and functionality.

  10. Safety of the recombinant cholera toxin B subunit, killed whole-cell (rBS-WC oral cholera vaccine in pregnancy.

    Directory of Open Access Journals (Sweden)

    Ramadhan Hashim

    Full Text Available INTRODUCTION: Mass vaccinations are a main strategy in the deployment of oral cholera vaccines. Campaigns avoid giving vaccine to pregnant women because of the absence of safety data of the killed whole-cell oral cholera (rBS-WC vaccine. Balancing this concern is the known higher risk of cholera and of complications of pregnancy should cholera occur in these women, as well as the lack of expected adverse events from a killed oral bacterial vaccine. METHODOLOGY/PRINCIPAL FINDINGS: From January to February 2009, a mass rBS-WC vaccination campaign of persons over two years of age was conducted in an urban and a rural area (population 51,151 in Zanzibar. Pregnant women were advised not to participate in the campaign. More than nine months after the last dose of the vaccine was administered, we visited all women between 15 and 50 years of age living in the study area. The outcome of pregnancies that were inadvertently exposed to at least one oral cholera vaccine dose and those that were not exposed was evaluated. 13,736 (94% of the target women in the study site were interviewed. 1,151 (79% of the 1,453 deliveries in 2009 occurred during the period when foetal exposure to the vaccine could have occurred. 955 (83% out of these 1,151 mothers had not been vaccinated; the remaining 196 (17% mothers had received at least one dose of the oral cholera vaccine. There were no statistically significant differences in the odds ratios for birth outcomes among the exposed and unexposed pregnancies. CONCLUSIONS/SIGNIFICANCE: We found no statistically significant evidence of a harmful effect of gestational exposure to the rBS-WC vaccine. These findings, along with the absence of a rational basis for expecting a risk from this killed oral bacterial vaccine, are reassuring but the study had insufficient power to detect infrequent events. TRIAL REGISTRATION: ClinicalTrials.gov NCT00709410.

  11. Enhanced killing of breast cancer cells by a d-amino acid analog of the winter flounder-derived pleurocidin NRC-03.

    Science.gov (United States)

    Hilchie, Ashley L; Haney, Evan F; Pinto, Devanand M; Hancock, Robert E W; Hoskin, David W

    2015-12-01

    Cationic antimicrobial peptides (CAPs) defend against pathogens and, in some cases, exhibit potent anticancer activities. We previously reported that the pleurocidin NRC-03 causes lysis of breast cancer and multiple myeloma cells. NRC-03 also reduces the EC50 of other cytotoxic compounds and prevents tumor growth in vivo. However, the therapeutic utility of NRC-03 may be limited by its susceptibility to degradation by proteases. The goal of this study was to characterize the anticancer activities of a d-amino acid analog of NRC-03 ([D]-NRC-03) that was predicted to be resistant to proteolytic degradation. Unlike NRC-03, [D]-NRC-03 was not degraded by human serum or trypsin and, in comparison to NRC-03, showed increased killing of breast cancer cells, including multidrug-resistant cells; however, [D]-NRC-03 was somewhat more cytotoxic than NRC-03 for several types of normal cells. Importantly, [D]-NRC-03 was more effective than NRC-03 in vivo since 4-fold less peptide was required for an equivalent inhibitory effect on the growth of breast cancer cell xenografts in immune-deficient mice. These findings demonstrate that a d-amino acid analog of NRC-03 overcomes a major limitation to the therapeutic use of NRC-03, namely peptide stability. Further modification of [D]-NRC-03 is required to improve its selectivity for cancer cells.

  12. Vine kill interval and temperature effects on Fusarium dry rot development in Russet Burbank

    Science.gov (United States)

    Fusarium dry rot disease development in potato storage is universal to all market sectors and regions. The objective of this 2-year study was to evaluate three possible management decisions that may impact Fusarium dry rot development in storage: a) vine kill to harvest time, b) harvested tuber pulp...

  13. In vivo label-free photoacoustic flow cytography and on-the-spot laser killing of single circulating melanoma cells

    Science.gov (United States)

    He, Yun; Wang, Lidai; Shi, Junhui; Yao, Junjie; Li, Lei; Zhang, Ruiying; Huang, Chih-Hsien; Zou, Jun; Wang, Lihong V.

    2016-12-01

    Metastasis causes as many as 90% of cancer-related deaths, especially for the deadliest skin cancer, melanoma. Since hematogenous dissemination of circulating tumor cells is the major route of metastasis, detection and destruction of circulating tumor cells are vital for impeding metastasis and improving patient prognosis. Exploiting the exquisite intrinsic optical absorption contrast of circulating melanoma cells, we developed dual-wavelength photoacoustic flow cytography coupled with a nanosecond-pulsed melanoma-specific laser therapy mechanism. We have successfully achieved in vivo label-free imaging of rare single circulating melanoma cells in both arteries and veins of mice. Further, the photoacoustic signal from a circulating melanoma cell immediately hardware-triggers a lethal pinpoint laser irradiation to kill it on the spot in a thermally confined manner without causing collateral damage. A pseudo-therapy study including both in vivo and in vitro experiments demonstrated the performance and the potential clinical value of our method, which can facilitate early treatment of metastasis by clearing circulating tumor cells from vasculature.

  14. Troxerutin, a natural flavonoid binds to DNA minor groove and enhances cancer cell killing in response to radiation.

    Science.gov (United States)

    Panat, Niranjan A; Singh, Beena G; Maurya, Dharmendra K; Sandur, Santosh K; Ghaskadbi, Saroj S

    2016-05-05

    Troxerutin, a flavonoid best known for its radioprotective and antioxidant properties is of considerable interest of study due to its broad pharmacological activities. The present study on troxerutin highlights its abilities to bind DNA and enhance cancer cell killing in response to radiation. Troxerutin showed strong binding with calf thymus DNA in vitro. Troxerutin-DNA interaction was confirmed by CD spectropolarimetry. The mode of binding of troxerutin to DNA was assessed by competing troxerutin with EtBr or DAPI, known DNA intercalator and a minor groove binder, respectively. DAPI fluorescence was drastically reduced with linear increase in troxerutin concentration suggesting possible binding of troxerutin to DNA minor groove. Further, computational studies of docking of troxerutin molecule on mammalian DNA also indicated possible troxerutin-DNA interaction at minor groove of DNA. Troxerutin was found to mainly localize in the nucleus of prostate cancer cells. It induced cytotoxicity in radioresistant (DU145) and sensitive (PC3) prostate cancer cells. When troxerutin pre-treated DU145 and PC3 cells were exposed to γ-radiation, cytotoxicity as estimated by MTT assay, was found to be further enhanced. In addition, the % subG1 population detected by propidium iodide staining also showed similar response when combined with radiation. A similar trend was observed in terms of ROS generation and DNA damage in DU145 cells when troxerutin and radiation were combined. DNA binding at minor groove by troxerutin may have contributed to strand breaks leading to increased radiation induced cell death.

  15. In vivo label-free photoacoustic flow cytography and on-the-spot laser killing of single circulating melanoma cells

    Science.gov (United States)

    He, Yun; Wang, Lidai; Shi, Junhui; Yao, Junjie; Li, Lei; Zhang, Ruiying; Huang, Chih-Hsien; Zou, Jun; Wang, Lihong V.

    2016-01-01

    Metastasis causes as many as 90% of cancer-related deaths, especially for the deadliest skin cancer, melanoma. Since hematogenous dissemination of circulating tumor cells is the major route of metastasis, detection and destruction of circulating tumor cells are vital for impeding metastasis and improving patient prognosis. Exploiting the exquisite intrinsic optical absorption contrast of circulating melanoma cells, we developed dual-wavelength photoacoustic flow cytography coupled with a nanosecond-pulsed melanoma-specific laser therapy mechanism. We have successfully achieved in vivo label-free imaging of rare single circulating melanoma cells in both arteries and veins of mice. Further, the photoacoustic signal from a circulating melanoma cell immediately hardware-triggers a lethal pinpoint laser irradiation to kill it on the spot in a thermally confined manner without causing collateral damage. A pseudo-therapy study including both in vivo and in vitro experiments demonstrated the performance and the potential clinical value of our method, which can facilitate early treatment of metastasis by clearing circulating tumor cells from vasculature. PMID:28000788

  16. The radioresistance to killing of A1-5 cells derives from activation of the Chk1 pathway

    Science.gov (United States)

    Hu, B.; Zhou, X. Y.; Wang, X.; Zeng, Z. C.; Iliakis, G.; Wang, Y.

    2001-01-01

    Checkpoints respond to DNA damage by arresting the cell cycle to provide time for facilitating repair. In mammalian cells, the G(2) checkpoint prevents the Cdc25C phosphatase from removing inhibitory phosphate groups from the mitosis-promoting kinase Cdc2. Both Chk1 and Chk2, the checkpoint kinases, can phosphorylate Cdc25C and inactivate its in vitro phosphatase activity. Therefore, both Chk1 and Chk2 are thought to regulate the activation of the G(2) checkpoint. Here we report that A1-5, a transformed rat embryo fibroblast cell line, shows much more radioresistance associated with a much stronger G(2) arrest response when compared with its counterpart, B4, although A1-5 and B4 cells have a similar capacity for nonhomologous end-joining DNA repair. These phenotypes of A1-5 cells are accompanied by a higher Chk1 expression and a higher phosphorylation of Cdc2. On the other hand, Chk2 expression increases slightly following radiation; however, it has no difference between A1-5 and B4 cells. Caffeine or UCN-01 abolishes the extreme radioresistance with the strong G(2) arrest and at the same time reduces the phosphorylation of Cdc2 in A1-5 cells. In addition, Chk1 but not Chk2 antisense oligonucleotide sensitizes A1-5 cells to radiation-induced killing and reduces the G(2) arrest of the cells. Taken together these results suggest that the Chk1/Cdc25C/Cdc2 pathway is the major player for the radioresistance with G(2) arrest in A1-5 cells.

  17. To kill, stay or flee: the effects of lions and landscape factors on habitat and kill site selection of cheetahs in South Africa.

    Science.gov (United States)

    Rostro-García, Susana; Kamler, Jan F; Hunter, Luke T B

    2015-01-01

    Understanding how animals utilize available space is important for their conservation, as it provides insight into the ecological needs of the species, including those related to habitat, prey and inter and intraspecific interactions. We used 28 months of radio telemetry data and information from 200 kill locations to assess habitat selection at the 3rd order (selection of habitats within home ranges) and 4th order (selection of kill sites within the habitats used) of a reintroduced population of cheetahs Acinonyx jubatus in Phinda Private Game Reserve, South Africa. Along with landscape characteristics, we investigated if lion Panthera leo presence affected habitat selection of cheetahs. Our results indicated that cheetah habitat selection was driven by a trade-off between resource acquisition and lion avoidance, and the balance of this trade-off varied with scale: more open habitats with high prey densities were positively selected within home ranges, whereas more closed habitats with low prey densities were positively selected for kill sites. We also showed that habitat selection, feeding ecology, and avoidance of lions differed depending on the sex and reproductive status of cheetahs. The results highlight the importance of scale when investigating a species' habitat selection. We conclude that the adaptability of cheetahs, together with the habitat heterogeneity found within Phinda, explained their success in this small fenced reserve. The results provide information for the conservation and management of this threatened species, especially with regards to reintroduction efforts in South Africa.

  18. To kill, stay or flee: the effects of lions and landscape factors on habitat and kill site selection of cheetahs in South Africa.

    Directory of Open Access Journals (Sweden)

    Susana Rostro-García

    Full Text Available Understanding how animals utilize available space is important for their conservation, as it provides insight into the ecological needs of the species, including those related to habitat, prey and inter and intraspecific interactions. We used 28 months of radio telemetry data and information from 200 kill locations to assess habitat selection at the 3rd order (selection of habitats within home ranges and 4th order (selection of kill sites within the habitats used of a reintroduced population of cheetahs Acinonyx jubatus in Phinda Private Game Reserve, South Africa. Along with landscape characteristics, we investigated if lion Panthera leo presence affected habitat selection of cheetahs. Our results indicated that cheetah habitat selection was driven by a trade-off between resource acquisition and lion avoidance, and the balance of this trade-off varied with scale: more open habitats with high prey densities were positively selected within home ranges, whereas more closed habitats with low prey densities were positively selected for kill sites. We also showed that habitat selection, feeding ecology, and avoidance of lions differed depending on the sex and reproductive status of cheetahs. The results highlight the importance of scale when investigating a species' habitat selection. We conclude that the adaptability of cheetahs, together with the habitat heterogeneity found within Phinda, explained their success in this small fenced reserve. The results provide information for the conservation and management of this threatened species, especially with regards to reintroduction efforts in South Africa.

  19. Killing Cancer Cells with the Help of Infrared Light – Photoimmunotherapy

    Science.gov (United States)

    Near-infrared photoimmunotherapy uses an antibody–photoabsorber conjugate that binds to cancer cells. When near-infrared light is applied, the cells swell and then burst, causing the cancer cell to die. Photoimmunotherapy is in clinical trials in patients with inoperable tumors.

  20. Effectiveness of cleaners and sanitizers in killing Salmonella Newport in the gut of a free-living nematode, Caenorhabditis elegans.

    Science.gov (United States)

    Kenney, Stephen J; Anderson, Gary L; Williams, Phillip L; Millner, Patricia D; Beuchat, Larry R

    2004-10-01

    Caenorhabditis elegans, a free-living nematode found in soil, has been shown to ingest human enteric pathogens, thereby potentially serving as a vector for preharvest contamination of fruits and vegetables. A study was undertaken to evaluate the efficacy of cleaners and sanitizers in killing Salmonella enterica serotype Newport in the gut of C. elegans. Adult worms were fed nalidixic acid-adapted cells of Escherichia coli OP50 (control) or Salmonella Newport for 24 h, washed, placed on paper discs, and incubated at temperatures of 4 or 20 degrees C and relative humidities of 33 or 98% for 24 h. Two commercial cleaners (Enforce and K Foam Lo) and four sanitizers (2% acetic acid, 2% lactic acid, Sanova, and chlorine [50 and 200 microg/ml]) were applied to worms for 0, 2, or 10 min. Populations of E. coli and Salmonella Newport (CFU per worm) in untreated and treated worms were determined by sonicating worms in 0.1% peptone and surface plating suspensions of released cells on tryptic soy agar containing nalidixic acid. Populations of Salmonella Newport in worms exposed to 33 or 98% relative humidity at 4 degrees or 33% relative humidity at 20 degrees C were significantly (P < or = 0.05) lower than the number surviving exposure to 98% relative humidity at 20 degrees C. In general, treatment of desiccated worms with cleaners and sanitizers was effective in significantly (P < or = 0.05) reducing the number of ingested Salmonella Newport. Results indicate that temperature and relative humidity influence the survival of Salmonella Newport in the gut of C. elegans, and cleaners and sanitizers may not eliminate the pathogen.

  1. Radiation related basic cancer research : research for radiation induced tumor cell killing

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Seung Hoon; Hong, Seok Il; Cho, Kyung Ja; Kim, Byung Gi; Lee, Kee Ho; Nam, Myung Jin

    1999-04-01

    The radioresistant clones was established from human U251 glioblastoma cell line through intermittently exposed to 3 Gy gamma-radiation for six months. Treatment of SNU-16 cells with various doses of radiation, TNF alpha and PMA resulted in a decrease in cell viability. The results prove that cell death of SNU16 is a apoptosis mediated by caspase-3. We have examined the expression of bcl-2 and c-myc in cervical cancer specimens and cervical intraepithelial neoplasia (CIN) to determine the role of coexpression of bcl-3 and c-myc during progression into cervical cancer. The frequent alterations in FHIT expression in many cervical carcinomas and their cell lines suggest that FHIT gene alterations are pla a role in cervical tumorigenesis. According to these correlation between the viability and apoptosis of RD cells, the proper range of the dosage for the investigation of differentiation potency in RD cells was assessed as 1 to 3Gy.

  2. Killing of p53-deficient hepatoma cells by parvovirus H-1 and chemotherapeutics requires promyelocytic leukemia protein

    Institute of Scientific and Technical Information of China (English)

    Maike Sieben; Markus Moehler; Kerstin Herzer; Maja Zeidler; Vera Heinrichs; Barbara Leuchs; Martin Schuler; Jan J Cornelis; Peter R Galle; Jean Rommelaere

    2008-01-01

    AIM: To evaluate the synergistic targeting and killing of human hepatocellular carcinoma (HCC) cells lacking p53 by the oncolytic autonomous parvovirus (PV) H-1 and chemotherapeutic agents and its dependence on functional promyelocytic leukemia protein (PML).METHODS: The role of p53 and PML in regulating cytotoxicity and gene transfer mediated by wild-type (wt)PV H-1 were explored in two pairs of isogenic human hepatoma cell lines with different p53 status.Furthermore,H-1 PV infection was combined with cytostatic drug treatment.RESULTS: While the HCC cells with different p53 status studied were all susceptible to H-1 PV-induced apoptosis,the cytotoxicity of H-1 PV was more pronounced in p53-negative than in p53-positive cells.Apoptosis rates in p53-negative cell lines treated by genotoxic drugs were further enhanced by a treatment with H-1 PV.In flow cytometric analyses,H-1 PV infection resulted in a reduction of the mitochondrial transmembrane potential.In addition,H-1 PV cells showed a significant increase in PML expression.Knocking down PML expression resulted in a striking reduction of the level of H-1 PV infected tumor cell death.CONCLUSION: H-1 PV is a suitable agent to circumvent the resistance of p53-negative HCC cells to genotoxic agents,and it enhances the apoptotic process which is dependent on functional PML.Thus,H-1 PV and its oncolytic vector derivatives may be considered as therapeutic options for HCC,particularly for p53-negative tumors.

  3. Working with the effects of traumatic bereavement by uxoricide (spouse killing) on young children's attachment behaviour.

    Science.gov (United States)

    Black, D

    1998-01-01

    This is a clinical description of the work of the Traumatic Stress Clinic, London with children where one parent kills the other, paying special attention to the disorder of attachment which occurs in a majority of the young children. It is suggested that this arises from the combination of the extreme trauma of witnessing the homicide, the frequent changes of carer, and the relative lack of therapeutic help.

  4. Anti-tumor activity of heat-killed Lactobacillus plantarum BF-LP284 on Meth-A tumor cells in BALB/c mice.

    Science.gov (United States)

    Shin, Ryoichi; Itoh, Yukie; Kataoka, Motoyuki; Iino-Miura, Shiori; Miura, Ryosuke; Mizutani, Takeo; Fujisawa, Tomohiko

    2016-09-01

    Probiotics exert numerous effects on human well-being. Here, heat-killed Lactobacillus plantarum BF-LP284 (H-Lp) was isolated as a potent immuno-modulator among 15 strains of lactobacilli in terms of TNF-α induction ability in peritoneal macrophages. In vitro TNF-α and IFN-γ induction in Peyer's patch (PP) cells was higher when incubated with H-Lp than with live L. plantarum BF-LP284 (L-Lp). Suppression of syngeneic Meth-A tumors in a murine model by oral administration of H-Lp was also greater than that of L-Lp and of controls. H-Lp stimulated IFN-γ production in spleen cells, which displayed inhibited tumor growth in Winn assays when treated with H-Lp. Moreover, H-Lp increased the ratio of CD3(+ )cells among peripheral blood mononuclear cells in Meth-A tumor-bearing mice, suggesting an H-Lp-mediated anti-tumor mechanism whereby immune cells that are activated by H-Lp in PP and acquire anti-tumor activity in the spleen migrate to tumor sites through lymphocyte homing to inhibit tumor growth.

  5. Treatment with 5-Aza-2'-Deoxycytidine Induces Expression of NY-ESO-1 and Facilitates Cytotoxic T Lymphocyte-Mediated Tumor Cell Killing.

    Directory of Open Access Journals (Sweden)

    Agnes S Klar

    Full Text Available NY-ESO-1 belongs to the cancer/testis antigen (CTA family and represents an attractive target for cancer immunotherapy. Its expression is induced in a variety of solid tumors via DNA demethylation of the promoter of CpG islands. However, NY-ESO-1 expression is usually very low or absent in some tumors such as breast cancer or multiple myeloma. Therefore, we established an optimized in vitro treatment protocol for up-regulation of NY-ESO-1 expression by tumor cells using the hypomethylating agent 5-aza-2'-deoxycytidine (DAC.We demonstrated de novo induction of NY-ESO-1 in MCF7 breast cancer cells and significantly increased expression in U266 multiple myeloma cells. This effect was time- and dose-dependent with the highest expression of NY-ESO-1 mRNA achieved by the incubation of 10 μM DAC for 72 hours. NY-ESO-1 activation was also confirmed at the protein level as shown by Western blot, flow cytometry, and immunofluorescence staining. The detection and quantification of single NY-ESO-1 peptides presented at the tumor cell surface in the context of HLA-A*0201 molecules revealed an increase of 100% and 50% for MCF7 and U266 cells, respectively. Moreover, the enhanced expression of NY-ESO-1 derived peptides at the cell surface was accompanied by an increased specific lysis of MCF7 and U266 cells by HLA-A*0201/NY-ESO-1(157-165 peptide specific chimeric antigen receptor (CAR CD8+ T cells. In addition, the killing activity of CAR T cells correlated with the secretion of higher IFN-gamma levels.These results indicate that NY-ESO-1 directed immunotherapy with specific CAR T cells might benefit from concomitant DAC treatment.

  6. Human NK cells differ more in their KIR2DL1-dependent thresholds for HLA-Cw6-mediated inhibition than in their maximal killing capacity.

    Directory of Open Access Journals (Sweden)

    Catarina R Almeida

    Full Text Available In this study we have addressed the question of how activation and inhibition of human NK cells is regulated by the expression level of MHC class I protein on target cells. Using target cell transfectants sorted to stably express different levels of the MHC class I protein HLA-Cw6, we show that induction of degranulation and that of IFN-γ secretion are not correlated. In contrast, the inhibition of these two processes by MHC class-I occurs at the same level of class I MHC protein. Primary human NK cell clones were found to differ in the amount of target MHC class I protein required for their inhibition, rather than in their maximum killing capacity. Importantly, we show that KIR2DL1 expression determines the thresholds (in terms of MHC I protein levels required for NK cell inhibition, while the expression of other receptors such as LIR1 is less important. Furthermore, using mathematical models to explore the dynamics of target cell killing, we found that the observed delay in target cell killing is exhibited by a model in which NK cells require some activation or priming, such that each cell can lyse a target cell only after being activated by a first encounter with the same or a different target cell, but not by models which lack this feature.

  7. Targeted alpha therapy in vivo: direct evidence for single cancer cell kill using {sup 149}Tb-rituximab

    Energy Technology Data Exchange (ETDEWEB)

    Beyer, G.J.; Soloviev, D.; Buchegger, F. [Division of Nuclear Medicine, University Hospital of Geneva, 24 Rue Micheli du Crest, 1211, Geneva 14 (Switzerland); Miederer, M. [Department of Molecular Pharmacology and Chemistry, Memorial Sloan-Kettering Cancer Center, New York (United States); Vranjes-Duric, S. [Laboratory of Radioisotopes, Vinca Institute of Nuclear Sciences, Belgrade (Czechoslovakia); Comor, J.J. [Laboratory of Physics, Vinca Institute of Nuclear Sciences, Belgrade (Czechoslovakia); Kuenzi, G.; Hartley, O. [Department of Medical Biochemistry, University Medical Center, Geneva (Switzerland); Senekowitsch-Schmidtke, R. [Clinic of Nuclear Medicine, Technical University of Munich, Munich (Germany)

    2004-04-01

    This study demonstrates high-efficiency sterilisation of single cancer cells in a SCID mouse model of leukaemia using rituximab, a monoclonal antibody that targets CD20, labelled with terbium-149, an alpha-emitting radionuclide. Radio-immunotherapy with 5.5 MBq labelled antibody conjugate (1.11 GBq/mg) 2 days after an intravenous graft of 5.10{sup 6} Daudi cells resulted in tumour-free survival for >120 days in 89% of treated animals. In contrast, all control mice (no treatment or treated with 5 or 300 {mu}g unlabelled rituximab) developed lymphoma disease. At the end of the study period, 28.4%{+-}4% of the long-lived daughter activity remained in the body, of which 91.1% was located in bone tissue and 6.3% in the liver. A relatively high daughter radioactivity concentration was found in the spleen (12%{+-}2%/g), suggesting that the killed cancer cells are mainly eliminated through the spleen. This promising preliminary in vivo study suggests that targeted alpha therapy with {sup 149}Tb is worthy of consideration as a new-generation radio-immunotherapeutic approach. (orig.)

  8. Stressing the ubiquitin-proteasome system without 20S proteolytic inhibition selectively kills cervical cancer cells.

    Directory of Open Access Journals (Sweden)

    Ravi K Anchoori

    Full Text Available Cervical cancer cells exhibit an increased requirement for ubiquitin-dependent protein degradation associated with an elevated metabolic turnover rate, and for specific signaling pathways, notably HPV E6-targeted degradation of p53 and PDZ proteins. Natural compounds with antioxidant properties including flavonoids and triterpenoids hold promise as anticancer agents by interfering with ubiquitin-dependent protein degradation. An increasing body of evidence indicates that their α-β unsaturated carbonyl system is the molecular determinant for inhibition of ubiquitin-mediated protein degradation up-stream of the catalytic sites of the 20S proteasome. Herein we report the identification and characterization of a new class of chalcone-based, potent and cell permeable chemical inhibitors of ubiquitin-dependent protein degradation, and a lead compound RAMB1. RAMB1 inhibits ubiquitin-dependent protein degradation without compromising the catalytic activities of the 20S proteasome, a mechanism distinct from that of Bortezomib. Treatment of cervical cancer cells with RAMB1 triggers unfolded protein responses, including aggresome formation and Hsp90 stabilization, and increases p53 steady state levels. RAMB1 treatment results in activation of lysosomal-dependent degradation pathways as a mechanism to compensate for increasing levels of poly-ubiquitin enriched toxic aggregates. Importantly, RAMB1 synergistically triggers cell death of cervical cancer cells when combined with the lysosome inhibitor Chloroquine.

  9. Targeting the Human Complement Membrane Attack Complex to Selectively Kill Prostate Cancer Cells

    Science.gov (United States)

    2013-10-01

    tract through the urethra. It must also shield the sperm from immune destruction within the vaginal tract while not eliminating cells within the...first of which plays a role in enhancing B cell immunity. iC3b’s other receptor binding partners, CR3 and CR4, have roles in clearance of pathogens by... Nature 444: 213–216. 25. Wiesmann, C., K. J. Katschke, J. Yin, K. Y. Helmy, M. Steffek, W. J. Fairbrother, S. A. McCallum, L. Embuscado, L. DeForge, P

  10. Rapid and efficient cancer cell killing mediated by high-affinity death receptor homotrimerizing TRAIL variants.

    Science.gov (United States)

    Reis, C R; van der Sloot, A M; Natoni, A; Szegezdi, E; Setroikromo, R; Meijer, M; Sjollema, K; Stricher, F; Cool, R H; Samali, A; Serrano, L; Quax, W J

    2010-10-21

    The tumour necrosis factor family member TNF-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in a variety of cancer cells through the activation of death receptors 4 (DR4) and 5 (DR5) and is considered a promising anticancer therapeutic agent. As apoptosis seems to occur primarily via only one of the two death receptors in many cancer cells, the introduction of DR selectivity is thought to create more potent TRAIL agonists with superior therapeutic properties. By use of a computer-aided structure-based design followed by rational combination of mutations, we obtained variants that signal exclusively via DR4. Besides an enhanced selectivity, these TRAIL-DR4 agonists show superior affinity to DR4, and a high apoptosis-inducing activity against several TRAIL-sensitive and -resistant cancer cell lines in vitro. Intriguingly, combined treatment of the DR4-selective variant and a DR5-selective TRAIL variant in cancer cell lines signalling by both death receptors leads to a significant increase in activity when compared with wild-type rhTRAIL or each single rhTRAIL variant. Our results suggest that TRAIL induced apoptosis via high-affinity and rapid-selective homotrimerization of each DR represent an important step towards an efficient cancer treatment.

  11. Glycan elongation beyond the mucin associated Tn antigen protects tumor cells from immune-mediated killing

    DEFF Research Database (Denmark)

    Mathiesen, Caroline Benedicte Kjærulff; Lavrsen, Kirstine; Wandall, Hans H.;

    2013-01-01

    Membrane bound mucins are up-regulated and aberrantly glycosylated during malignant transformation in many cancer cells. This results in a negatively charged glycoprotein coat which may protect cancer cells from immune surveillance. However, only limited data have so far demonstrated the critical...... steps in glycan elongation that make aberrantly glycosylated mucins affect the interaction between cancer cells and cytotoxic effector cells of the immune system. Tn (GalNAc-Ser/Thr), STn (NeuAcα2-6GalNAc-Ser/Thr), T (Galβ1–3GalNAc-Ser/Thr), and ST (NeuAcα2-6Galβ1–3GalNAc-Ser/Thr) antigens...... only the shortest possible mucin-like glycans (Tn and STn). Glyco-engineering was performed by zinc finger nuclease (ZFN) knockout (KO) of the Core 1 enzyme chaperone COSMC, thereby preventing glycan elongation beyond the initial GalNAc residue in O-linked glycans. We find that COSMC KO in the breast...

  12. Photodynamic killing of cancer cells by a Platinum(II) complex with cyclometallating ligand

    Science.gov (United States)

    Doherty, Rachel E.; Sazanovich, Igor V.; McKenzie, Luke K.; Stasheuski, Alexander S.; Coyle, Rachel; Baggaley, Elizabeth; Bottomley, Sarah; Weinstein, Julia A.; Bryant, Helen E.

    2016-03-01

    Photodynamic therapy that uses photosensitizers which only become toxic upon light-irradiation provides a strong alternative to conventional cancer treatment due to its ability to selectively target tumour material without affecting healthy tissue. Transition metal complexes are highly promising PDT agents due to intense visible light absorption, yet the majority are toxic even without light. This study introduces a small, photostable, charge-neutral platinum-based compound, Pt(II) 2,6-dipyrido-4-methyl-benzenechloride, complex 1, as a photosensitizer, which works under visible light. Activation of the new photosensitizer at low concentrations (0.1-1 μM) by comparatively low dose of 405 nm light (3.6 J cm-2) causes significant cell death of cervical, colorectal and bladder cancer cell lines, and, importantly, a cisplatin resistant cell line EJ-R. The photo-index of the complex is 8. We demonstrate that complex 1 induces irreversible DNA single strand breaks following irradiation, and that oxygen is essential for the photoinduced action. Neither light, nor compound alone led to cell death. The key advantages of the new drug include a remarkably fast accumulation time (diffusion-controlled, minutes), and photostability. This study demonstrates a highly promising new agent for photodynamic therapy, and attracts attention to photostable metal complexes as viable alternatives to conventional chemotherapeutics, such as cisplatin.

  13. Photodynamic killing of cancer cells by a Platinum(II) complex with cyclometallating ligand.

    Science.gov (United States)

    Doherty, Rachel E; Sazanovich, Igor V; McKenzie, Luke K; Stasheuski, Alexander S; Coyle, Rachel; Baggaley, Elizabeth; Bottomley, Sarah; Weinstein, Julia A; Bryant, Helen E

    2016-03-04

    Photodynamic therapy that uses photosensitizers which only become toxic upon light-irradiation provides a strong alternative to conventional cancer treatment due to its ability to selectively target tumour material without affecting healthy tissue. Transition metal complexes are highly promising PDT agents due to intense visible light absorption, yet the majority are toxic even without light. This study introduces a small, photostable, charge-neutral platinum-based compound, Pt(II) 2,6-dipyrido-4-methyl-benzenechloride, complex 1, as a photosensitizer, which works under visible light. Activation of the new photosensitizer at low concentrations (0.1-1 μM) by comparatively low dose of 405 nm light (3.6 J cm(-2)) causes significant cell death of cervical, colorectal and bladder cancer cell lines, and, importantly, a cisplatin resistant cell line EJ-R. The photo-index of the complex is 8. We demonstrate that complex 1 induces irreversible DNA single strand breaks following irradiation, and that oxygen is essential for the photoinduced action. Neither light, nor compound alone led to cell death. The key advantages of the new drug include a remarkably fast accumulation time (diffusion-controlled, minutes), and photostability. This study demonstrates a highly promising new agent for photodynamic therapy, and attracts attention to photostable metal complexes as viable alternatives to conventional chemotherapeutics, such as cisplatin.

  14. Spontaneous human squamous cell carcinomas are killed by a human cytotoxic T lymphocyte clone recognizing a wild-type p53-derived peptide

    DEFF Research Database (Denmark)

    Röpke, M; Hald, J; Guldberg, Per

    1996-01-01

    A cytotoxic T lymphocyte (CTL) clone generated in vitro from the peripheral blood of a healthy HLA-A2-positive individual against a synthetic p53 protein-derived wild-type peptide (L9V) was shown to kill squamous carcinoma cell lines derived from two head and neck carcinomas, which expressed mutant...

  15. Specific killing of P53 mutated tumor cell lines by a cross-reactive human HLA-A2-restricted P53-specific CTL line

    DEFF Research Database (Denmark)

    Würtzen, P A; Pedersen, L O; Poulsen, H S;

    2001-01-01

    53(65-73) and p53(187-197) peptides was obtained in the T-cell lines. Interestingly, cold target inhibition experiments demonstrated that the simultaneous recognition of the 2 peptides was the result of cross-reactivity, which was confirmed by killing experiments at the clonal CTL level. Furthermore...

  16. A designed equine herpes thymidine kinase (EHV4 TK) variant improves ganciclovir-induced cell-killing.

    Science.gov (United States)

    McSorley, Theresa; Ort, Stephan; Monnerjahn, Christian; Konrad, Manfred

    2014-02-01

    The limitations of the ganciclovir (GCV)/herpes simplex virus thymidine kinase (HSV1 TK: EC 2.7.1.21) system as a suicide gene therapy approach have been extensively studied over the years. In our study, we focused on improving the cytotoxic profile of the GCV/equine herpes virus-4 thymidine kinase (EHV4 TK: EC 2.7.1.21) system. Our approach involved the structure-guided mutagenesis of EHV4 TK in order to switch its ability to preferentially phosphorylate the natural substrate deoxythymidine (dT) to that of GCV. We performed steady-state kinetic analysis, genetic complementation in a thymidine kinase-deficient Escherichia coli strain, isothermal titration calorimetry, and analysis of GCV-induced cell killing through generation of HEK 293 stable cell-lines expressing EHV4 TK mutants and wild-type EHV4 TK. We found that the EHV4 TK S144H-GFP mutant preferentially phosphorylates GCV and confers increased GCV-induced cytotoxicity compared to wild-type EHV4 TK.

  17. Identification of protective pneumococcal T(H17 antigens from the soluble fraction of a killed whole cell vaccine.

    Directory of Open Access Journals (Sweden)

    Kristin L Moffitt

    Full Text Available Mucosal or parenteral immunization with a killed unencapsulated pneumococcal whole cell antigen (WCA with an adjuvant protects mice from colonization by a T(H17 CD4+ cell-mediated mechanism. Using preparative SDS gels, we separated the soluble proteins that compose the WCA in order to identify fractions that were immunogenic and protective. We screened these fractions for their ability to stimulate IL-17A secretion from splenocytes obtained from mice immunized with WCA and adjuvant. We identified 12 proteins within the stimulatory fractions by mass spectrometry; these proteins were then cloned, recombinantly expressed and purified using an Escherichia coli expression system. The ability of these proteins to induce IL-17A secretion was then evaluated by stimulation of mouse splenocytes. Of the four most stimulatory proteins, three were protective in a mouse pneumococcal serotype 6B colonization model. This work thus describes a method for identifying immunogenic proteins from the soluble fraction of pneumococcus and shows that several of the proteins identified protect mice from colonization when used as mucosal vaccines. We propose that, by providing protection against pneumococcal colonization, one or more of these proteins may serve as components of a multivalent pneumococcal vaccine.

  18. Enhanced killing activity of regulatory T cells ameliorates inflammation and autoimmunity.

    Science.gov (United States)

    Askenasy, Nadir

    2013-08-01

    Regulatory T cells (Treg) are pivotal suppressor elements in immune homeostasis with potential therapeutic applications in inflammatory and autoimmune disorders. Using Treg as vehicles for targeted immunomodulation, a short-lived Fas-ligand (FasL) chimeric protein (killer Treg) was found efficient in preventing the progression of autoimmune insulitis in NOD mice, and amelioration of chronic colitis and graft versus host disease. The main mechanisms of disease suppression by killer Treg are: a) in the acute phase induction of apoptosis in effector cells at the site of inflammation decreases the pathogenic burden, and b) persistent increase in FoxP3⁺ Treg with variable CD25 co-expression induced by FasL sustains disease suppression over extended periods of time. Reduced sensitivity of Treg to receptor-mediated apoptosis under inflammatory conditions makes them optimal vehicles for targeted immunotherapy using apoptotic agents.

  19. Talking about killing: Cell Phones, Collective Action, and Insurgent Violence in Iraq

    Science.gov (United States)

    2011-05-31

    technology, insurgency, collective action Jacob Shapiro, Nils Weidmann Harvard University Office of Sponsored Research 1350 Massachusetts Ave. Holyoke 727...at the district level changes as a result of the introduction of new cell phone towers. Second, using a novel identification strategy, we examine how... identification strategy, we examine how insurgent operation in the tower’s vicinity is affected by the introduction of coverage. Taken together, our

  20. Effect of telithromycin (HMR 3647) on polymorphonuclear neutrophil killing of Staphylococcus aureus in comparison with roxithromycin.

    Science.gov (United States)

    Vazifeh, D; Abdelghaffar, H; Labro, M T

    2002-05-01

    HMR 3647 (telithromycin), a new ketolide, is active on intracellular pathogens. It was previously demonstrated that it inhibits superoxide anion production in a time- and concentration-dependent manner, at concentrations which inhibit 50% of the control response of about 55 microg/ml (5 min) to 30 microg/ml (30 min); these values are similar to those obtained with roxithromycin, a classical erythromycin A derivative. Here we investigated whether these drugs modified the bactericidal activity of human polymorphonuclear neutrophils (PMN) on four strains of Staphylococcus aureus with different profiles of susceptibility to macrolides and ketolides. We found that the main factor involved in killing was the antibacterial potency of the drugs, although combinations of antibiotics with PMN were slightly more active than each component used alone against two of the four strains. In addition, high concentrations of the drugs, which impaired the PMN oxidative burst, did not impair PMN bactericidal activity. Likewise, some cytokines which enhance PMN oxidative metabolism did not modify PMN bactericidal activity in the presence or absence of macrolides or ketolides. These data suggest that oxygen-independent mechanisms contribute to the bactericidal activity of PMN on these strains of S. aureus. Both live and/or heat-killed bacteria impaired the uptake of telithromycin and roxithromycin (but not that of levofloxacin, a quinolone) in a concentration-dependent manner, owing to a modulation of PMN transductional systems involved in the activation of the macrolide carrier.

  1. Anti-breast cancer effects of live, heat-killed and cytoplasmic fractions of Enterococcus faecalis and Staphylococcus hominis isolated from human breast milk.

    Science.gov (United States)

    Hassan, Zubaida; Mustafa, Shuhaimi; Rahim, Raha Abdul; Isa, Nurulfiza Mat

    2016-03-01

    Development of tumour that is resistant to chemotherapeutics and synthetic drugs, coupled with their life-threatening side effects and the adverse effects of surgery and hormone therapies, led to increased research on probiotics' anticancer potentials. The current study investigated the potential of live, heat-killed cells (HKC) and the cytoplasmic fractions (CF) of Enterococcus faecalis and Staphylococcus hominis as anti-breast cancer agents. MCF-7 cell line was treated with 25, 50, 100 and 200 μg/mL each of live, HKC and CF of the bacteria; and cytotoxicity was evaluated for 24, 48 and 72 h using MTT assay. The morphological features of the treated cells were examined by fluorescence microscopy. The stage of cell cycle arrest and apoptosis were quantified by flow cytometry. The bacterial effect on non-malignant breast epithelial cell line, MCF-10A, was assessed using MTT assay for 24, 48 and 72 h. All the three forms of the bacteria caused a significant decrease in MCF-7 (up to 33.29%) cell proliferation in concentration- and time-dependent manner. Morphological features of apoptosis like cell death, cell shrinkage and membrane blebbing were observed. Flow cytometry analyses suggested that about 34.60% of treated MCF-7 was undergoing apoptosis. A strong anti-proliferative activity was efficiently induced through sub-G1 accumulation (up to 83.17%) in treated MCF-7 and decreased number in the G0/G1 phase (74.39%). MCF-10A cells treated with both bacteria showed no significant difference with the untreated (>90% viability). These bacteria can be used as good alternative nutraceutical with promising therapeutic indexes for breast cancer because of their non-cytotoxic effects to normal cells.

  2. Chaperone-Targeting Cytotoxin and Endoplasmic Reticulum Stress-Inducing Drug Synergize to Kill Cancer Cells

    Directory of Open Access Journals (Sweden)

    Joseph M. Backer

    2009-11-01

    Full Text Available Diverse physiological and therapeutic insults that increase the amount of unfolded or misfolded proteins in the endoplasmic reticulum (ER induce the unfolded protein response, an evolutionarily conserved protective mechanism that manages ER stress. Glucose-regulated protein 78/immunoglobulin heavy-chain binding protein (GRP78/BiP is an ER-resident protein that plays a central role in the ER stress response and is the only known substrate of the proteolytic A subunit (SubA of a novel bacterial AB5 toxin. Here, we report that an engineered fusion protein, epidermal growth factor (EGF-SubA, combining EGF and SubA, is highly toxic to growing and confluent epidermal growth factor receptor-expressing cancer cells, and its cytotoxicity is mediated by a remarkably rapid cleavage of GRP78/BiP. Systemic delivery of EGF-SubA results in a significant inhibition of human breast and prostate tumor xenografts in mouse models. Furthermore, EGF-SubA dramatically increases the sensitivity of cancer cells to the ER stress-inducing drug thapsigargin, and vice versa, demonstrating the first example of mechanism-based synergism in the action of a cytotoxin and an ER-targeting drug.

  3. Cisplatin in 5% Ethanol Eradicates Cisplatin-Resistant Lung Tumor by Killing Lung Cancer Side Population (SP Cells and Non-SP Cells

    Directory of Open Access Journals (Sweden)

    Qi eNiu

    2013-08-01

    Full Text Available Cancer side population (SP cells with cancer stem cell-like properties are thought to be responsible for lung cancer chemotherapy resistance and currently no drug can efficiently target them. Breast cancer resistance protein (BRCP/ABCG2 is a major drug transporter in protecting lung cancer SP cells from cytotoxic agents. We showed that a low concentration of ethanol, which inhibits many membrane proteins, inhibits ABCG2 in lung cancer SP cells. Furthermore, cytotoxic cisplatin (DDP in 5% (vol/vol ethanol kills SP plus non-SP cancer cells better than either treatment alone in eradicating chemoresistant lung tumors. We found that 5% ethanol did not reduce ABCG2 protein levels, but significantly reduced ABCG2 protein function by a Hoechst 33342 extrusion assay, an ATPase activity assay, and transmission electron microscopy. Further, DDP in 5% ethanol (5% ethanol-DDP induced apoptosis of the SP plus non-SP cancer cells both in vitro and in vivo. In DDP-resistant A549/DDP lung tumor-bearing Balb/C nude mice, intratumoral injection of 5% ethanol-DDP regressed tumors and significantly improved survivals compared with 5% ethanol, DDP alone, or control. Intratumoral injection of 5% ethanol-DDP helped eradicate tumors in 30% (3/10 of the mice after 4 weeks treatment. By killing SP and non-SP cancer cells, 5% ethanol-DDP could eradicate DDP-resistant lung tumor and extend survival, providing a novel way to improve chemoresistant lung cancer survival for clinic.

  4. Synergy of caspofungin with human polymorphonuclear granulocytes for killing Candida albicans

    OpenAIRE

    Tullio, Vivian; Mandras, Narcisa; Scalas, Daniela; Allizond, Valeria; Banche, Giuliana; Roana, Janira; Greco, Deborah; Castagno, Franco; Cuffini, Anna Maria; Carlone, Nicola A.

    2010-01-01

    The influence of caspofungin on polymorphonuclear leukocyte (PMN) phagocytosis and intracellular killing of Candida albicans was investigated. Caspofungin, at all of the concentrations tested (2, 3.2, and 8 μg/ml), significantly increased intracellular killing by PMNs through its direct action on both yeast cells and PMNs, indicating the potential ability of caspofungin to synergize with phagocytes for candidal killing. Caspofungin may therefore constitute an effective therapeutic option for ...

  5. Bacteriophage-derived CHAP domain protein, P128, kills Staphylococcus cells by cleaving interpeptide cross-bridge of peptidoglycan.

    Science.gov (United States)

    Sundarrajan, Sudarson; Raghupatil, Junjappa; Vipra, Aradhana; Narasimhaswamy, Nagalakshmi; Saravanan, Sanjeev; Appaiah, Chemira; Poonacha, Nethravathi; Desai, Srividya; Nair, Sandhya; Bhatt, Rajagopala Narayana; Roy, Panchali; Chikkamadaiah, Ravisha; Durgaiah, Murali; Sriram, Bharathi; Padmanabhan, Sriram; Sharma, Umender

    2014-10-01

    P128 is an anti-staphylococcal protein consisting of the Staphylococcus aureus phage-K-derived tail-associated muralytic enzyme (TAME) catalytic domain (Lys16) fused with the cell-wall-binding SH3b domain of lysostaphin. In order to understand the mechanism of action and emergence of resistance to P128, we isolated mutants of Staphylococcus spp., including meticillin-resistant Staphylococcus aureus (MRSA), resistant to P128. In addition to P128, the mutants also showed resistance to Lys16, the catalytic domain of P128. The mutants showed loss of fitness as shown by reduced rate of growth in vitro. One of the mutants tested was found to show reduced virulence in animal models of S. aureus septicaemia suggesting loss of fitness in vivo as well. Analysis of the antibiotic sensitivity pattern showed that the mutants derived from MRSA strains had become sensitive to meticillin and other β-lactams. Interestingly, the mutant cells were resistant to the lytic action of phage K, although the phage was able to adsorb to these cells. Sequencing of the femA gene of three P128-resistant mutants showed either a truncation or deletion in femA, suggesting that improper cross-bridge formation in S. aureus could be causing resistance to P128. Using glutathione S-transferase (GST) fusion peptides as substrates it was found that both P128 and Lys16 were capable of cleaving a pentaglycine sequence, suggesting that P128 might be killing S. aureus by cleaving the pentaglycine cross-bridge of peptidoglycan. Moreover, peptides corresponding to the reported cross-bridge of Staphylococcus haemolyticus (GGSGG, AGSGG), which were not cleaved by lysostaphin, were cleaved efficiently by P128. This was also reflected in high sensitivity of S. haemolyticus to P128. This showed that in spite of sharing a common mechanism of action with lysostaphin, P128 has unique properties, which allow it to act on certain lysostaphin-resistant Staphylococcus strains.

  6. A multifunctional probe with aggregation-induced emission characteristics for selective fluorescence imaging and photodynamic killing of bacteria over mammalian cells.

    Science.gov (United States)

    Gao, Meng; Hu, Qinglian; Feng, Guangxue; Tomczak, Nikodem; Liu, Rongrong; Xing, Bengang; Tang, Ben Zhong; Liu, Bin

    2015-04-02

    A multifunctional probe aggregation-induced emission-Zinc(II)-dipicolylamine (AIE-ZnDPA) is developed for selective targeting, fluorescence imaging, and photodynamic killing of both Gram-positive and Gram-negative bacteria over mammalian cells. The probe has significant advantages in simple probe design, enhanced fluorescence upon bacteria binding, excellent photostability, and broad-spectrum antibacterial activity with almost no harm to mammalian cells.

  7. "Kill" the messenger: Targeting of cell-derived microparticles in lupus nephritis.

    Science.gov (United States)

    Nielsen, Christoffer T; Rasmussen, Niclas S; Heegaard, Niels H H; Jacobsen, Søren

    2016-07-01

    Immune complex (IC) deposition in the glomerular basement membrane (GBM) is a key early pathogenic event in lupus nephritis (LN). The clarification of the mechanisms behind IC deposition will enable targeted therapy in the future. Circulating cell-derived microparticles (MPs) have been proposed as major sources of extracellular autoantigens and ICs and triggers of autoimmunity in LN. The overabundance of galectin-3-binding protein (G3BP) along with immunoglobulins and a few other proteins specifically distinguish circulating MPs in patients with systemic lupus erythematosus (SLE), and this is most pronounced in patients with active LN. G3BP co-localizes with deposited ICs in renal biopsies from LN patients supporting a significant presence of MPs in the IC deposits. G3BP binds strongly to glomerular basement membrane proteins and integrins. Accordingly, MP surface proteins, especially G3BP, may be essential for the deposition of ICs in kidneys and thus for the ensuing formation of MP-derived electron dense structures in the GBM, and immune activation in LN. This review focuses on the notion of targeting surface molecules on MPs as an entirely novel treatment strategy in LN. By targeting MPs, a double hit may be achieved by attenuating both the autoantigenic fueling of immune complexes and the triggering of the adaptive immune system. Thereby, early pathogenic events may be blocked in contrast to current treatment strategies that primarily target and modulate later events in the cellular and humoral immune response.

  8. Drug embedded PVP coated magnetic nanoparticles for targeted killing of breast cancer cells.

    Science.gov (United States)

    Rose, P Arsula; Praseetha, P K; Bhagat, Madhulika; Alexander, Princy; Abdeen, Sunitha; Chavali, Murthy

    2013-10-01

    Magnetic drug targeting is a drug delivery system that can be used in loco-regional cancer treatment. Coated magnetic particles, called carriers, are very useful for delivering chemotherapeutic drugs. Magnetic carriers were synthesized by co-precipitation of iron oxide followed by coating with polyvinyl pyrrolidone (PVP). Characterization was performed using X-ray diffraction, TEM, TGA, FTIR and UV-Vis Spectroscopy. Magnetite (Fe3O4) remained as the core of the carrier. The amount of PVP bound to the iron oxide nanoparticles was estimated by thermogravimetric analysis (TGA) and the attachment of PVP to the iron oxide nanoparticles confirmed by FTIR analysis. The loading efficiency of Epirubicin hydrochloride onto the PVP coated and uncoated iron oxide nanoparticles was measured at intervals such as 1 hr and 24 hrs by UV-Vis Spectroscopy. The binding of Epirubicin hydrochloride to the PVP coated and uncoated iron oxide nanoparticles were confirmed by FTIR analysis. The present findings showed that Epirubicin hydrochloride loaded PVP coated iron oxide nanoparticles are promising for magnetically targeted drug delivery. The drug displayed increased cell cytotoxicity at lower concentrations when conjugated with the nanoparticles than being administered conventionally as individual drugs.

  9. Vancomycin-modified mesoporous silica nanoparticles for selective recognition and killing of pathogenic gram-positive bacteria over macrophage-like cells.

    Science.gov (United States)

    Qi, Guobin; Li, Lili; Yu, Faquan; Wang, Hao

    2013-11-13

    Rapid, reliable recognition and detection of bacteria from an authentic specimen have been gained increasing interests in the past decades. Various materials have been designed and prepared for implementation of bacterial recognition and treatment in the artificial systems. However, in the complicated physiological condition, the macrophages always compromise the outcomes of bacterial detection and/or treatment. In this work, we demonstrated the vancomycin-modified mesoporous silica nanoparticles (MSNs is a subset of Van) for efficiently targeting and killing gram-positive bacteria over macrophage-like cells. Owing to the specific hydrogen bonding interactions of vancomycin toward the terminal d-alanyl-d-alanine moieties of gram-positive bacteria, the MSNs is a subset of Van exhibited enhanced recognition for gram-positive bacteria due to the multivalent hydrogen binding effect. Furthermore, the fluorescent molecules (FITC) were covalently decorated inside of mesopores of MSNs for tracking and visualizing the MSNs is a subset of Van during the detection/treatment processes. Upon incubation of FITC decorated MSNs with bacteria (i.e., S. aureus and E. coli as gram-positive and gram-negative bacteria, respectively) or macrophage-like cells (Raw 264.7), the fluorescence signals in S. aureus were 2-4 times higher than that in E. coli and no detectable fluorescence signals were observed in Raw 264.7 cells under the same condition. Finally, the MSNs is a subset of Van showed unambiguous antibacterial efficacy without decrease in cell viability of macrophage-like cells. This new strategy opens a new door for specific detection and treatment of pathogenic bacteria with minimized side effects.

  10. Effect of nikkomycin Z and 50% human serum on the killing activity of high-concentration caspofungin against Candida species using time-kill methodology.

    Science.gov (United States)

    Szilágyi, Judit; Földi, Richárd; Bayegan, Sedigh; Kardos, Gábor; Majoros, László

    2012-02-01

    Caspofungin and nikkomycin Z (NIK) efficacy alone and in combination were tested against seven Candida species showing or not showing paradoxical growth (PG) against caspofungin in time-kill test in RPMI-1640. Selected isolates against caspofungin and NIK were also tested in 50% serum. PG was always eliminated by NIK as well as by serum. In the serum, 1 and 16 μg/ml caspofungin yielded 0.14-4.0 and 0.34-4.0 log CFU decreases from the starting inocula for C. albicans, C. glabrata, C. tropicalis, and C. dubliniensis, respectively. CFU decrease (0.10-2.08 log) at 16 μg/ml, but not at lower caspofungin concentration was noted against C. parapsilosis, C. orthopsilosis, and C. metapsilosis. One C. parapsilosis isolate was not inhibited even by 16 μg/ml caspofungin. Caspofungin against C. albicans, C. glabrata, C. tropicalis, and C. dubliniensis maintained its activity in serum at even 1 μg/ml concentration. PG seems to an in vitro phenomenon, without clinical relevance.

  11. Porphyromonas gingivalis Evasion of Autophagy and Intracellular Killing by Human Myeloid Dendritic Cells Involves DC-SIGN-TLR2 Crosstalk

    Science.gov (United States)

    El-Awady, Ahmed R.; Miles, Brodie; Scisci, Elizabeth; Kurago, Zoya B.; Palani, Chithra D.; Arce, Roger M.; Waller, Jennifer L.; Genco, Caroline A.; Slocum, Connie; Manning, Matthew; Schoenlein, Patricia V.; Cutler, Christopher W.

    2015-01-01

    Signaling via pattern recognition receptors (PRRs) expressed on professional antigen presenting cells, such as dendritic cells (DCs), is crucial to the fate of engulfed microbes. Among the many PRRs expressed by DCs are Toll-like receptors (TLRs) and C-type lectins such as DC-SIGN. DC-SIGN is targeted by several major human pathogens for immune-evasion, although its role in intracellular routing of pathogens to autophagosomes is poorly understood. Here we examined the role of DC-SIGN and TLRs in evasion of autophagy and survival of Porphyromonas gingivalis in human monocyte-derived DCs (MoDCs). We employed a panel of P. gingivalis isogenic fimbriae deficient strains with defined defects in Mfa-1 fimbriae, a DC-SIGN ligand, and FimA fimbriae, a TLR2 agonist. Our results show that DC-SIGN dependent uptake of Mfa1+P. gingivalis strains by MoDCs resulted in lower intracellular killing and higher intracellular content of P. gingivalis. Moreover, Mfa1+P. gingivalis was mostly contained within single membrane vesicles, where it survived intracellularly. Survival was decreased by activation of TLR2 and/or autophagy. Mfa1+P. gingivalis strain did not induce significant levels of Rab5, LC3-II, and LAMP1. In contrast, P. gingivalis uptake through a DC-SIGN independent manner was associated with early endosomal routing through Rab5, increased LC3-II and LAMP-1, as well as the formation of double membrane intracellular phagophores, a characteristic feature of autophagy. These results suggest that selective engagement of DC-SIGN by Mfa-1+P. gingivalis promotes evasion of antibacterial autophagy and lysosome fusion, resulting in intracellular persistence in myeloid DCs; however TLR2 activation can overcome autophagy evasion and pathogen persistence in DCs. PMID:25679217

  12. Porphyromonas gingivalis evasion of autophagy and intracellular killing by human myeloid dendritic cells involves DC-SIGN-TLR2 crosstalk.

    Directory of Open Access Journals (Sweden)

    Ahmed R El-Awady

    2015-02-01

    Full Text Available Signaling via pattern recognition receptors (PRRs expressed on professional antigen presenting cells, such as dendritic cells (DCs, is crucial to the fate of engulfed microbes. Among the many PRRs expressed by DCs are Toll-like receptors (TLRs and C-type lectins such as DC-SIGN. DC-SIGN is targeted by several major human pathogens for immune-evasion, although its role in intracellular routing of pathogens to autophagosomes is poorly understood. Here we examined the role of DC-SIGN and TLRs in evasion of autophagy and survival of Porphyromonas gingivalis in human monocyte-derived DCs (MoDCs. We employed a panel of P. gingivalis isogenic fimbriae deficient strains with defined defects in Mfa-1 fimbriae, a DC-SIGN ligand, and FimA fimbriae, a TLR2 agonist. Our results show that DC-SIGN dependent uptake of Mfa1+P. gingivalis strains by MoDCs resulted in lower intracellular killing and higher intracellular content of P. gingivalis. Moreover, Mfa1+P. gingivalis was mostly contained within single membrane vesicles, where it survived intracellularly. Survival was decreased by activation of TLR2 and/or autophagy. Mfa1+P. gingivalis strain did not induce significant levels of Rab5, LC3-II, and LAMP1. In contrast, P. gingivalis uptake through a DC-SIGN independent manner was associated with early endosomal routing through Rab5, increased LC3-II and LAMP-1, as well as the formation of double membrane intracellular phagophores, a characteristic feature of autophagy. These results suggest that selective engagement of DC-SIGN by Mfa-1+P. gingivalis promotes evasion of antibacterial autophagy and lysosome fusion, resulting in intracellular persistence in myeloid DCs; however TLR2 activation can overcome autophagy evasion and pathogen persistence in DCs.

  13. Low pH immobilizes and kills human leukocytes and prevents transmission of cell-associated HIV in a mouse model

    Directory of Open Access Journals (Sweden)

    Markham Richard B

    2005-09-01

    results suggest that physiologic or microbicide-induced acid immobilization and killing of infected white blood cells may be effective in preventing sexual transmission of cell-associated HIV.

  14. Keyhole limpet hemocyanin augmented the killing activity, cytokine production and proliferation of NK cells, and inhibited the proliferation of Meth A sarcoma cells in vitro

    Directory of Open Access Journals (Sweden)

    Md. Moklesur Rahman Sarker

    2014-01-01

    Full Text Available Objective: Keyhole limpet hemocyanin (KLH is a popular tumor vaccine carrier protein and an immunostimulant. The present study aimed to investigate the immunoregulatory activity of KLH on cytotoxicity, cytokines production, and proliferation of natural killer (NK cells. Moreover, antiproliferative activity of KLH on Meth A sarcoma cells was studied. Materials and Methods: Cytotoxicity was determined with killing ability of NK cells against yeast artificial chromosome (YAC-1 cells. Interferon-gamma (IFN-γ and tumor necrosis factor-alpha (TNF-α productions by NK cells were measured by enzyme-linked immunosorbent assay (ELISA. Proliferations of NK and Meth A cells were determined by [ 3 H]thymidine incorporated proliferation and 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT methods, respectively. Results: KLH at 6.25, 12.5, and 25 μg/well augmented cytotoxicity of NK cells against YAC-1 cells by 2.5, three, and five-times, respectively. KLH at 25 μg/well enhanced IFN-γ and TNF-α productions by 17- and 23-folds, respectively. The proliferation of NK cells was three times stimulated by KLH. The proliferation of Meth A cells was markedly inhibited by all the doses; the highest (4-folds higher inhibition was observed at a dose of KLH (25 μg/well. Conclusion: The study demonstrated the anticancer activity of KLH acting through the induction of NK cells and inhibition of cancer cells. KLH, therefore, may be a good candidate for an anticancer agent alone or in combination with other chemotherapeutic agents.

  15. Heat Killed Lactobacillus reuteri GMNL-263 Reduces Fibrosis Effects on the Liver and Heart in High Fat Diet-Hamsters via TGF-β Suppression

    Directory of Open Access Journals (Sweden)

    Wei-Jen Ting

    2015-10-01

    Full Text Available Obesity is one of the major risk factors for nonalcoholic fatty liver disease (NAFLD, and NAFLD is highly associated with an increased risk of cardiovascular disease (CVD. Scholars have suggested that certain probiotics may significantly impact cardiovascular health, particularly certain Lactobacillus species, such as Lactobacillus reuteri GMNL-263 (Lr263 probiotics, which have been shown to reduce obesity and arteriosclerosis in vivo. In the present study, we examined the potential of heat-killed bacteria to attenuate high fat diet (HFD-induced hepatic and cardiac damages and the possible underlying mechanism of the positive effects of heat-killed Lr263 oral supplements. Heat-killed Lr263 treatments (625 and 3125 mg/kg-hamster/day were provided as a daily supplement by oral gavage to HFD-fed hamsters for eight weeks. The results show that heat-killed Lr263 treatments reduce fatty liver syndrome. Moreover, heat-killed Lactobacillus reuteri GMNL-263 supplementation in HFD hamsters also reduced fibrosis in the liver and heart by reducing transforming growth factor β (TGF-β expression levels. In conclusion, heat-killed Lr263 can reduce lipid metabolic stress in HFD hamsters and decrease the risk of fatty liver and cardiovascular disease.

  16. In vitro killing assays of antisera antibody sheep post-infected with Fasciola gigantica with the presence of macrophages cells against homologous and heterologous liver flukes

    Directory of Open Access Journals (Sweden)

    S.E Estuningsih

    1999-10-01

    Full Text Available The previous artificial infection known that the Indonesian Thin Tail (ITT sheep was resistance against the liver fluke of Fasciola gigantica, the resistances occurred in the early infection. In order to observe the immune resistance, some in vitro studies were undertaken in the laboratory, to assay the ability of the antisera antibody of ITT sheep post-infected with F. gigantica, with the presence of macrophages cells in killing the homologous and heterologous liver flukes. The viability of liver flukes were observed within 24-72 hours of incubation period by observing their motility (motile flukes were designated live and non-motile once were death. The results showed that after 72 hours incubation, the motilities of the Newly Excysted Juvenile (NEJ of F. gigantica incubated with the presence of post-infected sera and macrophages cells solution were significantly lower (P0.05. It seems that the occurrence of homologous antibody to the antigens is very important in the development of killing mechanism. The absence of homologous antibody did not reduce the number of flukes or the ability of macrophages cells in killing F. hepatica was not apparent.

  17. Immunity to Schistosoma mansoni in guinea-pigs vaccinated with radiation-attenuated cercariae. T-cell activation of macrophages for larval killing

    Energy Technology Data Exchange (ETDEWEB)

    Gordon, J.R.; McLaren, D.J.

    1988-02-01

    This study addresses macrophage activation in guinea-pigs vaccinated with radiation-attenuated cercariae of Schistosom mansoni. Peritoneal exudate macrophages elicited in vaccinated animals by mineral oil injection were activated to kill larval schistosomes in vitro. Killing efficiency is dependent upon the cell:target ratio employed and is enhanced by, but is not strictly dependent on, the presence of specific antibodies. Macrophages co-cultured with parasites release superoxide radicals and hydrogen peroxide, but the use of inhibitors has shown that neither of these reactive oxygen intermediates are the causal agents of cellular cytotoxicity in this system. Oil-elicited macrophages from naive guinea-pigs do not show comparable activation; they can, however, be activated in vitro by incubation with culture supernatant fluids from schistosome antigen-stimulated spleen, or lymph node cells harvested from vaccinated guinea-pigs. Naive macrophages activated in this way kill schistosomula in vitro and release the activation markers IL-l and superoxide anion. The macrophage-activating factor (MAF) present in spleen cell culture supernatant fluids has a MW of 35,000-55,000, but does not have the chemical characteristics of gamma-interferon.

  18. Interaction between Salmonella typhimurium and phagocytic cells in pigs - Phagocytosis, oxidative burst and killing in polymorphonuclear leukocytes and monocytes

    DEFF Research Database (Denmark)

    Riber, Ulla; Lind, Peter

    1999-01-01

    Interactions between Salmonella typhimurium and peripheral blood leucocytes from healthy, Salmonella-free pigs were investigated in vitro. Both granulocytes and monocytes phagocytized FITC-labelled heat-killed Salmonella bacteria as shown by flow cytometry. Phagocytosis in whole blood and isolated...

  19. Killing curve activity of ciprofloxacin is comparable to synergistic effect of beta-lactam-tobramycin combinations against Haemophilus species endocarditis strains

    DEFF Research Database (Denmark)

    Westh, H; Frimodt-Møller, N; Gutschik, E

    1992-01-01

    Nine Haemophilus species strains, all beta-lactamase negative, isolated from patients with endocarditis were tested in killing curve experiments. Antibiotics used were penicillin, amoxicillin, aztreonam alone and in combination with tobramycin, as well as ciprofloxacin alone. Synergism between beta......-lactams and tobramycin with reduction of colony counts to zero was seen after 24 h for H. influenzae, H. parainfluenzae and H. segnis strains. Ciprofloxacin was as effective as beta-lactam-tobramycin combinations. The H. aphrophilus strain was not killed as effectively as other strains by any of the antibiotics....

  20. Modulation of Th1/Th2 Immune Responses by Killed Propionibacterium acnes and Its Soluble Polysaccharide Fraction in a Type I Hypersensitivity Murine Model: Induction of Different Activation Status of Antigen-Presenting Cells

    Directory of Open Access Journals (Sweden)

    Carla Cristina Squaiella-Baptistão

    2015-01-01

    Full Text Available Propionibacterium acnes (P. acnes is a gram-positive anaerobic bacillus present in normal human skin microbiota, which exerts important immunomodulatory effects, when used as heat- or phenol-killed suspensions. We previously demonstrated that heat-killed P. acnes or its soluble polysaccharide (PS, extracted from the bacterium cell wall, suppressed or potentiated the Th2 response to ovalbumin (OVA in an immediate hypersensitivity model, depending on the treatment protocol. Herein, we investigated the mechanisms responsible for these effects, using the same model and focusing on the activation status of antigen-presenting cells (APCs. We verified that higher numbers of APCs expressing costimulatory molecules and higher expression levels of these molecules are probably related to potentiation of the Th2 response to OVA induced by P. acnes or PS, while higher expression of toll-like receptors (TLRs seems to be related to Th2 suppression. In vitro cytokines production in cocultures of dendritic cells and T lymphocytes indicated that P. acnes and PS seem to perform their effects by acting directly on APCs. Our data suggest that P. acnes and PS directly act on APCs, modulating the expression of costimulatory molecules and TLRs, and these differently activated APCs drive distinct T helper patterns to OVA in our model.

  1. Killing of Sarcoma Cells by Proapoptotic Bcl-XS: Role of the BH3 Domain and Regulation by Bcl-XL

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    Raj. S. Mitra

    2001-01-01

    Full Text Available Kaposi's sarcoma (KS is the most common tumor affecting AIDS patients with over 20% of these patients afflicted by this disease. Previous studies have demonstrated that KS tumor cells predominantly express the prosurvival protein Bcl-XL compared with Bcl-2. In the current study, we have used an adenoviral vector that expresses Bcl-XS, a functional inhibitor of Bcl-XL, to study the significance of Bcl-XL expression in the KS cell line (SLK or KS primary cultures. The results demonstrate that 75% to 80% of SLK or KS primary cells were killed by the Bcl-XS containing adenovirus whereas KS cells infected with control adenovirus showed no significant cell death or growth inhibition. Overexpression of Bcl-XL, but not Bcl-2, in SILK cells attenuated apoptosis induced by adenovirus Bcl-XS. Immunoprecipitation experiments revealed that adenoviral Bcl-XS associated with Bcl-XL, but not with Bcl-2. Mutational analysis showed that the α2 helical region of Bcl-XS containing the BH3 motif was critical for killing activity and interaction with Bcl-XL. These results suggest that Bcl-XS is a direct killer and Bcl-XL may act by interacting with and sequestering Bcl-XS. These studies also suggest that targeting Bcl-XL may be of therapeutic benefit for the treatment of tumors that are characterized by inappropriate expression of Bcl-XL.

  2. The proteins (12 and 15 kDa) isolated from heat-killed Lactobacillus plantarum L67 induces apoptosis in HT-29 cells.

    Science.gov (United States)

    Song, S; Oh, S; Lim, K T

    2015-03-01

    A number of scientific studies have revealed that Lactobacillus strains have beneficial bioactivities in the gastrointestinal tract. In this study, the production of intracellular reactive oxygen species (ROS) and the amounts of intracellular calcium, protein kinase C activity, cytochrome c, Bid, Bcl-2, Bax and the apoptosis-mediated proteins [caspase-8, caspase-3 and poly ADP ribose polymerase (PARP)] were evaluated to understand the induction of programmed cell death in HT-29 cells by Lactobacillus plantarum L67. The results obtained from this study indicated that the relative intensities of the apoptotic-related factors (intracellular ROS and intracellular calcium) and of apoptotic signals (Bax and t-Bid) increased with increasing concentrations of the membrane proteins isolated from heat-killed L. plantarum L67, whereas the relative intensities of cytochrome c, Bcl-2, caspase-8, caspase-3 and PARP decreased. This study determines whether proteins (12 and 15 kDa) isolated from heat-killed L. plantarum L67 induce programmed cell death in HT-29 cells. Proteins isolated from L. plantarum L67 can stimulate the apoptotic signals and then consequently induce programmed cell death in HT-29 cells. The results in this study suggest that the proteins isolated from L. plantarum L67 could be used as an antitumoural agent in probiotics and as a component of supplements or health foods.

  3. Heat Killed Attenuated Leishmania Induces Apoptosis of HepG2 Cells Through ROS Mediated p53 Dependent Mitochondrial Pathway

    Directory of Open Access Journals (Sweden)

    Dipayan Bose

    2016-03-01

    Full Text Available Background/Aims: Cytotoxic effect of attenuated Leishmania on liver cancer cells by inducing ROS generation. Methods: Spectrophotometric study to analyze cell death and levels of different active caspases. Flow cytometric study was done to analyze apoptosis induction and ROS generation and levels of different protein. Western blot analysis was performed to study the levels of protein. Confocal microscopy was done to ascertain the expression of different apoptotic markers. Results: We have now observed that attenuated Leishmania donovani UR6 also has potentiality towards growth inhibition of HepG2 cells and investigated the mechanism of action. The effect is associated with increased DNA fragmentation, rise in number of annexinV positive cells, and cell cycle arrest at G1 phase. The detection of unregulated levels of active PARP, cleaved caspases 3 and 9, cytosolic cytochrome C, Bax, and Bad, along with the observed downregulation of Bcl-2 and loss of mitochondrial membrane potential suggested the involvement of mitochondrial pathway. Enhanced ROS and p53 levels regulate the apoptosis of HepG2 cells. NAC was found to inhibit p53 production but PFT-α has no effect on ROS generation. In conclusion, Leishmania donovani UR6 efficiently induces apoptosis in HepG2 cells through ROS mediated p53 dependent mitochondrial pathway. Conclusion: It has been reported earlier that some parasites show prominent cytotoxic effect and prevent tumor growth. From our study we found that Leishmania donovani UR6 efficiently induced apoptosis in HepG2 cells through ROS mediated p53 dependent mitochondrial pathway. This study has rejuvenated the age old idea of bio-therapy.

  4. Innate invariant NKT cells recognize Mycobacterium tuberculosis-infected macrophages, produce interferon-gamma, and kill intracellular bacteria.

    Directory of Open Access Journals (Sweden)

    Isabel Sada-Ovalle

    2008-12-01

    Full Text Available Cellular immunity to Mycobacterium tuberculosis (Mtb requires a coordinated response between the innate and adaptive arms of the immune system, resulting in a type 1 cytokine response, which is associated with control of infection. The contribution of innate lymphocytes to immunity against Mtb remains controversial. We established an in vitro system to study this question. Interferon-gamma is produced when splenocytes from uninfected mice are cultured with Mtb-infected macrophages, and, under these conditions, bacterial replication is suppressed. This innate control of bacterial replication is dependent on CD1d-restricted invariant NKT (iNKT cells, and their activation requires CD1d expression by infected macrophages as well as IL-12 and IL-18. We show that iNKT cells, even in limiting quantities, are sufficient to restrict Mtb replication. To determine whether iNKT cells contribute to host defense against tuberculosis in vivo, we adoptively transferred iNKT cells into mice. Primary splenic iNKT cells obtained from uninfected mice significantly reduce the bacterial burden in the lungs of mice infected with virulent Mtb by the aerosol route. Thus, iNKT cells have a direct bactericidal effect, even in the absence of synthetic ligands such as alpha-galactosylceramide. Our finding that iNKT cells protect mice against aerosol Mtb infection is the first evidence that CD1d-restricted NKT cells mediate protection against Mtb in vivo.

  5. Autophagy inhibition contributes to the synergistic interaction between EGCG and doxorubicin to kill the hepatoma Hep3B cells.

    Directory of Open Access Journals (Sweden)

    Li Chen

    Full Text Available (--Epigallocatechin-3-O-gallate(EGCG, the highest catechins from green tea, has promisingly been found to sensitize the efficacy of several chemotherapy agents like doxorubicin (DOX in hepatocellular carcinoma (HCC treatment. However, the detailed mechanisms by which EGCG augments the chemotherapeutic efficacy remain unclear. Herein, this study was designed to determine the synergistic impacts of EGCG and DOX on hepatoma cells and particularly to reveal whether the autophagic flux is involved in this combination strategy for the HCC. Electron microscopy and fluorescent microscopy confirmed that DOX significantly increased autophagic vesicles in hepatoma Hep3B cells. Western blot and trypan blue assay showed that the increasing autophagy flux by DOX impaired about 45% of DOX-induced cell death in these cells. Conversely, both qRT-PCR and western blotting showed that EGCG played dose-dependently inhibitory role in autophagy signaling, and that markedly promoted cellular growth inhibition. Amazingly, the combined treatment caused a synergistic effect with 40 to 60% increment on cell death and about 45% augmentation on apoptosis versus monotherapy pattern. The DOX-induced autophagy was abolished by this combination therapy. Rapamycin, an autophagic agonist, substantially impaired the anticancer effect of either DOX or combination with EGCG treatment. On the other hand, using small interference RNA targeting chloroquine autophagy-related gene Atg5 and beclin1 to inhibit autophagy signal, hepatoma cell death was dramatically enhanced. Furthermore, in the established subcutaneous Hep3B cells xenograft tumor model, about 25% reduction in tumor growth as well as 50% increment of apoptotic cells were found in combination therapy compared with DOX alone. In addition, immunohistochemistry analysis indicated that the suppressed tendency of autophagic hallmark microtubule-associated protein light chain 3 (LC3 expressions was consistent with thus combined

  6. Melphalan, alone or conjugated to an FSH-β peptide, kills murine testicular cells in vitro and transiently suppresses murine spermatogenesis in vivo.

    Science.gov (United States)

    Amory, John K; Hong, SungWoo; Yu, Xiaozhong; Muller, Charles H; Faustman, Elaine; Goldstein, Alex

    2014-07-01

    New approaches to sterilizing male animals are needed to control captive and wild animal populations. We sought to develop a nonsurgical method of permanent sterilization for male animals by administering the gonadotoxicant melphalan conjugated to peptides derived from the β-chain of FSHβ. We hypothesized that conjugating melphalan to FSHβ peptides would magnify the gonadotoxic effects of melphalan while minimizing systemic toxicity. The ability of conjugates of melphalan and FSHβ peptides to kill murine testicular cells was first tested in vitro in a three-dimensional testicular cell coculture system. In this system, melphalan caused considerable cell death as measured both by increases in lactate dehydrogenase concentrations in the culture supernatant and direct visualization of the cultures. Of the conjugates tested, melphalan conjugated to a 20-amino acid peptide derived from human FSHβ consisting of amino acids 33 to 53 (FSHβ (33-53)-melphalan) was very potent, with cell cytotoxicity and lactate dehydrogenase release roughly one-half that of melphalan. The effects of melphalan and FSHβ (33-53)-melphalan on spermatogenesis were then tested in vivo in mature C56Bl/6 male mice. Four weeks after intraperitoneal injection, all mice treated with either FSHβ (33-53)-melphalan or melphalan had approximately 75% reductions in testicular spermatid counts compared with control animals. Testicular histology revealed significant reduction in mature spermatids and spermatocytes in most tubules. However, 12 weeks after the injection, testicular spermatid counts and histology were similar to controls, except in one animal receiving FSHβ (33-53)-melphalan that had no apparent spermatogenesis. We conclude that melphalan and FSHβ (33-53)-melphalan are potent gonadotoxicants in male mice resulting in marked suppression of spermatogenesis 4 weeks after a single intraperitoneal injection. However, this effect is transient in most mice as spermatogenesis is similar to

  7. C16-Ceramide Analog Combined with Pc 4 Photodynamic Therapy Evokes Enhanced Total Ceramide Accumulation, Promotion of DEVDase Activation in the Absence of Apoptosis, and Augmented Overall Cell Killing

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    Duska Separovic

    2011-01-01

    Full Text Available Because of the failure of single modality approaches, combination therapy for cancer treatment is a promising alternative. Sphingolipid analogs, with or without anticancer drugs, can improve tumor response. C16-pyridinium ceramide analog LCL30, was used in combination with photodynamic therapy (PDT, an anticancer treatment modality, to test the hypothesis that the combined treatment will trigger changes in the sphingolipid profile and promote cell death. Using SCCVII mouse squamous carcinoma cells, and the silicone phthalocyanine Pc 4 for PDT, we showed that combining PDT with LCL30 (PDT/LCL30 was more effective than individual treatments in raising global ceramide levels, as well as in reducing dihydrosphingosine levels. Unlike LCL30, PDT, alone or combined, increased total dihydroceramide levels. Sphingosine levels were unaffected by LCL30, but were abolished after PDT or the combination. LCL30-triggered rise in sphingosine-1-phosphate was reversed post-PDT or the combination. DEVDase activation was evoked after PDT or LCL30, and was promoted post- PDT/LCL30. Neither mitochondrial depolarization nor apoptosis were observed after any of the treatments. Notably, treatment with the combination resulted in augmented overall cell killing. Our data demonstrate that treatment with PDT/LCL30 leads to enhanced global ceramide levels and DEVDase activation in the absence of apoptosis, and promotion of total cell killing.

  8. Combining Heavy Ion Radiation and Artificial MicroRNAs to Target the Homologous Recombination Repair Gene Efficiently Kills Human Tumor Cells

    Energy Technology Data Exchange (ETDEWEB)

    Zheng Zhiming [Department of Neurosurgery, Provincial Hospital Affiliated to Shandong University, Shandong University, Jinan (China); Department of Radiation Oncology, School of Medicine, Winship Cancer Institute, Emory University, Atlanta, Georgia (United States); Wang Ping; Wang Hongyan; Zhang Xiangming [Department of Radiation Oncology, School of Medicine, Winship Cancer Institute, Emory University, Atlanta, Georgia (United States); Wang Minli [Division of Life Sciences, Universities Space Research Association, Houston, Texas (United States); Cucinotta, Francis A. [National Aeronautics and Space Administration, Lyndon B. Johnson Space Center, Houston, Texas (United States); Wang Ya, E-mail: ywang94@emory.edu [Department of Radiation Oncology, School of Medicine, Winship Cancer Institute, Emory University, Atlanta, Georgia (United States)

    2013-02-01

    Purpose: Previously, we demonstrated that heavy ions kill more cells at the same dose than X-rays because DNA-clustered lesions produced by heavy ions affect nonhomologous end-joining (NHEJ) repair but not homologous recombination repair (HRR). We have also shown that our designed artificial microRNAs (amiRs) could efficiently target XRCC4 (an essential factor for NHEJ) or XRCC2 (an essential factor for HRR) and sensitize human tumor cells to X-rays. Based on these data, we were interested in testing the hypothesis that combining heavy ions and amiRs to target HRR but not NHEJ should more efficiently kill human tumor cells. Methods and Materials: Human tumor cell lines (U87MG, a brain tumor cell line, and A549, a lung cancer cell line) and their counterparts, overexpressed with amiR to target XRCC2, XRCC4 or both, were used in this study. Survival sensitivities were examined using a clonogenic assay after these cells were exposed to X-rays or heavy ions. In addition, these cell lines were subcutaneously injected into nude mice to form xenografts and the tumor size was compared after the tumor areas were exposed to X-rays or heavy ions. Results: Although targeting either XRCC4 (NHEJ factor) or XRCC2 (HRR factor) sensitized the human tumor cells to X-rays, in vitro and the xenograft animal model, targeting only XRCC2 but not XRCC4 sensitized the human tumor cells to heavy ions in vitro and in the xenograft animal model. Conclusions: Combining heavy ions with targeting the HRR pathway, but not the NHEJ pathway, could significantly improve the efficiency of tumor cell death.

  9. Involvement of superoxide and myeloperoxidase in oxygen-dependent killing of Staphylococcus aureus by neutrophils.

    Science.gov (United States)

    Hampton, M B; Kettle, A J; Winterbourn, C C

    1996-09-01

    We have used a quantitative assay that measures independent rate constants for phagocytosis and killing of Staphylococcus aureus to investigate the involvement of superoxide and myeloperoxidase in bacterial killing by human neutrophils. To inhibit superoxide-dependent processes, superoxide dismutase was cross-linked to immunoglobulin G and the conjugate was attached to the surface of S. aureus via protein A in its cell wall. Myeloperoxidase was inhibited with azide, and myeloperoxidase-deficient neutrophils were used. Adding the NADPH oxidase inhibitor diphenyleneiodonium, to prevent superoxide production, decreased the killing rate to 25%, indicating that oxidative killing mechanisms predominate in this system. The rate constant for killing of S. aureus with superoxide dismutase attached was 70% of that for control bacteria linked to inactivated enzyme. Superoxide dismutase had no effect in the presence of diphenyleneiodonium. The rate of killing was decreased to 33% in the presence of azide and to 40% with myeloperoxidase-deficient neutrophils. Superoxide dismutase had no effect in the presence of azide. On the assumption that the oxidative and nonoxidative components of killing can be considered separately, the oxidative rate was decreased by almost half by superoxide dismutase and was about six times lower when myeloperoxidase was inactive. We conclude that myeloperoxidase-dependent processes are strongly favored by human neutrophils as their prime mechanism of oxidative killing of S. aureus and that superoxide makes a direct contribution to killing. Our results also suggest that superoxide acts in conjunction with a myeloperoxidase-dependent pathway.

  10. Selective killing of human immunodeficiency virus infected cells by non-nucleoside reverse transcriptase inhibitor-induced activation of HIV protease

    Directory of Open Access Journals (Sweden)

    Smeulders Liesbeth

    2010-10-01

    Full Text Available Abstract Background Current antiretroviral therapy against human immunodeficiency virus (HIV-1 reduces viral load and thereby prevents viral spread, but it cannot eradicate proviral genomes from infected cells. Cells in immunological sanctuaries as well as cells producing low levels of virus apparently contribute to a reservoir that maintains HIV persistence in the presence of highly active antiretroviral therapy. Thus, accelerated elimination of virus producing cells may represent a complementary strategy to control HIV infection. Here we sought to exploit HIV protease (PR related cytotoxicity in order to develop a strategy for drug induced killing of HIV producing cells. PR processes the viral Gag and Gag-Pol polyproteins during virus maturation, but is also implicated in killing of virus producing cells through off-target cleavage of host proteins. It has been observed previously that micromolar concentrations of certain non-nucleoside reverse transcriptase inhibitors (NNRTIs can stimulate intracellular PR activity, presumably by enhancing Gag-Pol dimerization. Results Using a newly developed cell-based assay we compared the degree of PR activation displayed by various NNRTIs. We identified inhibitors showing higher potency with respect to PR activation than previously described for NNRTIs, with the most potent compounds resulting in ~2-fold increase of the Gag processing signal at 250 nM. The degree of enhancement of intracellular Gag processing correlated with the compound's ability to enhance RT dimerization in a mammalian two-hybrid assay. Compounds were analyzed for their potential to mediate specific killing of chronically infected MT-4 cells. Levels of cytotoxicity on HIV infected cells determined for the different NNRTIs corresponded to the relative degree of drug induced intracellular PR activation, with CC50 values ranging from ~0.3 μM to above the tested concentration range (10 μM. Specific cytotoxicity was reverted by addition

  11. Effects of growth temperature on the ingestion and killing of clinical isolates of Listeria monocytogenes by human neutrophils.

    OpenAIRE

    Stecha, P F; Heynen, C A; Roll, J T; Brown, J F; Czuprynski, C J

    1989-01-01

    In this study, we compared three human isolates (F5380, Scott A, and Murray B) and one laboratory strain (EGD) of Listeria monocytogenes for their resistance to ingestion and killing by human neutrophils. We observed no substantial difference in killing among these strains when they were grown at 37 degrees C. Because it is likely that listerial growth occurs at lower temperatures during food-borne outbreaks of listeriosis, we also compared these strains after they were grown at 22 and 4 degr...

  12. Effect of a killed whole Neospora caninum tachyzoite vaccine on the crube abortion rate of Costa Rican dairy cows under field conditions

    NARCIS (Netherlands)

    Romero, J.J.; Perez, E.; Frankena, K.

    2004-01-01

    A standard field trial was carried out to assess the effect of a commercial Neospora-vaccine based on whole killed tachyzoites (Bovilis–Neoguard, Intervet®) on the abortion rate. Eight hundred and seventy-six cows, over 2.5 months pregnant, belonging to 25 Costa Rican dairy herds, were used in the a

  13. Tetrandrine, a Compound Common in Chinese Traditional Medicine, Preferentially Kills Breast Cancer Tumor Initiating Cells (TICs) In Vitro

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Wei; Debeb, Bisrat G.; Lacerda, Lara; Li, Jessica; Woodward, Wendy A., E-mail: wwoodward@mdanderson.org [Division of Radiation Oncology, University of Texas M.D. Anderson Cancer Center, Houston, TX 77030 (United States)

    2011-05-04

    Tetrandrine is a bisbenzylisoquinoline alkaloid found in Stephania tetrandra, a Chinese medicine commonly used as an anti-inflammatory. It has extensive pharmacological activity, including positive ion channel blockade and inhibition of multiple drug resistance proteins. These activities are very similar to that of salinomycin, a known drug targeting breast cancer initiation cells (TICs). Herein, we tested tetrandrine targeting of breast cancer TICs. SUM-149, an inflammatory breast cancer cell line and SUM-159, a non-inflammatory metaplastic breast cancer cell line were used in these studies. In proliferation assays using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl) -2H-tetrazolium (MTS), we found that the IC{sub 50} for inhibition of proliferation is 15.3 ± 4.1 μM for SUM-149 and 24.3 ± 2.1 μM for SUM-159 cells. Tetrandrine also inhibited mammosphere formation, a surrogate for breast cancer TICs growth in vitro with IC{sub 50} around 1 μM for SUM-149 and around 2 μM for SUM-159 cells. Tetrandrine has similar effects on the mammosphere formation from cells isolated from fresh patient sample. Moreover, tetrandrine decreases the aldehyde dehydrogenase (ALDH) positive population in SUM-159 by 45% ± 5.45% P = 0.005. In summary, tetrandrine demonstrates significant efficacy against in vitro surrogates for inflammatory and aggressive breast cancer TICs.

  14. Tetrandrine, a Compound Common in Chinese Traditional Medicine, Preferentially Kills Breast Cancer Tumor Initiating Cells (TICs In Vitro

    Directory of Open Access Journals (Sweden)

    Jessica Li

    2011-05-01

    Full Text Available Tetrandrine is a bisbenzylisoquinoline alkaloid found in Stephania tetrandra, a Chinese medicine commonly used as an anti-inflammatory. It has extensive pharmacological activity, including positive ion channel blockade and inhibition of multiple drug resistance proteins. These activities are very similar to that of salinomycin, a known drug targeting breast cancer initiation cells (TICs. Herein, we tested tetrandrine targeting of breast cancer TICs. SUM-149, an inflammatory breast cancer cell line and SUM-159, a non-inflammatory metaplastic breast cancer cell line were used in these studies. In proliferation assays using 3-(4,5-dimethylthiazol-2-yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H-tetrazolium (MTS, we found that the IC50 for inhibition of proliferation is 15.3 ± 4.1 µM for SUM-149 and 24.3 ± 2.1 µM for SUM-159 cells. Tetrandrine also inhibited mammosphere formation, a surrogate for breast cancer TICs growth in vitro with IC50 around 1 µM for SUM-149 and around 2 µM for SUM-159 cells. Tetrandrine has similar effects on the mammosphere formation from cells isolated from fresh patient sample. Moreover, tetrandrine decreases the aldehyde dehydrogenase (ALDH positive population in SUM-159 by 45% ± 5.45% P = 0.005. In summary, tetrandrine demonstrates significant efficacy against in vitro surrogates for inflammatory and aggressive breast cancer TICs.

  15. Enhanced killing of androgen-independent prostate cancer cells using inositol hexakisphosphate in combination with proteasome inhibitors.

    Science.gov (United States)

    Diallo, J-S; Betton, B; Parent, N; Péant, B; Lessard, L; Le Page, C; Bertrand, R; Mes-Masson, A-M; Saad, F

    2008-11-18

    Effective treatments for androgen-independent prostate cancer (AIPCa) are lacking. To address this, emerging therapeutics such as proteasome inhibitors are currently undergoing clinical trials. Inositol hexakisphosphate (IP6) is an orally non-toxic phytochemical that exhibits antitumour activity against several types of cancer including PCa. We have previously shown that treatment of PC3 cells with IP6 induces the transcription of a subset of nuclear factor-kappaB (NF-kappaB)-responsive and pro-apoptotic BCL-2 family genes. In this study, we report that although NF-kappaB subunits p50/p65 translocate to the nucleus of PC3 cells in response to IP6, inhibition of NF-kappaB-mediated transcription using non-degradable inhibitor of kappaB (IkappaB)-alpha does not modulate IP6 sensitivity. Treatment with IP6 also leads to increased protein levels of PUMA, BIK/NBK and NOXA between 4 and 8 h of treatment and decreased levels of MCL-1 and BCL-2 after 24 h. Although blocking transcription using actinomycin D does not modulate PC3 cell sensitivity to IP6, inhibition of protein translation using cycloheximide has a significant protective effect. In contrast, blocking proteasome-mediated protein degradation using MG-132 significantly enhances the ability of IP6 to reduce cellular metabolic activity in both PC3 and DU145 AIPCa cell lines. This effect of combined treatment on mitochondrial depolarisation is particularly striking and is also reproduced by another proteasome inhibitor (ALLN). The enhanced effect of combined MG132/IP6 treatment is almost completely inhibited by cycloheximide and correlates with changes in BCL-2 family protein levels. Altogether these results suggest a role for BCL-2 family proteins in mediating the combined effect of IP6 and proteasome inhibitors and warrant further pre-clinical studies for the treatment of AIPCa.

  16. Defects in the acquisition of tumor-killing capability of CD8+ cytotoxic T cells in streptozotocin-induced diabetic mice.

    Directory of Open Access Journals (Sweden)

    Shu-Ching Chen

    Full Text Available Emerging evidences have shown that diabetes mellitus not only raises risk but also heightens mortality rate of cancer. It is not clear, however, whether antitumor CD8+ cytotoxic T lymphocyte (CTL response is down-modulated in diabetic hosts. We investigated the impact of hyperglycemia on CTLs' acquisition of tumor-killing capability by utilizing streptozotocin-induced diabetic (STZ-diabetic mice. Murine diabetes was induced by intraperitoneal injection of STZ (200 mg/kg in C57BL/6 mice, 2C-T cell receptor (TCR transgenic and P14-TCR transgenic mice. The study found that, despite harboring intact proliferative capacity measured with CFSE labeling and MTT assay, STZ-diabetic CD8+ CTLs displayed impaired effector functions. After stimulation, STZ-diabetic CD8+ CTLs produced less perforin and TNFα assessed by intracellular staining, as well as expressed less CD103 protein. Furthermore, adoptive transfer of STZ-diabetic P14 CD8+ effector cells showed an insufficient recruitment to the B16.gp33 melanoma and inadequate production of perforin, granzyme B and TNFα determined by immunohistochemistry in the tumor milieu. As a result, STZ-diabetic CD8+ effector cells were neither able to eliminate tumor nor to improve survival of tumor-bearing mice. Taken together, our data suggest that CD8+ CTLs are crippled to infiltrate into tumors and thus fail to acquire tumor-killing capability in STZ-diabetic hosts.

  17. Effect of sociality and season on gray wolf (Canis lupus foraging behavior: implications for estimating summer kill rate.

    Directory of Open Access Journals (Sweden)

    Matthew C Metz

    Full Text Available BACKGROUND: Understanding how kill rates vary among seasons is required to understand predation by vertebrate species living in temperate climates. Unfortunately, kill rates are only rarely estimated during summer. METHODOLOGY/PRINCIPAL FINDINGS: For several wolf packs in Yellowstone National Park, we used pairs of collared wolves living in the same pack and the double-count method to estimate the probability of attendance (PA for an individual wolf at a carcass. PA quantifies an important aspect of social foraging behavior (i.e., the cohesiveness of foraging. We used PA to estimate summer kill rates for packs containing GPS-collared wolves between 2004 and 2009. Estimated rates of daily prey acquisition (edible biomass per wolf decreased from 8.4±0.9 kg (mean ± SE in May to 4.1±0.4 kg in July. Failure to account for PA would have resulted in underestimating kill rate by 32%. PA was 0.72±0.05 for large ungulate prey and 0.46±0.04 for small ungulate prey. To assess seasonal differences in social foraging behavior, we also evaluated PA during winter for VHF-collared wolves between 1997 and 2009. During winter, PA was 0.95±0.01. PA was not influenced by prey size but was influenced by wolf age and pack size. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that seasonal patterns in the foraging behavior of social carnivores have important implications for understanding their social behavior and estimating kill rates. Synthesizing our findings with previous insights suggests that there is important seasonal variation in how and why social carnivores live in groups. Our findings are also important for applications of GPS collars to estimate kill rates. Specifically, because the factors affecting the PA of social carnivores likely differ between seasons, kill rates estimated through GPS collars should account for seasonal differences in social foraging behavior.

  18. Irreducible Killing Tensors from Third Rank Killing-Yano Tensors

    OpenAIRE

    Popa, Florian Catalin; Tintareanu-Mircea, Ovidiu

    2006-01-01

    We investigate higher rank Killing-Yano tensors showing that third rank Killing-Yano tensors are not always trivial objects being possible to construct irreducible Killing tensors from them. We give as an example the Kimura IIC metric were from two rank Killing-Yano tensors we obtain a reducible Killing tensor and from third rank Killing-Yano tensors we obtain three Killing tensors, one reducible and two irreducible.

  19. Central memory Vgamma9Vdelta2 T lymphocytes primed and expanded by bacillus Calmette-Guérin-infected dendritic cells kill mycobacterial-infected monocytes.

    Science.gov (United States)

    Martino, Angelo; Casetti, Rita; Sacchi, Alessandra; Poccia, Fabrizio

    2007-09-01

    In humans, innate immune recognition of mycobacteria, including Mycobacterium tuberculosis and bacillus Calmette-Guérin (BCG), is a feature of cells as dendritic cells (DC) and gammadelta T cells. In this study, we show that BCG infection of human monocyte-derived DC induces a rapid activation of Vgamma9Vdelta2 T cells (the major subset of gammadelta T cell pool in human peripheral blood). Indeed, in the presence of BCG-infected DC, Vgamma9Vdelta2 T cells increase both their expression of CD69 and CD25 and the production of TNF-alpha and IFN-gamma, in contrast to DC treated with Vgamma9Vdelta2 T cell-specific Ags. Without further exogenous stimuli, BCG-infected DC expand a functionally cytotoxic central memory Vgamma9Vdelta2 T cell population. This subset does not display lymph node homing receptors, but express a high amount of perforin. They are highly efficient in the killing of mycobacterial-infected primary monocytes or human monocytic THP-1 cells preserving the viability of cocultured, infected DC. This study provides further evidences about the complex relationship between important players of innate immunity and suggests an immunoregulatory role of Vgamma9Vdelta2 T cells in the control of mycobacterial infection.

  20. Effectiveness of Linezolid, 127I-Linezolid and 131I-Linezolid Against Methicillin-Susceptible Staphylococcus Aureus by Time Kill Curve Methods

    Directory of Open Access Journals (Sweden)

    Hasan Demiroğlu

    2015-02-01

    Full Text Available Objective: Linezolid (LNZ is one of the most effective treatments against Gram positive bacteria. However LNZ resistant intermediate strains have recently emerged in worldwide. The aim of the study was to compare the minimum inhibitory concentration (MIC, minimum bactericidal concentration (MBC and minimum biofilm inhibitory concentration (MBIC of LNZ, 127I-LNZ and 131I-LNZ against methicillin susceptible Staphylococcus aureus ATCC 35556 (MSSA biofilms. Methods: LNZ radiolabeled with 131I and cold labeling study with 127I was performed. Radiolabeling and inactive labeling quality-control studies of LNZ were carried out by using TLC (Thin Layer Radiochromatography and HPLC (High Pressure Liquid Chromatography. LNZ, 127I-LNZ and 131I-LNZ against biofilm-forming MSSA was investigated, using a twofold serial broth microtiter method, biofilm challenge, and bacterial count recovery. Results: The binding yield was obtained to be about 86±2% for radiolabeled LNZ. Minimal inhibitory concentration (MIC and minimal bactericidal concentration for LNZ, 127I-LNZ and 131I-LNZ ranged from 1 to 2 µg/mL respectively. In time-kill studies LNZ, 127I-LNZ and 131I-LNZ were bactericidal against staphylococci, producing ≥3 Log10 decrease in viable counts (cfu/mL within 6 h at 2xMIC. Following the biofilm formation on polystyrene U-bottom microtiter plates to investigate the minimal biofilm inhibitory concentration (MBIC of LNZ, 127I-LNZ and 131I-LNZ was defined as the minimal concentration of antibiotic required to inhibite the biofilm. None of the LNZ, 127I-LNZ and 131I-LNZ killed 100% of biofilm associated cells. Mean cell survival in biofilms treated with 64 µg/mL LNZ, 127I-LNZ and 131I-LNZ (64 µg/mL was 48%, 49%, and 33%, respectively. Conclusion: Our results show that radiolabeled Linezolid demonstrated that 24 h of exposure to 64 µg/mL, promise in treating biofilm producing Staphylococcus aureus.

  1. Critical high-dimensional state transitions in cell populations or why cancers follow the principle ``What does not kill me makes me stronger''

    Science.gov (United States)

    Huang, Sui

    Transitions between high-dimensional attractor states in the quasi-potential landscape of the gene regulatory network, induced by environmental perturbations and/or facilitated by mutational rewiring of the network, underlie cell phenotype switching in development as well as in cancer progression, including acquisition of drug-resistant phenotypes. Considering heterogeneous cell populations as statistical ensembles of cells, and single-cell resolution gene expression profiling of cell populations undergoing a cell phenotype shift allow us now to map the topography of the landscape and its distortion. From snapshots of single-cell expression patterns of a cell population measured during major transitions we compute a quantity that identifies symmetry-breaking destabilization of attractors (bifurcation) and concomitant dimension-reduction of the state space manifold (landscape distortion) which precede critical transitions to new attractor states. The model predicts, and we show experimentally, the almost inevitable generation of aberrant cells associated with such critical transitions in multi-attractor landscapes: therapeutic perturbations which seek to push cancer cells to the apoptotic state, almost always produce ``rebellious'' cells which move in the ``opposite direction'': instead of dying they become more stem-cell-like and malignant. We show experimentally that the inadvertent generation of more malignant cancer cells by therapy indeed results from transition of surviving (but stressed) cells into unforeseen attractor states and not simply from selection of inherently more resistant cells. Thus, cancer cells follow not so much Darwin, as generally thought (survival of the fittest), but rather Nietzsche (What does not kill me makes me stronger). Supported by NIH (NCI, NIGMS), Alberta Innovates.

  2. Antibodies against invasive phenotype-specific antigens increase Mycobacterium avium subspecies paratuberculosis translocation across a polarized epithelial cell model and enhance killing by bovine macrophages

    Science.gov (United States)

    Everman, Jamie L.; Bermudez, Luiz E.

    2015-01-01

    Johne's disease, caused by Mycobacterium avium subspecies paratuberculosis (MAP), is a severe chronic enteritis which affects large populations of ruminants globally. Prevention strategies to combat the spread of Johne's disease among cattle herds involve adhering to strict calving practices to ensure young susceptible animals do not come in contact with MAP-contaminated colostrum, milk, or fecal material. Unfortunately, the current vaccination options available are associated with high cost and suboptimal efficacy. To more successfully combat the spread of Johne's disease to young calves, an efficient method of protection is needed. In this study, we examined passive immunization as a mode of introducing protective antibodies against MAP to prevent the passage of the bacterium to young animals via colostrum and milk. Utilizing the infectious MAP phenotype developed after bacterial exposure to milk, we demonstrate that in vitro opsonization with serum from Johne's-positive cattle results in enhanced translocation across a bovine MDBK polarized epithelial cell monolayer. Furthermore, immune serum opsonization of MAP results in a rapid host cell-mediated killing by bovine macrophages in an oxidative-, nitrosative-, and extracellular DNA trap-independent manner. This study illustrates that antibody opsonization of MAP expressing an infectious phenotype leads to the killing of the bacterium during the initial stage of macrophage infection. PMID:26301206

  3. Water disinfection using silver nanoparticle impregnated activated carbon: Escherichia coli cell-killing in batch and continuous packed column operation over a long duration.

    Science.gov (United States)

    Biswas, Pritam; Bandyopadhyaya, Rajdip

    2016-09-01

    Silver nanoparticles (Ag-NP) were selectively impregnated on the external surface of plasma treated activated carbon (AC) granules (referred to as Ag-AC hybrid, having 0.8 wt% of Ag), for achieving continuous disinfection of water in a single flow-column set-up. First, Ag-NPs (28 nm mean size) were synthesized by UV reduction. Subsequently, Escherichia coli cell-killing experiments were performed in both shake flask (i. e. batch-mode) and flow-column (i. e. continuous-mode) operations, using E. coli K12 (MTCC 1302) as a model organism. Batch results using 8 mg Ag-AC hybrid/ml of cell suspension showed that, 10(4) CFU/ml of cells were killed within 25 min contact time, with cell concentration decaying exponentially in time. Maintaining almost the same contact time as in the batch experiments, three columns packed with Ag-AC (all having a height of 25 cm but increasing diameters of 1, 5 and 8 cm, respectively) were used for monitoring cell-killing performance over a long duration. For all columns, inlet water having 10(4) CFU/ml E. coli could be completely disinfected to produce treated, outlet water having zero cell count. Specifically for the 8 cm diameter column, a maximum throughput of treating 1.62 L of contaminated water per hour could be maintained for at least up to 16 days. Moreover, the Ag concentration in the outlet water was only up to 29.8 μg/L at steady state, which is well within the recommended limit of 100 μg/L for drinking water. Hence, water disinfection for potable quality water (zero E. coli count and <100 μg/L Ag) can be achieved in a continuous manner over a long duration, with our packed Ag-AC column.

  4. Antimicrobial peptides effectively kill a broad spectrum of Listeria monocytogenes and Staphylococcus aureus strains independently of origin, sub-type, or virulence factor expression

    DEFF Research Database (Denmark)

    Gottlieb, Caroline Trebbien; Thomsen, L.E.; Ingmer, H.;

    2008-01-01

    ) to the human β-defensin 3 (HBD-3). All strains were inhibited by concentrations of hydrogen peroxide between 0.1% – 1.0%. Sub-selections of both species differed in expression of several virulence-related factors and in their ability to survive in human whole blood and kill the nematode virulence model...... Caenorhabditis elegans. For L. monocytogenes, proliferation in whole blood was paralleled by high invasion in Caco-2 cells and fast killing of C. elegans, however, no such pattern in phenotypic behavior was observed for S. aureus and none of the phenotypic differences were correlated to sensitivity to HDPs......Background Host defense peptides (HDPs), or antimicrobial peptides (AMPs), are important components of the innate immune system that bacterial pathogens must overcome to establish an infection and HDPs have been suggested as novel antimicrobial therapeutics in treatment of infectious diseases...

  5. How to kill creativity.

    Science.gov (United States)

    Amabile, T M

    1998-01-01

    In today's knowledge economy, creativity is more important than ever. But many companies unwittingly employ managerial practices that kill it. How? By crushing their employees' intrinsic motivation--the strong internal desire to do something based on interests and passions. Managers don't kill creativity on purpose. Yet in the pursuit of productivity, efficiency, and control--all worthy business imperatives--they undermine creativity. It doesn't have to be that way, says Teresa Amabile. Business imperatives can comfortably coexist with creativity. But managers will have to change their thinking first. Specifically, managers will need to understand that creativity has three parts: expertise, the ability to think flexibly and imaginatively, and motivation. Managers can influence the first two, but doing so is costly and slow. It would be far more effective to increase employees' intrinsic motivation. To that end, managers have five levers to pull: the amount of challenge they give employees, the degree of freedom they grant around process, the way they design work groups, the level of encouragement they give, and the nature of organizational support. Take challenge as an example. Intrinsic motivation is high when employees feel challenged but not overwhelmed by their work. The task for managers, therefore, becomes matching people to the right assignments. Consider also freedom. Intrinsic motivation--and thus creativity--soars when managers let people decide how to achieve goals, not what goals to achieve. Managers can make a difference when it comes to employee creativity. The result can be truly innovative companies in which creativity doesn't just survive but actually thrives.

  6. OBP-401-GFP telomerase-dependent adenovirus illuminates and kills high-metastatic more effectively than low-metastatic triple-negative breast cancer in vitro.

    Science.gov (United States)

    Yano, S; Takehara, K; Kishimoto, H; Tazawa, H; Urata, Y; Kagawa, S; Bouvet, M; Fujiwara, T; Hoffman, R M

    2017-02-01

    We previously described the development of a highly-invasive, triple-negative breast cancer (TNBC) variant using serial orthotopic implantation of MDA-MB-231 human breast cancer in nude mice. The isolated variant is highly invasive in the mammary gland and metastasized to lymph nodes in 10 of 12 mice compared with 2 of 12 of the parental cell line. OBP-401 is a telomerase-dependent cancer-specific, green fluorescent protein (GFP)-expressing adenovirus. OBP-401 was used to infect parental MDA-MB-231P cells and high-metastatic MDA-MB-231H and MDA-MB-231HLN isolated from a lymph node metastasis and MDA-MB-231HLM isolated from a lung metastasis. Time-course imaging showed that OBP-401 labeled MDA-MB-231HP, MDA-MB-231HLN, and MDA-MB-231HLM cells more brightly than MDA-MB-231 parental cells. OBP-401 killed MDA-MB-231H, MDA-MB-231HLN, and MDA-MB-231HLM cells more efficiently than MDA-MB-231P parental cells. These results indicate that OBP-401 could infect, label and then kill high-metastatic MDA-MB-231 more efficiently than low-metastatic MDA-MB-231.

  7. S. Typhimurium strategies to resist killing by cationic antimicrobial peptides.

    Science.gov (United States)

    Matamouros, Susana; Miller, Samuel I

    2015-11-01

    S. Typhimurium is a broad host range Gram-negative pathogen that must evade killing by host innate immune systems to colonize, replicate, cause disease, and be transmitted to other hosts. A major pathogenic strategy of Salmonellae is entrance, survival, and replication within eukaryotic cell phagocytic vacuoles. These phagocytic vacuoles and gastrointestinal mucosal surfaces contain multiple cationic antimicrobial peptides (CAMPs) which control invading bacteria. S. Typhimurium possesses several key mechanisms to resist killing by CAMPs which involve sensing CAMPs and membrane damage to activate signaling cascades that result in remodeling of the bacterial envelope to reduce its overall negative charge with an increase in hydrophobicity to decrease binding and effectiveness of CAMPs. Moreover Salmonellae have additional mechanisms to resist killing by CAMPs including an outer membrane protease which targets cationic peptides at the surface, and specific efflux pumps which protect the inner membrane from damage. This article is part of a Special Issue entitled: Bacterial Resistance to Antimicrobial Peptides.

  8. Enhanced Cancer Cell (HeLa Killing Efficacy of Mixed Αlpha and Gamma Iron Oxide Superparamagnetic Nanoparticles under Combined AC (Alternating Current Magnetic-Field and Photoexcitation

    Directory of Open Access Journals (Sweden)

    Md. Shariful Islam, Yoshihumi Kusumoto, Md. Abdulla Al-Mamun and Yuji Horie

    2011-12-01

    Full Text Available We synthesized mixed α and γ-Fe2O3 nanoparticles and investigated their toxic effects against HeLa cells under induced AC (alternating current magnetic-fields and photoexcited conditions at room temperature. The findings revealed that the cell-killing percentage was increased with increasing dose for all types of treatments. Finally, 99% cancer cells were destructed at 1.2 mL dose when exposed to combined AC magnetic-field and photoexcited conditions (T3 whereas 89 and 83 % of HeLa cells were killed under only AC magnetic-field induced (T1 or only photoexcited (T2 condition at the same dose.ABSTRAK: Campuran α dan zarah γ-Fe2O3 bersaiz nano disintesiskan dan kesan toksidnya terhadap sel HeLa dikaji dibawah aruhan medan magnet arus ulang-alik (alternating current (AC dan keadaan photoexcited (proses ransangan atom atau molekul suatu bahan dengan penyerapan tenaga sinaran pada suhu bilik. Penemuan mendedahkan bahawa peratusan sel yang musnah bertambah dengan pertambahan dos untuk semua jenis rawatan. Akhirnya, 99% sel kanser dimusnahkan pada kadar dos 1.2mL setelah didedahkan terhadap kombinasi medan magnet AC dan keadaan photoexcited (T3 dimana 89% dan 83% sel HeLa dimusnahkan dengan hanya di bawah aruhan medan magnet AC (T1 atau hanya pada keadaan photoexcited (T2 pada kadar dos yang sama.KEY WORDS : Cancer, Hyperthermia, Iron oxide nanoparticles, Heat dissipation,    Cytotoxicity, HeLa cell.

  9. Maximal killing of lymphoma cells by DNA damage-inducing therapy requires not only the p53 targets Puma and Noxa, but also Bim.

    Science.gov (United States)

    Happo, Lina; Cragg, Mark S; Phipson, Belinda; Haga, Jon M; Jansen, Elisa S; Herold, Marco J; Dewson, Grant; Michalak, Ewa M; Vandenberg, Cassandra J; Smyth, Gordon K; Strasser, Andreas; Cory, Suzanne; Scott, Clare L

    2010-12-09

    DNA-damaging chemotherapy is the backbone of cancer treatment, although it is not clear how such treatments kill tumor cells. In nontransformed lymphoid cells, the combined loss of 2 proapoptotic p53 target genes, Puma and Noxa, induces as much resistance to DNA damage as loss of p53 itself. In Eμ-Myc lymphomas, however, lack of both Puma and Noxa resulted in no greater drug resistance than lack of Puma alone. A third B-cell lymphoma-2 homology domain (BH)3-only gene, Bim, although not a direct p53 target, was up-regulated in Eμ-Myc lymphomas incurring DNA damage, and knockdown of Bim levels markedly increased the drug resistance of Eμ-Myc/Puma(-/-)Noxa(-/-) lymphomas both in vitro and in vivo. Remarkably, c-MYC-driven lymphoma cell lines from Noxa(-/-)Puma(-/-)Bim(-/-) mice were as resistant as those lacking p53. Thus, the combinatorial action of Puma, Noxa, and Bim is critical for optimal apoptotic responses of lymphoma cells to 2 commonly used DNA-damaging chemotherapeutic agents, identifying Bim as an additional biomarker for treatment outcome in the clinic.

  10. Effects of killing Helicobacter pylori quadruple therapy on peptic ulcer: A randomized double-blind clinical trial

    Institute of Scientific and Technical Information of China (English)

    Li-Ying Feng; Xi-Xian Yao; Shu-Lin Jiang

    2005-01-01

    AIM: To study the therapeutic efficacy of a Chinese and Western integrated regimen, killing Helicobacter pylori quadruple therapy on H pylori-associated peptic ulcers(PU).METHODS: With prospective and double-blind controlled method, seventy-five active PU patients with H pylori infection were randomized to receive one of the following three regimens: (1) new triple therapy (group A:lansoprazole 30 mg qd, plus clarithromycin 250 mg bid,plus amoxycillin 500 mg tid, each for 10 d); (2) killing Hp quadruple therapy(group B: the three above drugs plus killing H pylori capsule 6 capsules bid for 4 wk) and (3)placebo(group C: gastropine 3 tablets bid for 4 wk).H pylori eradication and ulcer healing quality were evaluated under an endoscope 4 wk after treatment. The patients were followed up for 5 years.RESULTS: Both the healing rate of PU and H pylori eradication rate in group B were significantly higher than those in group C (100% and 96.4% vs 20% and 0%,respectively, P<0.005), but there was no significant difference compared to those in group A (88% and92%, P>0.05). The healing quality of ulcer in group B was superior to that in groups C and A (P<0.05). The recurrence rate of PU in group B (4%) was lower than that in group A (10%) and group C (100%, P<0.01).CONCLUSION: Killing Helicobacter pylori quadruple therapy can not only promote the eradication of H pylori and healing quality of ulcer but also reduce recurrence rate of ulcer.

  11. Isolation and killing of candidate chronic myeloid leukemia stem cells by antibody targeting of IL-1 receptor accessory protein

    DEFF Research Database (Denmark)

    Järås, Marcus; Johnels, Petra; Hansen, Nils Gunder;

    2010-01-01

    will require full eradication of Ph chromosome-positive (Ph(+)) CML stem cells. Here we used gene-expression profiling to identify IL-1 receptor accessory protein (IL1RAP) as up-regulated in CML CD34(+) cells and also in cord blood CD34(+) cells as a consequence of retroviral BCR/ABL1 expression. To test...... their Ph-chromosome status. Interestingly, we found that the CML CD34(+)CD38(-)IL1RAP(+) cells were Ph(+), whereas CML CD34(+)CD38(-)IL1RAP(-) cells were almost exclusively Ph(-). By performing long-term culture-initiating cell assays on the two cell populations, we found that Ph(+) and Ph(-) candidate CML...

  12. Fungal and bacterial killing by neutrophils.

    Science.gov (United States)

    Ermert, David; Zychlinsky, Arturo; Urban, Constantin

    2009-01-01

    Neutrophils are professional phagocytes of the innate immune system that are essential to control bacterial and fungal infections. These cells engulf and kill invading microbes. Additionally, activated neutrophils are able to release neutrophil extracellular traps (NETs). These fibers consist of chromatin decorated with antimicrobial proteins to trap and kill microbes. Appropriate quantitative methods are required to understand the nature of interactions of neutrophils with pathogens. Here we present assays to measure killing mediated by phagocytosis, by NETs, by a combination of both, and by granular extract. As examples, we use Candida albicans for fungal and Shigella flexneri for bacterial pathogens.

  13. A Subpopulation of the K562 Cells Are Killed by Curcumin Treatment after G2/M Arrest and Mitotic Catastrophe

    Science.gov (United States)

    Martinez-Castillo, Macario; Bonilla-Moreno, Raul; Aleman-Lazarini, Leticia; Meraz-Rios, Marco Antonio; Orozco, Lorena; Cedillo-Barron, Leticia; Cordova, Emilio J.

    2016-01-01

    Curcumin is extensively investigated as a good chemo-preventive agent in the development of many cancers and particularly in leukemia, including treatment of chronic myelogenous leukemia and it has been proposed as an adjuvant for leukemia therapies. Human chronic myeloid leukemia cells (K562), were treated with 20 μM of curcumin, and we found that a subpopulation of these cells were arrested and accumulate in the G2/M phase of the cell cycle. Characterization of this cell subpopulation showed that the arrested cells presented nuclear morphology changes resembling those described for mitotic catastrophe. Mitotic cells displayed abnormal chromatin organization, collapse of the mitotic spindle and abnormal chromosome segregation. Then, these cells died in an apoptosis dependent manner and showed diminution in the protein levels of BCL-2 and XIAP. Moreover, our results shown that a transient activation of the nuclear factor κB (NFκB) occurred early in these cells, but decreased after 6 h of the treatment, explaining in part the diminution of the anti-apoptotic proteins. Additionally, P73 was translocated to the cell nuclei, because the expression of the C/EBPα, a cognate repressor of the P73 gene, was decreased, suggesting that apoptosis is trigger by elevation of P73 protein levels acting in concert with the diminution of the two anti-apoptotic molecules. In summary, curcumin treatment might produce a P73-dependent apoptotic cell death in chronic myelogenous leukemia cells (K562), which was triggered by mitotic catastrophe, due to sustained BAX and survivin expression and impairment of the anti-apoptotic proteins BCL-2 and XIAP. PMID:27832139

  14. T cells infiltrate the liver and kill hepatocytes in HLA-B(∗)57:01-associated floxacillin-induced liver injury.

    Science.gov (United States)

    Wuillemin, Natascha; Terracciano, Luigi; Beltraminelli, Helmut; Schlapbach, Christoph; Fontana, Stefano; Krähenbühl, Stephan; Pichler, Werner J; Yerly, Daniel

    2014-06-01

    Drug-induced liver injury is a major safety issue. It can cause severe disease and is a common cause of the withdrawal of drugs from the pharmaceutical market. Recent studies have identified the HLA-B(∗)57:01 allele as a risk factor for floxacillin (FLUX)-induced liver injury and have suggested a role for cytotoxic CD8(+) T cells in the pathomechanism of liver injury caused by FLUX. This study aimed to confirm the importance of FLUX-reacting cytotoxic lymphocytes in the pathomechanism of liver injury and to dissect the involved mechanisms of cytotoxicity. IHC staining of a liver biopsy from a patient with FLUX-induced liver injury revealed periportal inflammation and the infiltration of cytotoxic CD3(+) CD8(+) lymphocytes into the liver. The infiltration of cytotoxic lymphocytes into the liver of a patient with FLUX-induced liver injury demonstrates the importance of FLUX-reacting T cells in the underlying pathomechanism. Cytotoxicity of FLUX-reacting T cells from 10 HLA-B(∗)57:01(+) healthy donors toward autologous target cells and HLA-B(∗)57:01-transduced hepatocytes was analyzed in vitro. Cytotoxicity of FLUX-reacting T cells was concentration dependent and required concentrations in the range of peak serum levels after FLUX administration. Killing of target cells was mediated by different cytotoxic mechanisms. Our findings emphasize the role of the adaptive immune system and especially of activated drug-reacting T cells in human leukocyte antigen-associated, drug-induced liver injury.

  15. Cancer cells become susceptible to natural killer cell killing after exposure to histone deacetylase inhibitors due to glycogen synthase kinase-3-dependent expression of MHC class I-related chain A and B

    DEFF Research Database (Denmark)

    Skov, Søren; Pedersen, Marianne Terndrup; Andresen, Lars

    2005-01-01

    by small interfering RNA or by different inhibitors showed that GSK-3 activity is essential for the induced MICA/B expression. We thus present evidence that cancer cells which survive the direct induction of cell death by HDAC inhibitors become targets for NKG2D-expressing cells like NK cells, gammadelta T......We show that histone deacetylase (HDAC) inhibitors lead to functional expression of MHC class I-related chain A and B (MICA/B) on cancer cells, making them potent targets for natural killer (NK) cell-mediated killing through a NK group 2, member D (NKG2D) restricted mechanism. Blocking either...... apoptosis or oxidative stress caused by HDAC inhibitor treatment did not affect MICA/B expression, suggesting involvement of a separate signal pathway not directly coupled to induction of cell death. HDAC inhibitor treatment induced glycogen synthase kinase-3 (GSK-3) activity and down-regulation of GSK-3...

  16. Platelets kill intraerythrocytic malarial parasites and mediate survival to infection.

    Science.gov (United States)

    McMorran, Brendan J; Marshall, Vikki M; de Graaf, Carolyn; Drysdale, Karen E; Shabbar, Meriam; Smyth, Gordon K; Corbin, Jason E; Alexander, Warren S; Foote, Simon J

    2009-02-01

    Platelets play a critical role in the pathogenesis of malarial infections by encouraging the sequestration of infected red blood cells within the cerebral vasculature. But platelets also have well-established roles in innate protection against microbial infections. We found that purified human platelets killed Plasmodium falciparum parasites cultured in red blood cells. Inhibition of platelet function by aspirin and other platelet inhibitors abrogated the lethal effect human platelets exert on P. falciparum parasites. Likewise, platelet-deficient and aspirin-treated mice were more susceptible to death during erythrocytic infection with Plasmodium chabaudi. Both mouse and human platelets bind malarial-infected red cells and kill the parasite within. These results indicate a protective function for platelets in the early stages of erythrocytic infection distinct from their role in cerebral malaria.

  17. Are lead-free hunting rifle bullets as effective at killing wildlife as conventional lead bullets? A comparison based on wound size and morphology

    Energy Technology Data Exchange (ETDEWEB)

    Trinogga, Anna, E-mail: anna_trinogga@gmx.de; Fritsch, Guido; Hofer, Heribert; Krone, Oliver

    2013-01-15

    Fragmentation of the lead core of conventional wildlife hunting rifle bullets causes contamination of the target with lead. The community of scavenger species which feed on carcasses or viscera discarded by hunters are regularly exposed to these lead fragments and may die by acute or chronic lead intoxication, as demonstrated for numerous species such as white-tailed eagles (Haliaeetus albicilla) where it is among the most important sources of mortality. Not only does hunting with conventional ammunition deposit lead in considerable quantities in the environment, it also significantly delays or threatens the recovery of endangered raptor populations. Although lead-free bullets might be considered a suitable alternative that addresses the source of these problems, serious reservations have been expressed as to their ability to quickly and effectively kill a hunted animal. To assess the suitability of lead-free projectiles for hunting practice, the wounding potential of conventional bullets was compared with lead-free bullets under real life hunting conditions. Wound dimensions were regarded as good markers of the projectiles' killing potential. Wound channels in 34 killed wild ungulates were evaluated using computed tomography and post-mortem macroscopical examination. Wound diameters caused by conventional bullets did not differ significantly to those created by lead-free bullets. Similarly, the size of the maximum cross-sectional area of the wound was similar for both bullet types. Injury patterns suggested that all animals died by exsanguination. This study demonstrates that lead-free bullets are equal to conventional hunting bullets in terms of killing effectiveness and thus equally meet the welfare requirements of killing wildlife as painlessly as possible. The widespread introduction and use of lead-free bullets should be encouraged as it prevents environmental contamination with a seriously toxic pollutant and contributes to the conservation of a wide

  18. Photoexcited quantum dots for killing multidrug-resistant bacteria

    Science.gov (United States)

    Courtney, Colleen M.; Goodman, Samuel M.; McDaniel, Jessica A.; Madinger, Nancy E.; Chatterjee, Anushree; Nagpal, Prashant

    2016-05-01

    Multidrug-resistant bacterial infections are an ever-growing threat because of the shrinking arsenal of efficacious antibiotics. Metal nanoparticles can induce cell death, yet the toxicity effect is typically nonspecific. Here, we show that photoexcited quantum dots (QDs) can kill a wide range of multidrug-resistant bacterial clinical isolates, including methicillin-resistant Staphylococcus aureus, carbapenem-resistant Escherichia coli, and extended-spectrum β-lactamase-producing Klebsiella pneumoniae and Salmonella typhimurium. The killing effect is independent of material and controlled by the redox potentials of the photogenerated charge carriers, which selectively alter the cellular redox state. We also show that the QDs can be tailored to kill 92% of bacterial cells in a monoculture, and in a co-culture of E. coli and HEK 293T cells, while leaving the mammalian cells intact, or to increase bacterial proliferation. Photoexcited QDs could be used in the study of the effect of redox states on living systems, and lead to clinical phototherapy for the treatment of infections.

  19. Measurement of bacterial ingestion and killing by macrophages.

    Science.gov (United States)

    Campbell, P A; Canono, B P; Drevets, D A

    2001-05-01

    This unit presents fairly simple assays for measuring the binding of bacteria to macrophages, internalization of bacteria (also called ingestion or phagocytosis), and bacterial killing by macrophages. The first basic protocol describes how to measure the ability of macrophages to ingest bacteria. Because it is critical to remove residual extracellular organisms, the protocol presents two alternative steps to accomplish this: a washing procedure and a more stringent method in which cells are sedimented through sucrose. In addition, it is important to distinguish those bacteria truly ingested by a macrophage from those that are bound to, but not internalized by, the cell. A simple but effective way to do this is described in an alternate protocol. The unit also presents two ways to measure the ability of a macrophage to kill bacteria it has internalized. The first is a straightforward assay in which bacterial colonies are enumerated before and after a killing period; a subsequent colony count will indicate whether the bacteria grew within or were killed by the macrophage. The second protocol describes a way to measure bacterial viability based on bacterial metabolism, in which the ability of bacterial dehydrogenases to mediate the reduction of a tetrazolium salt to purple formazan is monitored by measuring absorbance spectrophotometrically.

  20. The response of normal and ataxia-telangiectasia cells to bleomycin: relationships between chromosome damage, cell cycle delay and cell killing.

    Science.gov (United States)

    Zampetti-Bosseler, F; Scott, D

    1985-08-01

    In agreement with our earlier observation (Scott and Zampetti-Bosseler, 1982) on X-irradiated normal and ataxia-telangiectasia (A-T) fibroblasts, we now report that after bleomycin or neocarzinostatin treatment also, A-T cells exhibit less G2 delay than normal cells. We confirm that A-T cells sustain more chromosome damage and lethality than normal cells after bleomycin. These observations support the hypothesis (Painter and Young, 1980) that A-T cells are defective in the recognition of certain lesions which normally lead to delays in progression through the cell cycle, during which they are repaired, and which, if unrepaired, lead to cell-lethal chromosome damage. However, we find that after bleomycin, as opposed to X-rays, the contribution of this type of lesion to cell death is minimal. The predominant lesions leading to cell death after bleomycin are not manifested at chromosome aberrations and do not lead to G2 delay or DNA-synthesis inhibition. A-T cells are defective in the recognition and/or repair of both types of lesion.

  1. Garlic compounds selectively kill childhood pre-B acute lymphoblastic leukemia cells in vitro without reducing T-cell function: Potential therapeutic use in the treatment of ALL

    Directory of Open Access Journals (Sweden)

    Greg Hodge

    2008-03-01

    Full Text Available Greg Hodge1, Stephen Davis2, Michael Rice1, Heather Tapp1, Ben Saxon1, Tamas Revesz11Haematology/Oncology Department, Women’s and Children’s Hospital, North Adelaide, Australia; 2Department of Mycology, Women’s and Children’s Hospital, North Adelaide, AustraliaAbstract: Drugs used for remission induction therapy for childhood precursor-B acute lymphoblastic leukemia (ALL are nonselective for malignant cells. Several garlic compounds have been shown to induce apoptosis of cancer cells and to alter lymphocyte function. To investigate the effect of garlic on the apoptosis of ALL cells and lymphocyte immune function, cells from newly diagnosed childhood ALL patients were cultured with several commonly used chemotherapeutic agents and several garlic compounds. Apoptosis, lymphocyte proliferation and T-cell cytokine production were determined using multiparameter flow cytometry. At concentrations of garlic compounds that did not result in significant increases in Annexin V and 7-AAD staining of normal lymphocytes, there was a significant increase in apoptosis of ALL cells with no alteration of T-cell proliferation as determined by CD25/CD69 upregulation or interferonγ, interleukin-2 or tumor necrosis factor-α intracellular cytokine production. In contrast, the presence of chemotherapeutic agents resulted in nonselective increases in both lymphocyte and ALL apoptosis and a decrease in T-cell proliferation and cytokine production. In conclusion, we show selective apoptosis of malignant cells by garlic compounds that do not alter T-cell immune function and indicate the potential therapeutic benefit of garlic compounds in the treatment of childhood ALL.Keywords: childhood precursor-B acute lymphoblastic leukemia, garlic, apoptosis, immune function, intracellular cytokines

  2. Specific targeted killing of ErbB2 positive breast cancer by retrovirus-mediated Immunocaspase-3 secreting T cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Lihong; YANG An-gang; JIA Lintao; ZHAO Jing; XU Yanming; WEN Weihong; BAO Wei; CAO Yunxin; SU Chengzhi; WANG Chengji

    2004-01-01

    In this study, Immunocaspase-3 gene was transfected into Jurkat T lymphocytes and the targeted proapoptotic protein Immunocaspase-3 was stably secreted.Its entry to ErbB2 positive SKBr3 breast carcinoma cell line was observed by indirect immunofluorescence staining.Growth of SKBr3 cells was significantly inhibited when they were cultured with medium containing Immunocaspase-3.Next, Immunocaspase-3 gene was cloned into retrovirus vector pLNCX, which was then transfected into PA317 cells to package. Packaged cells producing high titer pseudoviruses were acquired and the pseudoviruses were harvested to infect PBMCs, which had been stimulated to division. The latter were selected and administered to nude mice bearing SKBr3 tumors through tail vein. The results showed that the treatment contributed to an inhibition of tumor growth and prolonged the lifetime of nude mice bearing SKBr3 tumor.The efficiency of inhibition of tumor reached 73.25%, and the average lifetime of treated nude mice was 80.95% longer than that of control group. Immunohistochemical examination revealed the exclusive distribution of Immunocaspase-3proteins only in the tumor tissue samples; and TUNEL assay confirmed the occurrence of apoptosis in tumor cells. The present study suggests that Immunocaspase-3 secreted by T lymphocytes can selectively bind and enter into ErbB2 positive breast cancer cells, where it exhibits a proapoptotic activity and causes tumor suppression in an in vivo tumor model.

  3. Gadolinium-loaded chitosan nanoparticles for neutron-capture therapy: Influence of micrometric properties of the nanoparticles on tumor-killing effect.

    Science.gov (United States)

    Ichikawa, Hideki; Uneme, Takeshi; Andoh, Tooru; Arita, Yuya; Fujimoto, Takuya; Suzuki, Minoru; Sakurai, Yoshinori; Shinto, Hiroyuki; Fukasawa, Tomonori; Fujii, Fumihiko; Fukumori, Yoshinobu

    2014-06-01

    As a nanoparticulate device for controlled delivery of Gd in NCT, the authors have developed gadolinium-loaded chitosan nanoparticles (Gd-nanoCPs). In the present study, influence of micrometric properties such as particle size, particle-surface charge and Gd content of Gd-nanoCPs on tumor-killing effect by Gd-NCT was investigated with Gd-nanoCPs. Two types of Gd-nanoCPs with different mean particle size, zeta potential and Gd-content (Gd-nanoCP-400; 391nm, 28mV, 9wt% and Gd-nanoCP-200; 214nm, 19mV, 24wt%) could be prepared by using chitosans with different molecular weights. Gd-nanoCPs incorporating 1.2mg of natural Gd were injected intratumorally once or twice to mice subcutaneously-bearing B16F10 melanoma. Eight hours after the last administration, thermal neutron was irradiated to tumor region of the mice. Remarkable tumor-growth was observed in both hot and cold control groups. In contrast, Gd-NCT groups showed significant tumor-growth suppression effect, though their efficacy was found to depend on the micrometric properties of Gd-nanoCPs. In particular, the Gd-nanoCP-200 exhibited stronger tumor-killing effect than the Gd-nanoCP-400 at the same Gd dose and it was still similar to Gd-nanoCP-400 in tumor-growth suppressing effect even at the half of Gd dose of Gd-nanoCP-400. This significance in tumor-killing effect would be ascribed from a higher Gd retention in the tumor tissue and an improved distribution of Gd with intratumorally administered Gd-nanoCP-200. Indeed, the Gd concentration in tumor tissue at the time corresponding to the onset of thermal neutron irradiation was determined to be significantly higher in Gd-nanoCP-200, compared with Gd-nanoCP-400. These results demonstrated that appropriate modification of Gd-nanoCPs in micrometric properties would be an effective way to improve the retention of Gd in the tumor tissue after intratumoral injection, leading to the enhanced tumor-killing effect in Gd-NCT.

  4. Killing and replacing queen-laid eggs: low cost of worker policing in the honeybee.

    Science.gov (United States)

    Kärcher, Martin H; Ratnieks, Francis L W

    2014-07-01

    Worker honeybees, Apis mellifera, police each other's reproduction by killing worker-laid eggs. Previous experiments demonstrated that worker policing is effective, killing most (∼98%) worker-laid eggs. However, many queen-laid eggs were also killed (∼50%) suggesting that effective policing may have high costs. In these previous experiments, eggs were transferred using forceps into test cells, mostly into unrelated discriminator colonies. We measured both the survival of unmanipulated queen-laid eggs and the proportion of removal errors that were rectified by the queen laying a new egg. Across 2 days of the 3-day egg stage, only 9.6% of the queen-laid eggs in drone cells and 4.1% in worker cells were removed in error. When queen-laid eggs were removed from cells, 85% from drone cells and 61% from worker cells were replaced within 3 days. Worker policing in the honeybee has a high benefit to policing workers because workers are more related to the queen's sons (brothers, r = 0.25) than sister workers' sons (0.15). This study shows that worker policing also has a low cost in terms of the killing of queen-laid eggs, as only a small proportion of queen-laid eggs are killed, most of which are rapidly replaced.

  5. Silencing expression of the catalytic subunit of DNA-dependent protein kinase by small interfering RNA sensitizes human cells for radiation-induced chromosome damage, cell killing, and mutation

    Science.gov (United States)

    Peng, Yuanlin; Zhang, Qinming; Nagasawa, Hatsumi; Okayasu, Ryuichi; Liber, Howard L.; Bedford, Joel S.

    2002-01-01

    Targeted gene silencing in mammalian cells by RNA interference (RNAi) using small interfering RNAs (siRNAs) was recently described by Elbashir et al. (S. M. Elbashir et al., Nature (Lond.), 411: 494-498, 2001). We have used this methodology in several human cell strains to reduce expression of the Prkdc (DNA-PKcs) gene coding for the catalytic subunit of the DNA-dependent protein kinase (DNA-PKcs) that is involved in the nonhomologous end joining of DNA double-strand breaks. We have also demonstrated a radiosensitization for several phenotypic endpoints of radiation damage. In low-passage normal human fibroblasts, siRNA knock-down of DNA-PKcs resulted in a reduced capacity for restitution of radiation-induced interphase chromosome breaks as measured by premature chromosome condensation, an increased yield of acentric chromosome fragments at the first postirradiation mitosis, and an increased radiosensitivity for cell killing. For three strains of related human lymphoblasts, DNA-PKcs-targeted siRNA transfection resulted in little or no increase in radiosensitivity with respect to cell killing, a 1.5-fold decrease in induced mutant yield in TK6- and p53-null NH32 cells, but about a 2-fold increase in induced mutant yield in p53-mutant WTK1 cells at both the hypoxanthine quanine phosphoribosyl transferase (hprt) and the thymidine kinase loci.

  6. Carbofuran - A New and Effective Method of Illegal Killing of Otters (Lutra lutra in the Czech Republic

    Directory of Open Access Journals (Sweden)

    Václav Hlaváč

    2010-10-01

    Full Text Available Carbofuran - a poison recently used to illegally kill otters - is described from the Czech Republic. Six different cases of illegal poisoning of otters have been discovered since 2006 with a total number of 14 killed individuals. Apart from otters, many other animals - namely raptors and other carnivores - have been poisoned by carbofuran in the Czech Republic as well. The poisoning substance is now banned in the EU, however, it has been widely used in agriculture as insecticide and large supplies are generally available. This fact together with relative ease of using it can pose a threat to otter population, especially in areas with raising conflict between otter protection and fish farming. Therefore, it is important to report any suspicious cases of dead wildlife, to immediately carry out laboratory testing in such cases, and to inform relevant officials including the police. It will help monitor the problem and raise local awareness, and could possibly help to catch some persecutors. Taking these actions should be the first step in trying to stop poisoning wildlife by carbofuran.

  7. Effect of dietary heat-killed Lactobacillus brevis SBC8803 (SBL88™) on sleep: a non-randomised, double blind, placebo-controlled, and crossover pilot study.

    Science.gov (United States)

    Nakakita, Y; Tsuchimoto, N; Takata, Y; Nakamura, T

    2016-09-01

    We previously reported that dietary heat-killed Lactobacillus brevis SBC8803 affects sleep rhythms in mice. The present study evaluated the effect of consumption of heat-killed SBC8803 on sleep architecture in humans. A non-randomised, placebo-controlled, double blind, and crossover pilot study was conducted using volunteers who scored at a slightly high level (i.e. ≥6) on the Athens Insomnia Scale (AIS). Male subjects (n=17; age 41-69 y) consumed placebo or SBC8803 capsules (25 mg/day of heat-killed SBC8803) for 10 days. Electroencephalograms (EEG) were recorded using a mobile, one-channel system, providing objective data on sleep. Subjects' sleep journals and administration of the AIS provided subjective data on sleep. Three subjects were excluded from the statistical analysis. Analysis of the remaining 14 volunteers revealed no significant differences between placebo and SBC8803 consumption in either the AIS or the sleep EEG. The sleep journals revealed an improvement in 'waking' for the SBC8803 consumption periods (P=0.047), and there was a marginally significant effect on 'drowsiness during the following day' (P=0.067). Effects on the EEG delta power value (μV(2)/min) were revealed by a stratified analysis based on age, AIS, and the Beck Depression Inventory (BDI). Specifically, effects were found among subjects in their 40s who consumed the SBC8803 capsules (P=0.049) and among subjects with a BDI score less than the all-subjects average (13.3) (P=0.045). A marginally significant effect was found among subjects with an AIS score less than the all-subjects average (11.6) (P=0.065). The delta power value of 5 subjects with both BDI and AIS scores less than the average increased significantly (P=0.017). While the number of subjects was limited, a beneficial effect on sleep due to consumption of heat-killed L. brevis SBC8803 was found in subjects with slightly challenged sleep.

  8. Circulating preproinsulin signal peptide-specific CD8 T cells restricted by the susceptibility molecule HLA-A24 are expanded at onset of type 1 diabetes and kill β-cells.

    Science.gov (United States)

    Kronenberg, Deborah; Knight, Robin R; Estorninho, Megan; Ellis, Richard J; Kester, Michel G; de Ru, Arnoud; Eichmann, Martin; Huang, Guo C; Powrie, Jake; Dayan, Colin M; Skowera, Ania; van Veelen, Peter A; Peakman, Mark

    2012-07-01

    Type 1 diabetes results from T cell-mediated β-cell destruction. The HLA-A*24 class I gene confers significant risk of disease and early onset. We tested the hypothesis that HLA-A24 molecules on islet cells present preproinsulin (PPI) peptide epitopes to CD8 cytotoxic T cells (CTLs). Surrogate β-cell lines secreting proinsulin and expressing HLA-A24 were generated and their peptide ligandome examined by mass spectrometry to discover naturally processed and HLA-A24-presented PPI epitopes. A novel PPI epitope was identified and used to generate HLA-A24 tetramers and examine the frequency of PPI-specific T cells in new-onset HLA-A*24(+) patients and control subjects. We identified a novel naturally processed and HLA-A24-presented PPI signal peptide epitope (PPI(3-11); LWMRLLPLL). HLA-A24 tetramer analysis reveals a significant expansion of PPI(3-11)-specific CD8 T cells in the blood of HLA-A*24(+) recent-onset patients compared with HLA-matched control subjects. Moreover, a patient-derived PPI(3-11)-specific CD8 T-cell clone shows a proinflammatory phenotype and kills surrogate β-cells and human HLA-A*24(+) islet cells in vitro. These results indicate that the type 1 diabetes susceptibility molecule HLA-A24 presents a naturally processed PPI signal peptide epitope. PPI-specific, HLA-A24-restricted CD8 T cells are expanded in patients with recent-onset disease. Human islet cells process and present PPI(3-11), rendering themselves targets for CTL-mediated killing.

  9. Death ligand TRAIL, secreted by CD1a+ and CD14+ cells in blister fluids, is involved in killing keratinocytes in toxic epidermal necrolysis.

    Science.gov (United States)

    de Araujo, Elisabeth; Dessirier, Valérie; Laprée, Geneviève; Valeyrie-Allanore, Laurence; Ortonne, Nicolas; Stathopoulos, Efstathios N; Bagot, Martine; Bensussan, Armand; Mockenhaupt, Maja; Roujeau, Jean-Claude; Tsapis, Andreas

    2011-02-01

    Toxic epidermal necrolysis (TEN) is characterized by an acute detachment and destruction of keratinocytes, affecting large areas of the skin. It is often related to adverse drug reactions. Previous studies have shown that effector CD8+ T cells, which accumulate in the blister fluid, are functionally cytotoxic and act through a classical perforin/granzyme B pathway. It has recently been shown that these cytotoxic T cells also secrete granulysin peptide, which is lethal to keratinocytes. These cytotoxic T cells exert their killer activity against autologous keratinocytes in the presence of the drug. However, they are unlikely to be the only effectors of TEN. We therefore searched for soluble death factors in the blister fluids that might kill keratinocytes. We found that the amounts of interferon-γ, TRAIL and TNF-α proteins were significantly greater in TEN blister fluids than in all controls (normal sera, TEN sera, burns and Eosinophilic pustular folliculitis blister fluids) and TNF-like weak inducer of apoptosis (TWEAK) amounts are also greater in all controls except burns. We showed that these proteins acted in synergy to induce the death of keratinocytes in vitro. We also found that TRAIL and TWEAK were secreted by CD1a+ and CD14+ cells present in the blister fluids. Thus, in addition to MHC class I-restricted cytotoxic T lymphocytes (CTLs), which lyse keratinocytes, ligands secreted by non-lymphoid cells capable of inducing keratinocyte death in an MHC class I-independent manner, also seem to be present in the blister fluids of patients with TEN.

  10. CD8 T cell-mediated killing of orexinergic neurons induces a narcolepsy-like phenotype in mice.

    Science.gov (United States)

    Bernard-Valnet, Raphaël; Yshii, Lidia; Quériault, Clémence; Nguyen, Xuan-Hung; Arthaud, Sébastien; Rodrigues, Magda; Canivet, Astrid; Morel, Anne-Laure; Matthys, Arthur; Bauer, Jan; Pignolet, Béatrice; Dauvilliers, Yves; Peyron, Christelle; Liblau, Roland S

    2016-09-27

    Narcolepsy with cataplexy is a rare and severe sleep disorder caused by the destruction of orexinergic neurons in the lateral hypothalamus. The genetic and environmental factors associated with narcolepsy, together with serologic data, collectively point to an autoimmune origin. The current animal models of narcolepsy, based on either disruption of the orexinergic neurotransmission or neurons, do not allow study of the potential autoimmune etiology. Here, we sought to generate a mouse model that allows deciphering of the immune mechanisms leading to orexin(+) neuron loss and narcolepsy development. We generated mice expressing the hemagglutinin (HA) as a "neo-self-antigen" specifically in hypothalamic orexin(+) neurons (called Orex-HA), which were transferred with effector neo-self-antigen-specific T cells to assess whether an autoimmune process could be at play in narcolepsy. Given the tight association of narcolepsy with the human leukocyte antigen (HLA) HLA-DQB1*06:02 allele, we first tested the pathogenic contribution of CD4 Th1 cells. Although these T cells readily infiltrated the hypothalamus and triggered local inflammation, they did not elicit the loss of orexin(+) neurons or clinical manifestations of narcolepsy. In contrast, the transfer of cytotoxic CD8 T cells (CTLs) led to both T-cell infiltration and specific destruction of orexin(+) neurons. This phenotype was further aggravated upon repeated injections of CTLs. In situ, CTLs interacted directly with MHC class I-expressing orexin(+) neurons, resulting in cytolytic granule polarization toward neurons. Finally, drastic neuronal loss caused manifestations mimicking human narcolepsy, such as cataplexy and sleep attacks. This work demonstrates the potential role of CTLs as final effectors of the immunopathological process in narcolepsy.

  11. Diphtheria Toxin/Human B-Cell Activating Factor Fusion Protein Kills Human Acute Lymphoblastic Leukemia BALL-1 Cells: An Experimental Study

    Institute of Scientific and Technical Information of China (English)

    Xin-pu Gao; Zheng-min Liu; Yu-lian Jiao; Bin Cui; Yue-ting Zhu; Jie Zhang; Lai-cheng Wang; Yue-ran Zhao

    2012-01-01

    Objective:This study aimed to express a fusion protein of diphtheria toxin and human B ceil-activating factor (DT388sBAFF) in Escherichia coli (E.coli) and investigate its activity in human B-lineage acute lymphoblastic leukemia 1 cells (BALL-1).Methods:A fragment of DT388sBAFF fusion gene was separated from plasmid pUC57-DT388sBAFF digested with Nde Ⅰ and Xho Ⅰ,and inserted into the expression vector pcold Ⅱ digested with the same enzymes.Recombinants were screened by the colony polymerase chain reaction (PCR) and restriction map.The recombinant expression vector was transformed into BL21 and its expression was induced by isopropyl β-D-1-thiogalactopyranoside (IPTG).The recombinant protein was identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot,and then purified by Ni2+-NTA affinity chromatography.The expression level of B cell-activating factor receptor (BAFF-R) on BALL-1 cells was assessed by real-time PCR.The receptor binding capacity of recombinant protein was determined by cell fluorescent assay.The specific cytotoxicity of recombinant protein on BALL-1 cells was detected by 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) assay.Results:The expression level of recombinant protein was 50% of total bacterial proteins in E.coli,and the recombinant protein could bind to BAFF-R-positive BALL-1 cells and thereby produce a cytotoxic effect on the cells.Conclusion:The fusion protein expression vector DT388sBAFF was successfully constructed and the recombinant protein with selective cytotoxicity against BALL-1 cells was obtained,providing foundation for further study of the therapy of human B-lineage acute lymphoblastic leukemia.

  12. Sulforaphane enhances the anticancer activity of taxanes against triple negative breast cancer by killing cancer stem cells.

    Science.gov (United States)

    Burnett, Joseph P; Lim, Gi; Li, Yanyan; Shah, Ronak B; Lim, Rebekah; Paholak, Hayley J; McDermott, Sean P; Sun, Lichao; Tsume, Yasuhiro; Bai, Shuhua; Wicha, Max S; Sun, Duxin; Zhang, Tao

    2017-05-28

    Triple negative breast cancer (TNBC) typically exhibits rapid progression, high mortality and faster relapse rates relative to other breast cancer subtypes. In this report we examine the combination of taxanes (paclitaxel or docetaxel) with a breast cancer stem cell (CSC)-targeting agent sulforaphane for use against TNBC. We demonstrate that paclitaxel or docetaxel treatment induces IL-6 secretion and results in expansion of CSCs in TNBC cell lines. Conversely, sulforaphane is capable of preferentially eliminating CSCs, by inhibiting NF-κB p65 subunit translocation, downregulating p52 and consequent downstream transcriptional activity. Sulforaphane also reverses taxane-induced aldehyde dehydrogenase-positive (ALDH+) cell enrichment, and dramatically reduces the size and number of primary and secondary mammospheres formed. In vivo in an advanced treatment orthotopic mouse xenograft model together with extreme limiting dilution analysis (ELDA), the combination of docetaxel and sulforaphane exhibits a greater reduction in primary tumor volume and significantly reduces secondary tumor formation relative to either treatment alone. These results suggest that treatment of TNBCs with cytotoxic chemotherapy would be greatly benefited by the addition of sulforaphane to prevent expansion of and eliminate breast CSCs.

  13. Effects of raising frogs and putting pest-killing lamps in paddy fields on the prevention of rice pests and diseases

    Science.gov (United States)

    Teng, Qing; Hu, Xue-Feng; Luo, Fan; Cao, Ming-Yang

    2014-05-01

    Frogs in paddy fields become less and less due to applying large amounts of pesticides and human hunting for a long time, which causes the aggravation of rice pests and diseases. A field experiment was carried out in the suburb of Shanghai to study the effects of artificially raising frogs and putting frequency oscillation pest-killing lamps in paddy fields on the prevention of rice pests and diseases. The field experiment includes three treatments. Treatment I: 150 frogs, each 20 g in weight, per 100 m2 were put in the fields; Treatment II: a frequency oscillation pest-killing lamp was put in the fields; Treatment III: no frogs and pest-killing lamps were put in the fields. All the experimental fields were operated based on the organic faming system. The amount of organic manure, 7500 kg/hm2, was applied to the fields as base fertilizer before sowing in early June, 2013. No any chemical fertilizers and pesticides were used during the entire period of rice growth. Each treatment is in triplicate and each plot is 67 m2 in area. The results are as follows: (1) During the entire growth period, the incidences of rice pests and diseases with Treatment I and II are significantly lower than those with CK (Treatment III). The incidence of chilo suppressalis with Treatment I, II and III is 0, 0.46% and 1.69%, respectively; that of cnaphalocrocis medinalis is 7.67%, 6.62% and 10.10%, respectively; that of rice sheath blight is 0, 11.11% and 5.43%, respectively; that of rice planthopper is 4.25 per hill, 5.75 per hill and 11 per hill, respectively. (2) The grain yield of the three treatments is significantly different. That of Treatment I, II and III is 5157.73 kg/hm2, 4761.60 kg/hm2 and 3645.14kg/hm2 on average, respectively. (3) Affected by frog activities, the contents of NH4-N, available P and available K in the soil with Treatment I are significantly raised. All the above suggest that artificially raising frogs in paddy fields could effectively prevent rice pests and

  14. Fc-mediated activity of EGFR x c-Met bispecific antibody JNJ-61186372 enhanced killing of lung cancer cells.

    Science.gov (United States)

    Grugan, Katharine D; Dorn, Keri; Jarantow, Stephen W; Bushey, Barbara S; Pardinas, Jose R; Laquerre, Sylvie; Moores, Sheri L; Chiu, Mark L

    2017-01-01

    Epidermal growth factor receptor (EGFR) mutant non-small cell lung cancers acquire resistance to EGFR tyrosine kinase inhibitors through multiple mechanisms including c-Met receptor pathway activation. We generated a bispecific antibody targeting EGFR and c-Met (JNJ-61186372) demonstrating anti-tumor activity in wild-type and mutant EGFR settings with c-Met pathway activation. JNJ-61186372 was engineered with low fucosylation (<10 %), resulting in enhanced antibody-dependent cell-mediated cytotoxicity and FcγRIIIa binding. In vitro and in vivo studies with the single-arm EGFR or c-Met versions of JNJ-61186372 identified that the Fc-activity of JNJ-61186372 is mediated by binding of the anti-EGFR arm and required for inhibition of EGFR-driven tumor cells. In a tumor model driven by both EGFR and c-Met, treatment with Fc-silent JNJ-61186372 or with c-Met single-arm antibody reduced tumor growth inhibition compared to treatment with JNJ-61186372, suggesting that the Fc function of JNJ-61186372 is essential for maximal tumor inhibition. Moreover in this same model, downregulation of both EGFR and c-Met receptors was observed upon treatment with Fc-competent JNJ-61186372, suggesting that the Fc interactions are necessary for down-modulation of the receptors in vivo and for efficacy. These Fc-mediated activities, in combination with inhibition of both the EGFR and c-Met signaling pathways, highlight the multiple mechanisms by which JNJ-61186372 combats therapeutic resistance in EGFR mutant patients.

  15. Inhibition of HSP90 by AUY922 Preferentially Kills Mutant KRAS Colon Cancer Cells by Activating Bim through ER Stress.

    Science.gov (United States)

    Wang, Chun Yan; Guo, Su Tang; Wang, Jia Yu; Liu, Fen; Zhang, Yuan Yuan; Yari, Hamed; Yan, Xu Guang; Jin, Lei; Zhang, Xu Dong; Jiang, Chen Chen

    2016-03-01

    Oncogenic mutations of KRAS pose a great challenge in the treatment of colorectal cancer. Here we report that mutant KRAS colon cancer cells are nevertheless more susceptible to apoptosis induced by the HSP90 inhibitor AUY922 than those carrying wild-type KRAS. Although AUY922 inhibited HSP90 activity with comparable potency in colon cancer cells irrespective of their KRAS mutational statuses, those with mutant KRAS were markedly more sensitive to AUY922-induced apoptosis. This was associated with upregulation of the BH3-only proteins Bim, Bik, and PUMA. However, only Bim appeared essential, in that knockdown of Bim abolished, whereas knockdown of Bik or PUMA only moderately attenuated apoptosis induced by AUY922. Mechanistic investigations revealed that endoplasmic reticulum (ER) stress was responsible for AUY922-induced upregulation of Bim, which was inhibited by a chemical chaperone or overexpression of GRP78. Conversely, siRNA knockdown of GRP78 or XBP-1 enhanced AUY922-induced apoptosis. Remarkably, AUY922 inhibited the growth of mutant KRAS colon cancer xenografts through activation of Bim that was similarly associated with ER stress. Taken together, these results suggest that AUY922 is a promising drug in the treatment of mutant KRAS colon cancers, and the agents that enhance the apoptosis-inducing potential of Bim may be useful to improve the therapeutic efficacy.

  16. Inefficacy of vancomycin and teicoplanin in eradicating and killing Staphylococcus epidermidis biofilms in vitro.

    Science.gov (United States)

    Claessens, J; Roriz, M; Merckx, R; Baatsen, P; Van Mellaert, L; Van Eldere, J

    2015-04-01

    Biofilm-associated bacteria display a decreased susceptibility towards antibiotics. Routine assessment of antibiotic susceptibility of planktonic bacteria therefore offers an insufficient prediction of the biofilm response. In this study, in vitro biofilms of eight clinical Staphylococcus epidermidis strains were subjected to treatment with vancomycin, teicoplanin, oxacillin, rifampicin and gentamicin. In addition, the biofilms were subjected to combinations of an antibiotic with rifampicin. The effects on the biofilms were assessed by crystal violet staining to determine the total biofilm biomass, staining with XTT to determine bacterial cell viability, and microscopy. Combining these methods showed that treatment of S. epidermidis biofilms with glycopeptides increased the total biofilm biomass and that these antibiotics were not effective in killing bacteria embedded in biofilms. The decreased killing efficacy was more pronounced in biofilms produced by strains that were classified as 'strong' biofilm producers. Rifampicin, oxacillin and gentamicin effectively killed biofilm-associated bacteria of all tested strains. Combining antibiotics with rifampicin increased the killing efficacy without influencing the total biofilm biomass. When vancomycin or teicoplanin were combined with rifampicin, the increase in biofilm biomass was neutralised and also the killing efficacy was influenced in a positive way. We conclude that the combined methodology used in this study showed that glycopeptides were not effective in eradicating S. epidermidis biofilms but that combination with rifampicin improved the killing efficacy in vitro.

  17. Comparison microbial killing efficacy between sonodynamic therapy and photodynamic therapy

    Science.gov (United States)

    Drantantiyas, Nike Dwi Grevika; Astuti, Suryani Dyah; Nasution, Aulia M. T.

    2016-11-01

    Biofilm is a way used by bacteria to survive from their environmental conditions by forming colony of bacteria. Specific characteristic in biofilm formation is the availability of matrix layer, known as extracellular polymer substance. Treatment using antibiotics may lead bacteria to be to resistant. Other treatments to reduce microbial, like biofilm, can be performed by using photodynamic therapy. Successful of this kind of therapy is induced by penetration of light and photosensitizer into target cells. The sonodynamic therapy offers greater penetrating capability into tissues. This research aimed to use sonodynamic therapy in reducing biofilm. Moreover, it compares also the killing efficacy of photodynamic therapy, sonodynamic therapy, and the combination of both therapeutic schemes (known as sono-photodynamic) to achieve higher microbial killing efficacy. Samples used are Staphylococcus aureus biofilm. Treatments were divided into 4 groups, i.e. group under ultrasound treatment with variation of 5 power levels, group of light treatment with exposure of 75s, group of combined ultrasound-light with variation of ultrasound power levels, and group of combined lightultrasound with variation of ultrasound power levels. Results obtained for each treatment, expressed in % efficacy of log CFU/mL, showed that the treatment of photo-sonodynamic provides greater killing efficacy in comparison to either sonodynamic and sono-photodynamic. The photo-sonodynamic shows also greater efficacy to photodynamic. So combination of light-ultrasound (photo-sonodynamic) can effectively kill microbial biofilm. The combined therapy will provide even better efficacy using exogenous photosensitizer.

  18. Killing rates for caspofungin against Candida albicans after brief and continuous caspofungin exposure in the presence and absence of serum.

    Science.gov (United States)

    Kovács, Renátó; Gesztelyi, Rudolf; Perlin, David S; Kardos, Gábor; Domán, Marianna; Berényi, Réka; Majoros, László

    2014-10-01

    It was previously demonstrated that brief (≤1 h) exposures to echinocandins are as effective to kill Candida albicans cells as continuous 24-h exposure. However, killing rates after continuous and short (1 h) echinocandin exposures to C. albicans have not yet been evaluated in RPMI-1640 with and without 50 % serum. We evaluated four echinocandin susceptible C. albicans bloodstream isolates, ATCC 10231 type strain and an echinocandin-resistant isolate (DPL20, FKS F645P). Caspofungin MICs, time-kill and postantifungal effect (PAFE) tests were performed in RPMI-1640 with and without 50 % serum. Killing rates (k values) in time-kill and PAFE experiments were determined for each strain and concentration. In time-kill experiments, colony count decreases were isolate- and concentration-dependent at 0.25, 1, 4, 8, 16 and 32 mg/L in RPMI-1640, but concentration-independent at 1, 4, 8, 16 and 32 mg/L in 50 % serum. One-hour caspofungin exposure at 4, 16 and 32 mg/L resulted in CFU decreases comparable with the results obtained in time-kill experiments in RPMI-1640, but 50 % serum at 4, 16 and 32 mg/L allowed growth of all isolates (k values were negative) (P caspofungin exposure. As only a short growth inhibition without killing was observed in 50 % serum, clinical relevance of caspofungin PAFE in vivo is questionable.

  19. Three-dimensional (3D) plasmonic hot spots for label-free sensing and effective photothermal killing of multiple drug resistant superbugs.

    Science.gov (United States)

    Jones, Stacy; Sinha, Sudarson Sekhar; Pramanik, Avijit; Ray, Paresh Chandra

    2016-11-03

    Drug resistant superbug infection is one of the foremost threats to human health. Plasmonic nanoparticles can be used for ultrasensitive bio-imaging and photothermal killing by amplification of electromagnetic fields at nanoscale "hot spots". One of the main challenges to plasmonic imaging and photothermal killing is design of a plasmonic substrate with a large number of "hot spots". Driven by this need, this article reports design of a three-dimensional (3D) plasmonic "hot spot"-based substrate using gold nanoparticle attached hybrid graphene oxide (GO), free from the traditional 2D limitations. Experimental results show that the 3D substrate has capability for highly sensitive label-free sensing and generates high photothermal heat. Reported data using p-aminothiophenol conjugated 3D substrate show that the surface enhanced Raman spectroscopy (SERS) enhancement factor for the 3D "hot spot"-based substrate is more than two orders of magnitude greater than that for the two-dimensional (2D) substrate and five orders of magnitude greater than that for the zero-dimensional (0D) p-aminothiophenol conjugated gold nanoparticle. 3D-Finite-Difference Time-Domain (3D-FDTD) simulation calculations indicate that the SERS enhancement factor can be greater than 10(4) because of the bent assembly structure in the 3D substrate. Results demonstrate that the 3D-substrate-based SERS can be used for fingerprint identification of several multi-drug resistant superbugs with detection limits of 5 colony forming units per mL. Experimental data show that 785 nm near infrared (NIR) light generates around two times more photothermal heat for the 3D substrate with respect to the 2D substrate, and allows rapid and effective killing of 100% of the multi-drug resistant superbugs within 5 minutes.

  20. It's not just conflict that motivates killing of orangutans.

    Directory of Open Access Journals (Sweden)

    Jacqueline T Davis

    Full Text Available We investigated why orangutans are being killed in Kalimantan, Indonesia, and the role of conflict in these killings. Based on an analysis of interview data from over 5,000 respondents in over 450 villages, we also assessed the socio-ecological factors associated with conflict and non-conflict killings. Most respondents never kill orangutans. Those who reported having personally killed an orangutan primarily did so for non-conflict reasons; for example, 56% of these respondents said that the reason they had killed an orangutan was to eat it. Of the conflict-related reasons for killing, the most common reasons orangutans were killed was fear of orangutans or in self-defence. A similar pattern was evident among reports of orangutan killing by other people in the villages. Regression analyses indicated that religion and the percentage of intact forest around villages were the strongest socio-ecological predictors of whether orangutans were killed for conflict or non-conflict related reasons. Our data indicate that between 44,170 and 66,570 orangutans were killed in Kalimantan within the respondents' active hunting lifetimes: between 12,690 and 29,024 for conflict reasons (95%CI and between 26,361 and 41,688 for non-conflict reasons (95% CI. These findings confirm that habitat protection alone will not ensure the survival of orangutans in Indonesian Borneo, and that effective reduction of orangutan killings is urgently needed.

  1. Extracts from Calendula officinalis offer in vitro protection against H2 O2 induced oxidative stress cell killing of human skin cells.

    Science.gov (United States)

    Alnuqaydan, Abdullah M; Lenehan, Claire E; Hughes, Rachel R; Sanderson, Barbara J

    2015-01-01

    The in vitro safety and antioxidant potential of Calendula officinalis flower head extracts was investigated. The effect of different concentrations (0.125, 0.5, 1.0, 2.0 and 5.0% (v/v)) of Calendula extracts on human skin cells HaCaT in vitro was explored. Doses of 1.0% (v/v) (0.88 mg dry weight/mL) or less showed no toxicity. Cells were also exposed to the Calendula extracts for either 4, 24 or 48 h before being exposed to an oxidative insult (hydrogen peroxide H2 O2 ) for 1 h. Using the MTT cytotoxicity assay, it was observed that two independent extracts of C. officinalis gave time-dependent and concentration-dependent H2 O2 protection against induced oxidative stress in vitro using human skin cells. Pre-incubation with the Calendula extracts for 24 and 48 h increased survival relative to the population without extract by 20% and 40% respectively following oxidative challenge. The antioxidant potential of the Calendula extracts was confirmed using a complimentary chemical technique, the DPPH(●) assay. Calendula extracts exhibited free radical scavenging abilities. This study demonstrates that Calendula flower extracts contain bioactive and free radical scavenging compounds that significantly protect against oxidative stress in a human skin cell culture model.

  2. Effect of a killed whole Neospora caninum tachyzoite vaccine on the crude abortion rate of Costa Rican dairy cows under field conditions.

    Science.gov (United States)

    Romero, J J; Pérez, E; Frankena, K

    2004-09-02

    A standard field trial was carried out to assess the effect of a commercial Neospora-vaccine based on whole killed tachyzoites (Bovilis-Neoguard, Intervet) on the abortion rate. Eight hundred and seventy-six cows, over 2.5 months pregnant, belonging to 25 Costa Rican dairy herds, were used in the analysis. For each cow vaccinated, a cow of the same herd, breed and age category, was selected as control. The period of administration of treatments extended from June to November of 2000. The treatments were administered in two, 5-ml doses 1 month apart, the first dose given between day 75 and 90 of gestation. The incidence of abortion among all treated cows was of 16.0% (140/876). The treatment specific incidence was 11.2% (49/438) and 20.8% (91/438) for the vaccinated and the placebo group, respectively. The prevented fraction by vaccination amounted to 0.46 (95% CI: 0.26, 0.61), and the cumulative incidence ratio for the vaccinated group was 0.54. The Cox hazard ratio was 0.51(95% CI: 0.37, 0.72), meaning that the force of abortion is reduced twice in the vaccinated group. The results of this study, the first one following this type of design, shows that the killed whole Neospora caninum tachyzoite preparation had a reasonable effect on the abortion rate in Costa Rican dairy cattle.

  3. Selective killing of K-ras-transformed pancreatic cancer cells by targeting NAD(P)H oxidase

    Institute of Scientific and Technical Information of China (English)

    Peng Wang; Yi-Chen Sun; Wen-Hua Lu; Peng Huang; and Yumin Hu

    2015-01-01

    Introduction:Oncogenic activation of the K-ras gene occurs in>90%of pancreatic ductal carcinoma and plays a critical role in the pathogenesis of this malignancy. Increase of reactive oxygen species (ROS) has also been observed in a wide spectrum of cancers. This study aimed to investigate the mechanistic association between K-ras–induced transformation and increased ROS stress and its therapeutic implications in pancreatic cancer. Methods:ROS level, NADPH oxidase (NOX) activity and expression, and cel invasion were examined in human pancreatic duct epithelial E6E7 cel s transfected with K-rasG12V compared with parental E6E7 cel s. The cytotoxic effect and antitumor effect of capsaicin, a NOX inhibitor, were also tested in vitro and in vivo. Results:K-ras transfection caused activation of the membrane-associated redox enzyme NOX and elevated ROS generation through the phosphatidylinositol 3′-kinase (PI3K) pathway. Importantly, capsaicin preferential y inhibited the enzyme activity of NOX and induced severe ROS accumulation in K-ras–transformed cel s compared with parental E6E7 cel s. Furthermore, capsaicin effectively inhibited cel proliferation, prevented invasiveness of K-ras–transformed pancreatic cancer cel s, and caused minimum toxicity to parental E6E7 cel s. In vivo, capsaicin exhibited antitumor activity against pancreatic cancer and showed oxidative damage to the xenograft tumor cel s. Conclusions:K-ras oncogenic signaling causes increased ROS stress through NOX, and abnormal ROS stress can selectively kil tumor cel s by using NOX inhibitors. Our study provides a basis for developing a novel therapeutic strategy to effectively kil K-ras–transformed cel s through a redox-mediated mechanism.

  4. An actinomycete isolate from solitary wasp mud nest having strong antibacterial activity and kills the Candida cells due the shrinkage and the cytosolic loss

    Directory of Open Access Journals (Sweden)

    Vijay eKumar

    2014-08-01

    Full Text Available An actinomycetes strain designated as MN 2(6 was isolated from the solitary wasp mud nest. The isolate was identified using polyphasic taxonomy. It produced the extensive branched brown substrate and white aerial hyphae that changed into grayish black. The aerial mycelia produced the spiral spore chains with rugose spore surface. The growth was observed between temperature range of 27-37°C, pH 8-10 and below salt concentration of 6% (w/v. The comparative analysis of 16S rRNA gene sequence and phylogenetic relationship showed that strain MN 2(6 lies in clade with Streptomyces hygroscopicus subsp. hygroscopicus NRRL 2387T, Streptomyces sporocinereus NBRC 100766T and Streptomyces demainii NRRL B-1478T with which it shares a 16S rRNA gene sequence similarity of 99.3%. The strain MN 2(6 can be differentiated from type strains based on phenotypic characteristics. The strain MN 2(6 showed most promising activity against Gram-positive, Gram-negative bacteria, acid-fast bacilli and Candida species suggesting broad-spectrum characteristics of the active metabolite. Evaluation of anti-candidal activity of the metabolite of strain MN 2(6 by scanning electron microscopy (SEM revealed changed external morphology of yeast. It kills the Candida cells due to the shrinkage and the cytosolic loss. However, further studies are required to elucidate the structure of the active metabolite produced by the isolate MN 2(6

  5. Evaluation of the effects of a plasma activated medium on cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Mohades, S.; Laroussi, M., E-mail: mlarouss@odu.edu; Sears, J.; Barekzi, N.; Razavi, H. [Plasma Engineering and Medicine Institute, Old Dominion University, Norfolk, Virginia 23529 (United States)

    2015-12-15

    The interaction of low temperature plasma with liquids is a relevant topic of study to the field of plasma medicine. This is because cells and tissues are normally surrounded or covered by biological fluids. Therefore, the chemistry induced by the plasma in the aqueous state becomes crucial and usually dictates the biological outcomes. This process became even more important after the discovery that plasma activated media can be useful in killing various cancer cell lines. Here, we report on the measurements of concentrations of hydrogen peroxide, a species known to have strong biological effects, produced by application of plasma to a minimum essential culture medium. The activated medium is then used to treat SCaBER cancer cells. Results indicate that the plasma activated medium can kill the cancer cells in a dose dependent manner, retain its killing effect for several hours, and is as effective as apoptosis inducing drugs.

  6. Evaluation of the effects of a plasma activated medium on cancer cells

    Science.gov (United States)

    Mohades, S.; Laroussi, M.; Sears, J.; Barekzi, N.; Razavi, H.

    2015-12-01

    The interaction of low temperature plasma with liquids is a relevant topic of study to the field of plasma medicine. This is because cells and tissues are normally surrounded or covered by biological fluids. Therefore, the chemistry induced by the plasma in the aqueous state becomes crucial and usually dictates the biological outcomes. This process became even more important after the discovery that plasma activated media can be useful in killing various cancer cell lines. Here, we report on the measurements of concentrations of hydrogen peroxide, a species known to have strong biological effects, produced by application of plasma to a minimum essential culture medium. The activated medium is then used to treat SCaBER cancer cells. Results indicate that the plasma activated medium can kill the cancer cells in a dose dependent manner, retain its killing effect for several hours, and is as effective as apoptosis inducing drugs.

  7. Inhibition of pre-B cell colony-enhancing factor (PBEF/NAMPT/visfatin) decreases the ability of human neutrophils to generate reactive oxidants but does not impair bacterial killing.

    Science.gov (United States)

    Roberts, Kate J; Cross, Andrew; Vasieva, Olga; Moots, Robert J; Edwards, Steven W

    2013-09-01

    NAMPT, also known as PBEF and visfatin, can act extracellularly as a cytokine-like molecule or intracellularly as a NAMPT, regulating NAD biosynthesis in the NAD salvage pathway. Inhibitors of NAMPT have anti-inflammatory and anticancer activity and are finding use as therapeutic agents. In view of the importance of NAD metabolism in neutrophil function, we determined the effects of NAMPT inhibition on a variety of neutrophil functions associated with their role in host protection against infections. Incubation of human neutrophils with the NAMPT inhibitor APO866 decreased neutrophil NAD(P)/H levels in a dose- and time-dependent manner but without a concomitant change in cell viability. NAMPT inhibition did not affect the expression of a number of cell-surface receptors involved in adhesion and opsono-phagocytosis, but the respiratory burst was decreased significantly. Whereas opsono-phagocytosis of Staphylococcus aureus was unaffected by NAMPT inhibition, intraphagosomal oxidant production was decreased. However, the killing efficiency of neutrophils was unaffected. These data indicate that therapeutic NAMPT inhibition is unlikely to have deleterious effects on host protection against infections, in spite of this ability to down-regulate neutrophil respiratory burst activity significantly.

  8. Theriocide: Naming Animal Killing

    Directory of Open Access Journals (Sweden)

    Piers Beirne

    2014-08-01

    Full Text Available In this essay I recommend ‘theriocide’ as the name for those diverse human actions that cause the deaths of animals. Like the killing of one human by another, theriocide may be socially acceptable or unacceptable, legal or illegal. It may be intentional or unintentional and may involve active maltreatment or passive neglect. Theriocide may occur one-on-one, in small groups or in large-scale social institutions. The numerous and sometimes intersecting sites of theriocide include intensive rearing regimes; hunting and fishing; trafficking; vivisection; militarism; pollution; and human-induced climate change. If the killing of animals by humans is as harmful to them as homicide is to humans, then the proper naming of such deaths offers a remedy, however small, to the extensive privileging of human lives over those of other animals. Inevitably, the essay leads to a shocking question: Is theriocide murder?

  9. Killing of Klebsiella pneumoniae by human alveolar macrophages.

    Science.gov (United States)

    Hickman-Davis, Judy M; O'Reilly, Philip; Davis, Ian C; Peti-Peterdi, Janos; Davis, Glenda; Young, K Randall; Devlin, Robert B; Matalon, Sadis

    2002-05-01

    We investigated putative mechanisms by which human surfactant protein A (SP-A) effects killing of Klebsiella pneumoniae by human alveolar macrophages (AMs) isolated from bronchoalveolar lavagates of patients with transplanted lungs. Coincubation of AMs with human SP-A (25 microg/ml) and Klebsiella resulted in a 68% decrease in total colony forming units by 120 min compared with AMs infected with Klebsiella in the absence of SP-A, and this SP-A-mediated effect was abolished by preincubation with N(G)-monomethyl-L-arginine. Incubation of transplant AMs with SP-A increased intracellular Ca(2+) concentration ([Ca(2+)](i)) by 70% and nitrite and nitrate (NO(x)) production by 45% (from 0.24 +/- 0.02 to 1.3 +/- 0.21 nmol small middle dot 10(6) AMs(-1).h(-1)). Preincubation with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester inhibited the increase in [Ca(2+)](i) and abrogated the SP-A-mediated Klebsiella phagocytosis and killing. In contrast, incubation of AMs from normal volunteers with SP-A decreased both [Ca(2+)](i) and NO(x) production and did not result in killing of Klebsiella. Significant killing of Klebsiella was also seen in a cell-free system by sustained production of peroxynitrite (>1 microM/min) at pH 5 but not at pH 7.4. These findings indicate that SP-A mediates pathogen killing by AMs from transplant lungs by stimulating phagocytosis and production of reactive oxygen-nitrogen intermediates.

  10. Increased Lytic Efficiency of Bovine Macrophages Trained with Killed Mycobacteria

    Science.gov (United States)

    Juste, Ramon A.; Alonso-Hearn, Marta; Garrido, Joseba M.; Abendaño, Naiara; Sevilla, Iker A.; Gortazar, Christian; de la Fuente, José; Dominguez, Lucas

    2016-01-01

    Innate immunity is evolutionarily conserved in multicellular organisms and was considered to lack memory until very recently. One of its more characteristic mechanisms is phagocytosis, the ability of cells to engulf, process and eventually destroy any injuring agent. We report the results of an ex vivo experiment in bovine macrophages in which improved clearance of Mycobacterium bovis (M. bovis) was induced by pre-exposure to a heat killed M. bovis preparation. The effects were independent of humoral and cellular adaptive immune responses and lasted up to six months. Specifically, our results demonstrate the existence of a training effect in the lytic phase of phagocytosis that can be activated by killed mycobacteria, thus suggesting a new mechanism of vaccine protection. These findings are compatible with the recently proposed concept of trained immunity, which was developed to explain the observation that innate immune responses provide unspecific protection against pathogens including other than those that originally triggered the immune response. PMID:27820836

  11. Killing of naive T cells by CD95L-transfected dendritic cells (DC): in vivo study using killer DC-DC hybrids and CD4(+) T cells from DO11.10 mice.

    Science.gov (United States)

    Kusuhara, Masahiro; Matsue, Keiko; Edelbaum, Dale; Loftus, Julie; Takashima, Akira; Matsue, Hiroyuki

    2002-04-01

    Dendritic cells (DC) play the dual task of initiating cellular immunity against potentially harmful foreign antigens (Ag), while maintaining immunological tolerance to self-Ag and environmental Ag. As an approach to induce Ag-specific suppression, we and others introduced CD95 ligand (L) cDNA into DC. The resulting "killer" DC delivered apoptotic signals, instead of activation signals, to primed CD4(+) T cells in vitro and induced Ag-specific immunosuppression in vivo. To study the impact of killer DC on naive T cells, the fate of Ag-reactive T cells and the extent of their depletion after killer DC treatment, we performed in vitro and in vivo reconstitution experiments using: (a) killer DC-DC hybrids created between CD95L-transduced XS106 DC clone (A/J origin) and splenic DC from BALB/c mice, (b) CD4(+) T cells isolated from DO11.10 transgenic mice (BALB/c background), and (c) OVA(323-339) peptide as relevant Ag. Ovalbumin (OVA)-pulsed killer DC-DC hybrids inhibited DO11.10 T cell activation triggered by conventional DC, instead of inducing their activation. Rapid apoptosis of T cells was observed after co-culture with OVA-pulsed killer DC-DC hybrids, but not with non-pulsed killer DC-DC hybrids or OVA-pulsed control DC-DC hybrids. For in vivo reconstitution, (BALB/cxA/J)F1 mice received subcutaneous administration of killer DC-DC hybrids, followed by intravenous inoculation of DO11.10 T cells. Killer DC-DC hybrids migrated preferentially to draining lymph nodes albeit with relatively low efficiency (0.5-1% recovery) and they induced significant, but incomplete (30-40%) killing of DO11.10 T cells in this location. These results document the abilities of CD95L-transduced DC to trigger apoptosis of naive T cells in an Ag-specific manner, to overrule T cell activation signals delivered by conventional DC, and to reduce local frequencies of Ag-reactive T cells in vivo. Our data also uncover two major limitations (relatively low homing efficiency and incomplete

  12. Electroencephalographic evaluation of the effectiveness of blunt trauma to induce loss of consciousness for on-farm killing of chickens and turkeys.

    Science.gov (United States)

    Cors, J-C; Gruber, A D; Günther, R; Meyer-Kühling, B; Esser, K-H; Rautenschlein, S

    2015-02-01

    Euthanasia of small numbers of birds in case of injury or other illness directly on the farm may be necessary for welfare reasons. This should be done without transportation of the moribund animals in order to minimize pain and distress. Blood loss has to be avoided to minimize the risk of contaminating the environment. Cervical dislocation in combination with a blunt trauma may be an appropriate way to achieve this aim but the bird's age and body weight may influence the practicability of this method in the field. In this study, we evaluated broilers, broiler breeders, and turkeys of different age groups and weights up to nearly 16 kg for the efficacy of blunt trauma to induce unconsciousness, allowing subsequent killing of the bird without pain. The effect of blunt trauma on the brain was determined by electroencephalography (EEG). Auditory evoked potentials (AEPs) were recorded for each animal. Convulsions or tonic seizures were observed in all investigated animals after blunt trauma, including strong wing movements, torticollis, and stretching of legs. The EEG results demonstrate that the blunt trauma induced by a single, sufficiently strong hit placed in the frontoparietal region of the head led to a reduction or loss of the AEP in all groups of birds. These results clearly indicate a loss of sensibility and induction of unconsciousness, which would allow painless killing of the birds immediately after the induction of the blunt trauma.

  13. Burkholderia pseudomallei Capsule Exacerbates Respiratory Melioidosis but Does Not Afford Protection against Antimicrobial Signaling or Bacterial Killing in Human Olfactory Ensheathing Cells.

    Science.gov (United States)

    Dando, Samantha J; Ipe, Deepak S; Batzloff, Michael; Sullivan, Matthew J; Crossman, David K; Crowley, Michael; Strong, Emily; Kyan, Stephanie; Leclercq, Sophie Y; Ekberg, Jenny A K; St John, James; Beacham, Ifor R; Ulett, Glen C

    2016-07-01

    Melioidosis, caused by the bacterium Burkholderia pseudomallei, is an often severe infection that regularly involves respiratory disease following inhalation exposure. Intranasal (i.n.) inoculation of mice represents an experimental approach used to study the contributions of bacterial capsular polysaccharide I (CPS I) to virulence during acute disease. We used aerosol delivery of B. pseudomallei to establish respiratory infection in mice and studied CPS I in the context of innate immune responses. CPS I improved B. pseudomallei survival in vivo and triggered multiple cytokine responses, neutrophil infiltration, and acute inflammatory histopathology in the spleen, liver, nasal-associated lymphoid tissue, and olfactory mucosa (OM). To further explore the role of the OM response to B. pseudomallei infection, we infected human olfactory ensheathing cells (OECs) in vitro and measured bacterial invasion and the cytokine responses induced following infection. Human OECs killed >90% of the B. pseudomallei in a CPS I-independent manner and exhibited an antibacterial cytokine response comprising granulocyte colony-stimulating factor, tumor necrosis factor alpha, and several regulatory cytokines. In-depth genome-wide transcriptomic profiling of the OEC response by RNA-Seq revealed a network of signaling pathways activated in OECs following infection involving a novel group of 378 genes that encode biological pathways controlling cellular movement, inflammation, immunological disease, and molecular transport. This represents the first antimicrobial program to be described in human OECs and establishes the extensive transcriptional defense network accessible in these cells. Collectively, these findings show a role for CPS I in B. pseudomallei survival in vivo following inhalation infection and the antibacterial signaling network that exists in human OM and OECs.

  14. Human alpha-lactalbumin made lethal to tumor cells (HAMLET) kills human glioblastoma cells in brain xenografts by an apoptosis-like mechanism and prolongs survival.

    Science.gov (United States)

    Fischer, Walter; Gustafsson, Lotta; Mossberg, Ann-Kristin; Gronli, Janne; Mork, Sverre; Bjerkvig, Rolf; Svanborg, Catharina

    2004-03-15

    Malignant brain tumors present a major therapeutic challenge because no selective or efficient treatment is available. Here, we demonstrate that intratumoral administration of human alpha-lactalbumin made lethal to tumor cells (HAMLET) prolongs survival in a human glioblastoma (GBM) xenograft model, by selective induction of tumor cell apoptosis. HAMLET is a protein-lipid complex that is formed from alpha-lactalbumin when the protein changes its tertiary conformation and binds oleic acid as a cofactor. HAMLET induces apoptosis in a wide range of tumor cells in vitro, but the therapeutic effect in vivo has not been examined. In this study, invasively growing human GBM tumors were established in nude rats (Han:rnu/rnu Rowett, n = 20) by transplantation of human GBM biopsy spheroids. After 7 days, HAMLET was administered by intracerebral convection-enhanced delivery for 24 h into the tumor area; and alpha-lactalbumin, the native, folded variant of the same protein, was used as a control. HAMLET reduced the intracranial tumor volume and delayed the onset of pressure symptoms in the tumor-bearing rats. After 8 weeks, all alpha-lactalbumin-treated rats had developed pressure symptoms, but the HAMLET-treated rats remained asymptomatic. Magnetic resonance imaging scans revealed large differences in tumor volume (456 versus 63 mm(3)). HAMLET caused apoptosis in vivo in the tumor but not in adjacent intact brain tissue or in nontransformed human astrocytes, and no toxic side effects were observed. The results identify HAMLET as a new candidate in cancer therapy and suggest that HAMLET should be additionally explored as a novel approach to controlling GBM progression.

  15. Rapid and transient activation of gamma/delta T cells to interferon gamma production, NK cell-like killing and antigen processing during acute virus infection

    Science.gov (United States)

    Gamma/delta T cells are the majority peripheral blood T cells in young cattle. The role of gamma/delta T cells in innate responses against infection with foot-and-mouth disease virus (FMDV) was analyzed on 5 consecutive days following infection. Before infection, bovine gamma/delta T cells expressed...

  16. CD8+ lymphocytes that kill allogeneic and xenogeneic major histocompatibility complex class I targets.

    Science.gov (United States)

    Harms, J S; Splitter, G A

    1995-09-01

    CD8+ CTLs generated in a two-way MLR should lyse target cells only if these targets share a class I MHC allele with the original stimulators. Using cattle PBMCs in a two-way MLR, we generated CD8+ CTLs that kill allogeneic and xenogeneic cell lines. We have named these cells MLK cells. PBMCs isolated from two unrelated animals were cultured together. After 14 days microfluorimetry analysis was performed on the MLK cells with results showing > 90% CD8+ cells. RFLP analysis revealed these cells to be predominately of one animal. MLK cells were then used as effector cells in cytotoxicity assays with syngeneic, allogeneic, and xenogeneic target cells. MLK cells were able to kill all targets. Incubating MLK cells with mAbs to CD8 markedly reduced killing, suggesting a TCR-mediated cytolytic pathway. Effective cytolysis of these targets by MLK cells was dependent on class I expression. MHC class I expression-impaired mutants of allogeneic and xenogeneic targets were not susceptible to cytolysis. Comparisons to LAK cells revealed similarities in phenotype and function to the NK1.1-, CD8+ subset.

  17. Vibriocidal antibody responses to a bivalent killed whole-cell oral cholera vaccine in a phase III trial in Kolkata, India.

    Directory of Open Access Journals (Sweden)

    Suman Kanungo

    Full Text Available BACKGROUND: During the development of a vaccine, identification of the correlates of protection is of paramount importance for establishing an objective criterion for the protective performance of the vaccine. However, the ascertainment of correlates of immunity conferred by any vaccine is a difficult task. METHODS: While conducting a phase three double-blind, cluster-randomized, placebo-controlled trial of a bivalent killed whole-cell oral cholera vaccine in Kolkata, we evaluated the immunogenicity of the vaccine in a subset of participants. Randomly chosen participants (recipients of vaccine or placebo were invited to provide blood samples at baseline, 14 days after the second dose and one year after the first dose. At these time points, serum geometric mean titers (GMT of vibriocidal antibodies and seroconversion rates for vaccine and placebo arms were calculated and compared across the age strata (1 to 5 years, 5 to 15 years and more than 15 years as well as for all age groups. RESULTS: Out of 137 subjects included in analysis, 69 were vaccinees and 68 received placebo. There were 5•7 and 5•8 geometric mean fold (GMF rises in titers to Vibrio cholerae Inaba and Ogawa, respectively at 14 days after the second dose, with 57% and 61% of vaccinees showing a four-fold or greater titer rise, respectively. After one year, the titers to Inaba and Ogawa remained 1•7 and 2•8 fold higher, respectively, compared to baseline. Serum vibriocidal antibody response to V. cholerae O139 was much lower than that to Inaba or Ogawa. No significant differences in the GMF-rises were observed among the age groups. CONCLUSIONS: The reformulated oral cholera vaccine induced a statistically significant anti-O1 Inaba and O1 Ogawa vibriocidal antibody response 14 days after vaccination, which although declined after one year remained significantly higher than baseline. Despite this decline, the vaccine remained protective five years after vaccination.

  18. 5-Fluorouracil-loaded polylactic acid nanoparticles have a killing effect on gastric cancer cell lines in vitro%5-氟尿嘧啶聚乳酸载药纳米微粒对胃癌细胞株的体外杀伤作用

    Institute of Scientific and Technical Information of China (English)

    李晓利; 牛敏; 张铭; 张娜娜; 施瑶

    2015-01-01

    背景:5-氟尿嘧啶在胃癌治疗中占据重要地位,但是长期服用容易出现骨髓抑制、白细胞减少等不良反应。聚乳酸及共聚物载药微粒材料生物相容性较高,分解物不会在机体内发生聚集。  目的:探讨聚乳酸载药纳米微粒对胃癌细胞株的体外杀伤作用机制。  方法:选取10只小鼠进行实验,利用超声乳化方法制备5-氟尿嘧啶聚乳酸载药纳米微粒,并采用噻唑蓝比色法配制1×10-7,1×10-6,1×10-5,1×10-4 mol/L的5-氟尿嘧啶聚乳酸载药纳米微粒,检测其对小鼠胃癌细胞的体外杀伤效应,并且计算出药物的抑制率浓度以及对胃癌细胞的生长抑制能力等以及凋亡的诱导作用。  结果与结论:透射电镜下观察5-氟尿嘧啶聚乳酸载药纳米微粒:形态完好,分布相对均匀,不出现粘连等现象,用药24 h药物浓度可达到50%,72 h后能够达到62.9%;不同浓度下单一5-氟尿嘧啶和5-氟尿嘧啶聚乳酸载药纳米微粒和小鼠胃癌细胞联合培养48 h后,细胞的活性随着药物浓度的提高出现下降趋势,并且5-氟尿嘧啶聚乳酸载药纳米微粒细胞抑制能力显著高于5-氟尿嘧啶(P OBJECTIVE:To investigate the in vitro cytotoxicity mechanism of 5-fluorouracil-loaded polylactic acid nanoparticles on gastric cancer cel lines. METHODS:Ten mice were selected in this study. 5-fluorouracil-loaded polylactic acid nanoparticles (1×10-7, 1×10-6, 1×10-5, 1×10-4 mol/L) were prepared using ultrasonic emulsification method. Kiling effect of polylactic acid nanoparticles on gastric cancer cel lines in vitrowere detected. Then, the inhibition rate was calculated at different concentrations. RESULTS AND CONCLUSIONS: Under the transmission electron microscope, 5-fluorouracil-loaded polylactic acid nanoparticles had good shape and relatively evenly distributed with no adhesions. After drug administration, the drug concentration was

  19. The enhanced effect of lupeol on the destruction of gastric cancer cells by NK cells.

    Science.gov (United States)

    Wu, Xiao-Ting; Liu, Jun-Quan; Lu, Xiao-Ting; Chen, Fu-Xing; Zhou, Zhong-Hai; Wang, Tao; Zhu, Sheng-Ping; Fei, Su-Juan

    2013-06-01

    Lupeol, a triterpene, was reported to possess beneficial effects as a therapeutic and preventive agent for a range of disorders. Many studies have confirmed that lupeol possesses strong activities such as antioxidative, antiinflammatory, antiarthritic, antimutagenic, and antimalarial, both in vitro and in vivo, and at its effective therapeutic doses exhibit no toxicity to normal cells and tissues. Lupeol was observed to inhibit the proliferation of gastric tumour cells in a dose-dependent manner, as assessed by MTT assay, and induce the proliferation of NK cells, as assessed by flow cytometry and Western blotting. The killing effect of NK cells on gastric tumour cells was assessed by LDH. Our experiment demonstrated that lupeol at appropriate concentrations could promote the proliferation of NK cells, inhibit the proliferation of gastric cancer cell lines BGC823, N87 and HGC27, and increase the killing effect of NK cells on gastric cancer cells. We speculated that lupeol might increase the expression of PFP, IFN-γ, and CD107a via the activation of PI3K/Akt and Wnt/β-catenin signalling pathways. Lupeol could serve as a potential agent against gastric cancer; however, further in-depth in vivo studies are still required.

  20. Effects of External Electric Field on AlN Precipitation and Recrystallization Texture of Deep-drawing 08Al Killed Steel Sheet

    Institute of Scientific and Technical Information of China (English)

    Xiang ZHAO; Zhuochao HU; Liang ZUO

    2006-01-01

    The effects of an electric field on AlN precipitation and recrystallization texture were investigated. Cold-rolled 08Al killed steel sheets were annealed at 550℃ according to the two-step processes, for various maintaining times, with and without applying an electric field. It was found that the electric field promotes the precipitation of the second phase (AlN particles), strengthens the γ-fiber and weakens the α-fiber texture component in the recrystallized specimens. A possible explanation for the reinforcement of γ-fiber texture by the electric field is that the second phase AlN particle promotes the growth of γ-fiber at the expense of differently oriented grains.

  1. Adenovirus mediated fusion gene system driven by KDR promoter kills selectively pancreatic cancer cells%双自杀基因重组腺病毒对胰腺癌细胞特异性杀伤作用

    Institute of Scientific and Technical Information of China (English)

    闫振宇; 陈旭; 孔恒; 黄宗海; 俞金龙; 厉周

    2008-01-01

    目的 研究腺病毒介导的KDR启动子驱动CD/TK融合基因系统(Ad-KDR-CDTK)对胰腺癌细胞Capan-2特异性的杀伤作用.方法 重组腺病毒体外感染表达KDR的Capaw2细胞株,用不表达KDR的肝癌细胞HepG2做对照.观察其感染效率并以RT-PCR方法 检测转基因细胞CDTK的表达,然后给予不同浓度的前药更昔洛韦(ganciclovir,GCV)和5-氟胞嘧啶(5-fluorocy-tosine,5-FC),MTT法观察该体系对Capan-2和HepG2细胞生长增殖的影响及其旁观者效应;电镜观察细胞的病变;流式细胞仪检测细胞周期的变化和DNA含量的变化.建立Capan-2裸鼠皮下移植瘤模型,瘤内注射Ad-KDR-CD/TK,腹腔注射前药GCV(50 mg·kg-1·d-1)和5-FC(500 mg·kg-1·d-1)14 d,观察肿瘤生长抑制效应.结果 腺病毒对两种细胞株的感染率相似,其感染率随腺病毒滴度的增高而递增.RT-PCR方法 检测发现转染Ad-KDR-CDTK的Capan-2细胞有目的 基因表达.MTT法检测显示前药呈剂量依赖性抑制Capan-2生长,而不表达KDR的肝癌细胞HepG2对前药不敏感,且观察到该体系对Capan-2明显的旁观者效应.电镜下可见Capan-2有凋亡改变.用流式细胞仪测定用药组出现典型的凋亡峰;细胞周期分析显示治疗后细胞G0-G1期比率增多,G2-M及S期细胞减少.在Capan-2裸鼠移植瘤模型中,该双自杀基因系统能够显著抑制肿瘤的生长.结论 KDR启动子可调控双自杀基因体系选择性杀伤胰腺癌细胞Capan-2,诱导胰腺癌细胞凋亡,并可显著抑制人胰腺癌裸鼠移植瘤的生长.%Objective To evaluate the selectively killing effect of adenovirus (Ad) mediated double suicide gene driven by KDR promoter on pancreatic cancer cell Capan-2. Methods KDR-ex-pressing Capan-2 and non-KDR-expressing HepG2 were infected by Ad-KDR-CDTK. The infection rate was observed and the expression of CDTK was detected by RT-PCR. Followed by treatment with 5-FC and GCV,the killing effects were evaluated and bystander effects

  2. THE KILLING OF COLON BACILLI BY ULTRAVIOLET LIGHT.

    Science.gov (United States)

    Wyckoff, R W

    1932-01-20

    The survival ratios of colon bacilli subjected to several monochromatic ultraviolet radiations follow semilogarithmic straight lines. For each wave length approximate observations have been made of the energy involved in cell destruction. This energy varies somewhat with frequency in the ultraviolet region; it is furthermore nearly one hundred times as great as the amount of X-ray energy required to bring about the same killing. Preliminary experiments show no measurable difference either in rate of killing or in lethal energy between B. coli and B. aertrycke. Parallel results have already been obtained with X-rays and electrons. The data from colon bacilli are interpreted in terms of the assumptions employed for X-rays. They indicate that though bacterial death should result from a single quantum absorption, millions more such absorptions seemingly are without injurious effect on cell growth and multiplication. The "sensitive volume" within which, according to this picture, the lethal quantum must be stopped proves to be about the same as that of a single protein molecule. If this is the correct description of the phenomena of ultraviolet killing, it seems strange that the millions of non-deadly quanta absorbed per bacillus should not show themselves by altered growth rates or in other ways. That they apparently do not suggests the inapplicability of the statistical picture. The death rate under this kind of radiation then would be primarily an expression of the relative sensitivities of the bacterial population. Additional experiments are required to determine this question.

  3. Killing effect of EGFR-TKI combined with 125I seed implantation therapy onⅢB-Ⅳ stage lung cancer tissue

    Institute of Scientific and Technical Information of China (English)

    Ai-Sheng Xiang

    2016-01-01

    Objective:To analyze the killing effect of EGFR-TKI combined with 125I seed implantation therapy onⅢB-Ⅳ stage lung cancer tissue.Methods:A total of 78 patients withⅢB-Ⅳ stage lung cancer were randomly divided into observation group and control group (n=39), control group received EGFR-TKI treatment and observation group received EGFR-TKI combined with 125I seed implantation therapy. Differences in apoptosis gene, invasion gene and autophagy gene expression in lung tissue were compared between two groups after 1 month of treatment.Results:Apoptosis genesPDCD5, bax andbcl-xSmRNA expression levels in lung tissue of observation group after 1 month of treatment were higher than those of control group whileBag-1, survivin andbcl-xL mRNA expression levels were lower than those of control group; invasion genesCD147, EGFRandDDX17 mRNA expression levels were lower than those of control group while Bin1, E-cadherin andOvol2mRNA expression levels were higher than those of control group; autophagy genes ARHI, Beclin1, Atg5, LC3B, pULK andPI3KC3 mRNA expression levels were higher than those of control group.Conclusions: EGFR-TKI combined with 125I seed implantation therapy can enhance the tumor killing effect on patients withⅢB-Ⅳ stage lung cancer, and contribute to the optimization of overall condition and the extension of survival time.

  4. Chlordiazepoxide and diazepam induced mouse killing by rats.

    Science.gov (United States)

    Leaf, R C; Wnek, D J; Gay, P E; Corcia, R M; Lamon, S

    1975-10-14

    Chlordiazepoxide HCl, at dose levels from 2.5 mg/kg to 80 mg/kg, significantly increased the low base rates of mouse killing (3-9%) observed in large samples (N = 100/dose) of Holtzman strain albino male rats. Maximal killing rates were obtained at doses from 7.5 mg/kg to 20 mg/kg. Diazepam was equally effective, and several times more potent than chlordiazepoxide. Pentobarbital did not increase killing. Killing induced by chlordiazepoxide was blocked by d-amphetamine SO4, but not by l-amphetamine, at dose levels similar to those that block undrugged killing in this strain (ED50 = 1.5 mg/kg). Unlike pilocarpine-induced killing, the effects of chlordiazepoxide were not increased or decreased significantly by either peripherally or centrally active anticholinergic drugs, over wide dose ranges of these agents; nor were the effects of chlordiazepoxide increased by repeated daily administration.

  5. Kill the Killers: terapia con células Natural Killer en pacientes pediátricos con cáncer refractario Kill the Killers: Natural Killer cells therapy against paediatric refractory solid tumours

    Directory of Open Access Journals (Sweden)

    J. Valentín Quiroga

    2012-09-01

    Full Text Available Introducción: En el momento actual, los tumores sólidos refractarios al tratamiento convencional constituyen la principal causa de muerte en la edad pediátrica. Por tanto, es necesario desarrollar y consolidar nuevos tratamientos. Las células Natural Killer (NK constituyen la primera línea de defensa del sistema inmune frente al desarrollo de células tumorales. Planteamos una nueva estrategia de terapia celular antitumoral en niños con cánceres refractarios, inmunoterapia con células NK estimuladas con interleucina 15 (IL-15. Pacientes y Métodos: En 22 pacientes pediátricos con tumores sólidos refractarios y en controles sanos determinamos mediante citometría de flujo multiparamétrica y fluorescencia resuelta en el tiempo, el fenotipo y la actividad citotóxica de las células NK, respectivamente. En ratones inmunodeficientes desarrollamos un modelo de neuroblastoma metastático muy agresivo y terapia de rescate con células Natural Killer estimuladas con IL-15. Resultados: Los pacientes pediátricos con cáncer refractario tienen un mayor porcentaje de células NK bright y una menor actividad citotóxica. La estimulación con IL-15 mejora la citotoxicidad in vitro y disminuye la carga tumoral in vivo. Conclusiones: Las células NK estimuladas con IL-15 constituyen una prometedora estrategia antitumoral.Introduction: Refractory solid tumours lead children deaths. To change this statement, new treatments should be developed. Natural Killer cells constitute the first line of defence against tumour cells. We propose a new strategy for antitumor cell therapy in children with refractory solid malignancies: IL-15 stimulated NK cells. Patients and methods: 22 paediatric patients suffering refractory solid tumours participate in this study. We compare NK cell subsets and K562 cytotoxicity in patients and sex-age pair's healthy controls. We use multiparametric flow and time-resolved fluorescent, respectively. In immunocompromised mice we

  6. Small CD4 Mimetics Prevent HIV-1 Uninfected Bystander CD4+ T Cell Killing Mediated by Antibody-dependent Cell-mediated Cytotoxicity

    Directory of Open Access Journals (Sweden)

    Jonathan Richard

    2016-01-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 infection causes a progressive depletion of CD4+ T cells. Despite its importance for HIV-1 pathogenesis, the precise mechanisms underlying CD4+ T-cell depletion remain incompletely understood. Here we make the surprising observation that antibody-dependent cell-mediated cytotoxicity (ADCC mediates the death of uninfected bystander CD4+ T cells in cultures of HIV-1-infected cells. While HIV-1-infected cells are protected from ADCC by the action of the viral Vpu and Nef proteins, uninfected bystander CD4+T cells bind gp120 shed from productively infected cells and are efficiently recognized by ADCC-mediating antibodies. Thus, gp120 shedding represents a viral mechanism to divert ADCC responses towards uninfected bystander CD4+ T cells. Importantly, CD4-mimetic molecules redirect ADCC responses from uninfected bystander cells to HIV-1-infected cells; therefore, CD4-mimetic compounds might have therapeutic utility in new strategies aimed at specifically eliminating HIV-1-infected cells.

  7. Alternative to antibiotics against Pseudomonas aeruginosa: Effects of Glycyrrhiza glabra on membrane permeability and inhibition of efflux activity and biofilm formation in Pseudomonas aeruginosa and its in vitro time-kill activity.

    Science.gov (United States)

    Chakotiya, Ankita Singh; Tanwar, Ankit; Narula, Alka; Sharma, Rakesh Kumar

    2016-09-01

    The multi-drug resistance offered by Pseudomonas aeruginosa to antibiotics can be attributed towards its propensity to develop biofilm, modification in cell membrane and to efflux antibacterial drugs. The present study explored the activity of Glycyrrhiza glabra and one of its pure compounds, glycyrrhizic acid against P. aeruginosa and their mechanism of action in terms of the effect on membrane permeability, efflux activity, and biofilm formation were determined. Minimum inhibitory concentrations were determined by using broth dilution technique. The minimum bactericidal concentrations were assessed on agar plate. The MIC of the extract and glycyrrhizic acid was found to be 200 and 100 μg ml(-1), respectively. The MBC was found to be 800 and 400 μg ml(-1) in the case of extract and glycyrrhizic acid, respectively. Time -dependent killing efficacy was also estimated. Flowcytometric analysis with staining methods was used to determine the effect of extract and glycyrrhizic acid at 2 × MIC on different physiological parameters and compared it with the standard (antibiotic). The growth of P. aeruginosa was significantly inhibited by extract and the pure compound. The herbal extract and the glycyrrhic acid were also found to effective in targeting the physiological parameters of the bacteria that involve cell membrane permeabilization, efflux activity, and biofilm formation. This study reports the antipseudomonal action of Glycyrrhiza glabra and one of its compound and provides insight into their mode of action.

  8. Effects of bortezomib in sensitizing human prostate cancer cell lines to NK-mediated cytotoxicity

    Institute of Scientific and Technical Information of China (English)

    Wei Hu; Rui-Rui Zheng; Hui-Xia Cui; Dan Yue; Yong Wang; You-Hong Jiang

    2012-01-01

    The proteasome inhibitor,bortezomib,has been demonstrated to sensitize tumor cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis.Natural killer (NK) cells represent potent antitumor effector cells.They also express TRAIL.Therefore.we investigated whether bortezomib could sensitize tumor cells to NK cell-mediated killing,and have the same effect in human prostate cancer cell lines (LNCaP and DU145).We found that bortezomib strongly inhibits proliferation in both cell lines.Furthermore,compared with LNCaP cells,DU 145 cells are more sensitive to bortezomib-induced apoptosis.However,bortezomib is unable to sensitize these two cell lines to NK cell-mediated killing in short-term assays.In long-term assays,we found that killing mediated by activated NK cells following bortezornib treatment leads to greater antitumor effects than either treatment alone.In addition,treatment with bortezomib causes these cells to upregulate apoptosis-related mRNA as well as death receptors and downregulate the major histocompatibility class (MHC)-Ⅰ molecule on the cell surface of DU145 cells.In contrast,LNCaP cells are not sensitized by this treatment.Death receptors and the MHC-Ⅰ molecule did not change in this cell line.These-data suggest that bortezomib can be used to sensitize prostate cancer cells to NK cell-mediated killing and improve current cancer therapies.This therapeutic strategy may be more effective in patients with androgen-insensitive prostate cancer.

  9. Deprive to kill: Glutamine closes the gate to anticancer monocarboxylic drugs

    OpenAIRE

    Cardaci, Simone; Ciriolo, Maria Rosa

    2012-01-01

    Killing properties of antitumor drugs can be enhanced by strategies targeting biochemical adaptations of cancer cells. Recently, we reported that depriving cancer cells of glutamine is a feasible approach to enhance antitumor effects of the alkylating analog of pyruvic acid, 3-bromopyruvate, which rely on the induction of autophagic cell death by metabolic-oxidative stress. 3-bromopyruvate chemopotentiation is the result of its increased intracellular uptake mediated by the monocarboxylate tr...

  10. 食管癌抗原联合金葡素诱导杀瘤性细胞的实验研究%Study on Tumor-Killing Cells Induced by Esophageal Cancer Antigen and Staphlococcal Enterotoxin

    Institute of Scientific and Technical Information of China (English)

    李树义; 刘洪梅; 张庆波

    2011-01-01

    目的:体外应用超抗原金葡素C型(SEC)联合食管癌可溶性抗原(TSA)刺激外周血淋巴细胞,诱导产生细胞毒T细胞(CTLs),对肿瘤细胞进行杀伤,探讨其治疗肿瘤的可行性.方法:外周血淋巴细胞经TSA、超抗原SEC联合作用,进行体外培养.用酶联免疫吸附测定法(ELISA)法和细胞毒试验测定细胞因子含量和杀伤活性.结果:经TSA与超抗原SEC联合刺激,淋巴细胞增殖活性增强,在刺激后72 h达到峰值;与空白对照组相比,SEC、TSA以及SEC与TSA联合刺激,人肿瘤坏死因子α(TNF-α)活性从第1天开始升高,在3 d 达峰值;并且联合刺激的淋巴细胞组诱导的效应细胞对靶细胞杀伤活性显著高于单纯淋巴细胞组(P<0.05),对TSA来源的食管癌细胞具有选择性杀伤作用.结论:肿瘤可溶性抗原与超抗原SEC联合应用能诱导效应细胞明显增殖、活化、并产生高效特异性的抗肿瘤效果.%Objective:To study the effect of tumor soluble antigen (TSA) and staphlococcal enterotoxin(SEC) on the cyctotoxic T lymphocyte(CTL), and to evaluate the new cell immunotherapy on tumor. Methods: Lymphocyte cells were isolated from peripheral blood and then stimulated in vitro with soluble antigen and superantigen SEC, test levels of cytokine and killing activity by enzyme-linked immunosorbent assay (ELISA) and cytotoxic assay. Results: Proliferation activity of lymphocyte cells group stimulated by TSA and SEC was the strongest, peaked at 72h. Compared with the control group, the levels of tumor necrosis factor-α (TNF-α) in the study groups begun to increase after 1d, and peaked at 3d. The killing activity of lymphocyte cell group stimulated by TSA and SEC against target cells were significantly higher than the control group(P<0.05). Conclusions:The seffector cells stimulated by TSA and SEC induce effective proliferation,activation and high antitumor efficiency and show a potential kind of operative antitumor immunotherapy.

  11. Analysing the Wrongness of Killing

    DEFF Research Database (Denmark)

    Di Nucci, Ezio

    2014-01-01

    This article provides an in-depth analysis of the wrongness of killing by comparing different versions of three influential views: the traditional view that killing is always wrong; the liberal view that killing is wrong if and only if the victim does not want to be killed; and Don Marquis‟ future...... of value account of the wrongness of killing. In particular, I illustrate the advantages that a basic version of the liberal view and a basic version of the future of value account have over competing alternatives. Still, ultimately none of the views analysed here are satisfactory; but the different...... reasons why those competing views fail provide important insights into the ethics of killing....

  12. Killing Spinors -- Beyond Supergravity

    CERN Document Server

    Palomo-Lozano, Alberto

    2012-01-01

    This is a doctoral thesis on the application of techniques originally developed in the programme of characterisation of supersymmetric solutions to Supergravity theories, to finding alternative backgrounds. We start by discussing the concept of a Killing spinor, and how these are paramount to the process of classifying of these aforementioned supersymmetric solutions. Moreover, these geometric objects also have applications when considered in different scenarios (the 'beyond' in the title). In particular, techniques based on a parallelising rule for a spinorial field can be used for obtaining solutions to Einstein-Maxwell-De Sitter theories, as well as a (partial) classification of Lorentzian Einstein-Weyl manifolds, a problem of geometrical interest. The annexe contain an introduction and summary in Spanish language. The appendices discuss the tensorial and spinorial conventions employed, some relevant geometrical information on the scalar manifolds for the matter contents of interest, as well as for the nul...

  13. Killing Symmetry on Finsler Manifold

    CERN Document Server

    Ootsuka, Takayoshi; Ishida, Muneyuki

    2016-01-01

    Killing vector fields $K$ are defined on Finsler manifold. The Killing symmetry is reformulated simply as $\\delta K^\\flat =0$ by using the Killing non-linear 1-form $K^\\flat$ and the spray operator $\\delta$ with the Finsler non-linear connection. $K^\\flat$ is related to the generalization of Killing tensors on Finsler manifold, and the condition $\\delta K^\\flat =0$ gives an analytical method of finding higher derivative conserved quantities, which may be called hidden conserved quantities. We show two examples: the Carter constant on Kerr spacetime and the Runge-Lentz vectors in Newtonian gravity.

  14. Combinatorial stresses kill pathogenic Candida species.

    Science.gov (United States)

    Kaloriti, Despoina; Tillmann, Anna; Cook, Emily; Jacobsen, Mette; You, Tao; Lenardon, Megan; Ames, Lauren; Barahona, Mauricio; Chandrasekaran, Komelapriya; Coghill, George; Goodman, Daniel; Gow, Neil A R; Grebogi, Celso; Ho, Hsueh-Lui; Ingram, Piers; McDonagh, Andrew; de Moura, Alessandro P S; Pang, Wei; Puttnam, Melanie; Radmaneshfar, Elahe; Romano, Maria Carmen; Silk, Daniel; Stark, Jaroslav; Stumpf, Michael; Thiel, Marco; Thorne, Thomas; Usher, Jane; Yin, Zhikang; Haynes, Ken; Brown, Alistair J P

    2012-10-01

    Pathogenic microbes exist in dynamic niches and have evolved robust adaptive responses to promote survival in their hosts. The major fungal pathogens of humans, Candida albicans and Candida glabrata, are exposed to a range of environmental stresses in their hosts including osmotic, oxidative and nitrosative stresses. Significant efforts have been devoted to the characterization of the adaptive responses to each of these stresses. In the wild, cells are frequently exposed simultaneously to combinations of these stresses and yet the effects of such combinatorial stresses have not been explored. We have developed a common experimental platform to facilitate the comparison of combinatorial stress responses in C. glabrata and C. albicans. This platform is based on the growth of cells in buffered rich medium at 30°C, and was used to define relatively low, medium and high doses of osmotic (NaCl), oxidative (H(2)O(2)) and nitrosative stresses (e.g., dipropylenetriamine (DPTA)-NONOate). The effects of combinatorial stresses were compared with the corresponding individual stresses under these growth conditions. We show for the first time that certain combinations of combinatorial stress are especially potent in terms of their ability to kill C. albicans and C. glabrata and/or inhibit their growth. This was the case for combinations of osmotic plus oxidative stress and for oxidative plus nitrosative stress. We predict that combinatorial stresses may be highly significant in host defences against these pathogenic yeasts.

  15. Cytotoxic macrophage-released tumour necrosis factor-alpha (TNF-α) as a killing mechanism for cancer cell death after cold plasma activation

    Science.gov (United States)

    Kaushik, Nagendra Kumar; Kaushik, Neha; Min, Booki; Choi, Ki Hong; Hong, Young June; Miller, Vandana; Fridman, Alexander; Choi, Eun Ha

    2016-03-01

    The present study aims at studying the anticancer role of cold plasma-activated immune cells. The direct anti-cancer activity of plasma-activated immune cells against human solid cancers has not been described so far. Hence, we assessed the effect of plasma-treated RAW264.7 macrophages on cancer cell growth after co-culture. In particular, flow cytometer analysis revealed that plasma did not induce any cell death in RAW264.7 macrophages. Interestingly, immunofluorescence and western blot analysis confirmed that TNF-α released from plasma-activated macrophages acts as a tumour cell death inducer. In support of these findings, activated macrophages down-regulated the cell growth in solid cancer cell lines and induced cell death in vitro. Together our findings suggest plasma-induced reactive species recruit cytotoxic macrophages to release TNF-α, which blocks cancer cell growth and can have the potential to contribute to reducing tumour growth in vivo in the near future.

  16. An impedance-based cytotoxicity assay for real-time and label-free assessment of T-cell-mediated killing of adherent cells.

    Science.gov (United States)

    Peper, Janet Kerstin; Schuster, Heiko; Löffler, Markus W; Schmid-Horch, Barbara; Rammensee, Hans-Georg; Stevanović, Stefan

    2014-03-01

    The in vitro assessment of T-cell-mediated cytotoxicity plays an important and increasingly relevant role both in preclinical target evaluation and during immunomonitoring to accompany clinical trials employing targeted immunotherapies. For a long time, the gold standard for this purpose has been the chromium release assay (CRA). This end point assay, however, shows several disadvantages including the inevitable use of radioactivity. Based on electrical impedance measurements (using the xCELLigence system), we have established a label-free assay, facilitating the real-time monitoring of T-cell-mediated cytotoxicity. The coculture of peptide-specific T-cell lines with peptide-loaded target cells reproducibly led to a decrease in impedance due to induced apoptosis and detachment of target cells. Comparing our results to the standard CRA assay, we could demonstrate that impedance-based measurements show comparable results after short incubation periods (6h) but outperform the CRA both in reproducibility and sensitivity after prolonged incubation (24h), enabling the detection of target cell lysis with an effector to target ratio as low as 0.05:1. The impedance-based assay represents a valuable and highly sensitive tool for label-free real-time high throughput analysis of T-cell-mediated cytotoxicity.

  17. Expression of thymidine kinase mediated by a novel non-viral delivery system under the control of vascular endothelial growth factor receptor 2 promoter selectively kills human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Ying Wang; Hui-Xiong Xu; Ming-De Lu; Qing Tang

    2008-01-01

    AIM: To investigate the killing efficiency of a recombinant plasmid containing a thymidine kinase (TK) domain insert driven by the vascular endothelial growth factor receptor 2 (VEGFR2) promoter (KDR) on vascular endothelial cells.METHODS: The KDR-TK fragment was extracted from pBluescript 11 KDR-TK plasmid by enzymatic digestion with XhoI and Sa/I. The enhanced green fluorescence protein (EGFP) carrier was extracted from pEGFP by the same procedure. The KDR-TK was inserted into the pEGFP carrier to construct pEGFP-KDR-TK. Using ultrasound irradiation and microbubble,pEGFP-KDR-TK was transferred into human umbilical vein endothelial cells (HUVECs). The transient infection rate was estimated by green fluorescent protein (GFP)expression. Transfected HUVECs, non-transfected HUVECs, and HepG2 cells were cultured in the presence of different concentrations of ganciclovir (GCV), and the killing efficacy of HSV-TK/GCV was analyzed by 3-[4,5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTr) assay.RESULTS: The recombinant pEGFP-KDR-TK was successfully constructed by inserting the KDR-TK fragment into the pEGFP carrier. Transfected HUVECs showed cytoplasmic green fluorescence, and the transient transfection rate was about 20.3%. Pools of G418-resistant cells exhibited a higher sensitivity to the prodrug/GCV compared to non-transfected HUVECs or non-transfected HepG2 cells, respectively.CONCLUSION: KDR promoter and the suicide gene/prodrug system mediated by diagnostic ultrasound combined with microbubble can significantly kill HUVECs.Such therapy may present a novel and attractive approach to target gene therapy on tumor vessels.

  18. Enhancement of the immune response against Salmonella infection of mice by heat-killed multispecies combinations of lactic acid bacteria.

    Science.gov (United States)

    Chen, Chih-Yuan; Tsen, Hau-Yang; Lin, Chun-Li; Lin, Chien-Ku; Chuang, Li-Tsen; Chen, Chin-Shuh; Chiang, Yu-Cheng

    2013-11-01

    Heat-killed lactic acid bacteria (LAB) has advantages over live LAB in that it has a long shelf-life and is therefore easy to store and transport. From four LAB strains selected by immunomodulatory activity and adherent properties, we prepared the heat-killed multispecies combination of LAB (MLAB) and the cell walls from MLAB under two conditions (100 °C for 30 min and 121 °C for 15 min). Different effects on the adherent properties of these four LAB strains were observed, depending on the heating conditions. With mouse macrophage cells, the two heat-killed MLABs (HMLABs) showed significantly higher induction activities on the production of interleukin 12 (IL-12) than their individual strains did. Heat-killed MLABs and cell-wall preparations were able to reduce the Salmonella invasion of Caco-2 and mouse macrophage cells. Feeding mice with HMLAB could inhibit the Salmonella invasion of mice significantly. For these mice, the expression level of pro-inflammatory cytokines, such as TNF-α and IL-6, in mouse serum was reduced while that of the anti-inflammatory cytokine, i.e. IL-10, was enhanced. The HMLABs developed in this study showed higher protective effect against Salmonella invasion either of Caco-2 cells or of mice, relative to the heat-killed lactobacilli, which consisted of Lactobacillus acidophilus strains selected at random. In conclusion, the HMLABs were potentially useful for the protection of mice against Salmonella infection and the induced inflammation.

  19. TGF-β-induced CD4+Foxp3+ T cells attenuate acute graft-versus-host disease via suppressing expansion and killing of effector CD8+ cells

    OpenAIRE

    Gu, Jian; Lu, Ling; Chen, Maogen; Xu, Lili; Lan, Qin; Li, Qiang; Liu, Zhongmin; Chen, Guihua; Wang, Ping; Wang, Xuehao; Brand, David; Olsen, Nancy; Zheng, Song Guo

    2014-01-01

    TGF-β-induced CD4+Foxp3+ T cells (iTregs) have been identified as important prevention and treatment strategies for cell therapy in autoimmune diseases and other disorders. However, the potential use of iTregs as a treatment modality for acute graft-verse-host disease (GVHD) has not been realized because iTregs may be unstable and less suppressive in this disease. Here we restudied the ability of iTregs to prevent and treat acute GVHD in two different mouse models. Our results showed that so ...

  20. Tailored Nanoparticle Codelivery of antimiR-21 and antimiR-10b Augments Glioblastoma Cell Kill by Temozolomide: Toward a "Personalized" Anti-microRNA Therapy.

    Science.gov (United States)

    Ananta, Jeyarama S; Paulmurugan, Ramasamy; Massoud, Tarik F

    2016-09-01

    Glioblastoma remains an aggressive brain malignancy with poor prognosis despite advances in multimodal therapy that include standard use of Temozolomide. MicroRNA-21 (miR-21) and microRNA-10b (miR-10b) are oncomiRs overexpressed in glioblastoma, promoting many aspects of cancer biology. We hypothesized that PLGA nanoparticles carrying antisense miR-21 (antimiR-21) and antisense miR-10b (antimiR-10b) might beneficially knockdown endogenous miR-21 and miR-10b function and reprogram cells prior to Temozolomide treatment. PLGA nanoparticles were effective in intracellular delivery of encapsulated oligonucleotides. Concentrations of delivered antimiR-21 and antimiR-10b were optimized and specifically tailored to copy numbers of intracellular endogenous microRNAs. Coinhibition of miR-21 and miR-10b significantly reduced the number of viable cells (by 24%; p < 0.01) and increased (2.9-fold) cell cycle arrest at G2/M phase upon Temozolomide treatment in U87 MG cells. Cell-tailored nanoparticle-assisted concurrent silencing of miR-21 and miR-10b prior to Temozolomide treatment is an effective molecular therapeutic strategy in cell culture, warranting the need for further studies prior to future in vivo "personalized" medicine applications.

  1. Effects of disrupting the polyketide synthase gene WdPKS1 in Wangiella [Exophiala] dermatitidis on melanin production and resistance to killing by antifungal compounds, enzymatic degradation, and extremes in temperature

    Directory of Open Access Journals (Sweden)

    Mandal Piyali

    2006-06-01

    Full Text Available Abstract Background Wangiella dermatitidis is a human pathogenic fungus that is an etiologic agent of phaeohyphomycosis. W. dermatitidis produces a black pigment that has been identified as a dihydroxynaphthalene melanin and the production of this pigment is associated with its virulence. Cell wall pigmentation in W. dermatitidis depends on the WdPKS1 gene, which encodes a polyketide synthase required for generating the key precursor for dihydroxynaphthalene melanin biosynthesis. Results We analyzed the effects of disrupting WdPKS1 on dihydroxynaphthalene melanin production and resistance to antifungal compounds. Transmission electron microscopy revealed that wdpks1Δ-1 yeast had thinner cell walls that lacked an electron-opaque layer compared to wild-type cells. However, digestion of the wdpks1Δ-1 yeast revealed small black particles that were consistent with a melanin-like compound, because they were acid-resistant, reacted with melanin-binding antibody, and demonstrated a free radical signature by electron spin resonance analysis. Despite lacking the WdPKS1 gene, the mutant yeast were capable of catalyzing the formation of melanin from L-3,4-dihyroxyphenylalanine. The wdpks1Δ-1 cells were significantly more susceptible to killing by voriconazole, amphotericin B, NP-1 [a microbicidal peptide], heat and cold, and lysing enzymes than the heavily melanized parental or complemented strains. Conclusion In summary, W. dermatitidis makes WdPKS-dependent and -independent melanins, and the WdPKS1-dependent deposition of melanin in the cell wall confers protection against antifungal agents and environmental stresses. The biological role of the WdPKS-independent melanin remains unclear.

  2. Pathophysiological and immune cell responses in calves prior to and following lung challenge with formalin-killed Pasteurella multocida biotype A:3 and protection studies involving subsequent homologous live challenge.

    Science.gov (United States)

    Dowling, A; Hodgson, J C; Dagleish, M P; Eckersall, P D; Sales, J

    2004-08-01

    Pneumonic pasteurellosis is a common respiratory infection in cattle that has major economic and welfare implications world-wide and the incidence in the UK due to Pasteurella multocida, currently the same as that associated with Mannheimia haemolytica, is increasing. Whereas much is known regarding the pathogenesis of M. haemolytica infections little information is available on the pathogenic process of pasteurellosis initiated by P. multocida. In the present work calf systemic and innate immune responses to intratracheal challenge with formalin-killed P. multocida biotype A:3 and to subsequent experimental lung infection with live P. multocida were investigated. Eight-week-old calves were challenged intratracheally on day 0 with either 10(9) colony forming units (cfu) of formalin-killed P. multocida biotype A:3 in 300 ml saline (n = 10) or 300 ml saline alone (n = 10), followed, at day 21, by challenge with 10(9) cfu live P. multocida. Pathophysiological and lung phagocyte responses were assessed by clinical monitoring, sequential lung lavage and blood sampling. Results for samples obtained before, during and after challenge showed clinical and acute phase protein responses to both bacterial culture and saline control treatments, although higher responses were associated with bacterial challenge. Phagocytosis of P. multocida during 1h incubation periods with lavaged cells in vitro was unaffected by exposure in vivo to killed P. multocida and there was evidence that P. multocida was able to survive intracellularly during this assay. There was no indication that lung exposure to formalin-killed P. multocida conferred protection against subsequent homologous live challenge.

  3. Short communication: Antiproliferative effect of 8 different Lactobacillus strains on K562 cells.

    Science.gov (United States)

    Tuo, Yanfeng; Jiang, Shujuan; Qian, Fang; Mu, Guangqing; Liu, Peng; Guo, Yuanji; Ma, Changlu

    2015-01-01

    Some strains of Lactobacillus genus have antiproliferative activities against cancer cells. However, until now, the exact effector molecules of Lactobacillus strains with anticancer activity have not been identified. The aim of the present study was to explore which fraction of the Lactobacillus cells exerts the highest antiproliferative effect. For this purpose, the heat-killed bacterial cells, bacterial cell wall extract, and genomic DNA of 8 Lactobacillus strains were prepared to assess their antiproliferative activities against human myeloid leukemia cell lines K562. The heat-killed bacterial cells of the 8 lactobacilli strains exerted antiproliferative effect on K562 cells, and the inhibition rates exerted by the heat-killed bacterial cells of the strains G15AL, M5AL, SB31AL, SB5AL, and T3AL were significantly higher than those exerted by the cell walls and genomic DNA of the strains. The bacterial DNA of G15AL exerted higher antiproliferative effect on K562 cells. The exact effector molecules and the effect mechanism of the strains should be further explored for the application of these strains as probiotic strains or bioactive probiotic molecules.

  4. Killing by neutrophil extracellular traps: fact or folklore?

    Science.gov (United States)

    Menegazzi, Renzo; Decleva, Eva; Dri, Pietro

    2012-02-02

    Neutrophil extracellular traps (NETs) are DNA structures released by dying neutrophils and claimed to constitute a new microbicidal mechanism. Killing by NET-forming cells is ascribed to these structures because it is prevented by preincubation with DNase, which has been shown to dismantle NETs, before addition of the target microorganisms. Curiously, the possibility that the microorganisms ensnared in NETs are alive has not been considered. Using Staphylococcus aureus and Candida albicans blastospores, we demonstrate that the microorganisms captured by NETs and thought to be killed are alive because they are released and recovered in cell medium by incubation with DNase. It is concluded that NETs entrap but do not kill microbes.

  5. The effect of the volatile oil of Manchurian Wildginger on the killing of Demodex in vitro%细辛精油体外抑杀蠕形螨的实验研究

    Institute of Scientific and Technical Information of China (English)

    刘继鑫; 李朝品; 孙艳宏; 孙艳; 刘伯阳; 刘秀财; 郭家; 张浩

    2014-01-01

    Objective To observe the effect of the volatile oil of Manchurian Wildginger on the killing of demodex in vitro. Methods The volatile oil of Manchurian Wildginger and its diluents of 1∶2, 1∶4, 1∶8, 1∶16 and 1∶32 proportion were used to kill demodex in vitro, and the saline was used as control group. The mites were collected with adhesive cellophane tape technique. The killing effect on demodex with the different con-centrations volatile oils was observed under microscope. Results The pure volatile oil had high effect in killing D.f and D.b in vitro, and the killing effect to the D.f was better with statistical significance (t=9.172,P<0.05). However, the killing effect showed declining obviously with the increasing of dilutes multiple. Conclusion The pure volatile oil of Manchurian Wildginger has favorable effect on the killing of demodex in vitro.%目的:观察细辛精油体外杀螨效果。方法用细辛精油、1∶2、1∶4、1∶8、1∶16和1∶32稀释的细辛进行抑杀实验,并设定生理盐水为空白对照。采用透明胶带粘贴法获取蠕形螨,镜下观察不同浓度的细辛对两种人体蠕形螨的抑杀作用。结果细辛精油对两种蠕形螨均有明显的抑杀作用,且对毛囊蠕形螨的抑杀效果优于皮质蠕形螨,差异有统计学意义(t=9.172, P<0.05),但随稀释倍数的增加抑杀效果明显降低。结论细辛精油具有良好的体外抑杀蠕形螨的作用。

  6. Effect of platinum nanoparticles on cell death induced by ultrasound in human lymphoma U937 cells.

    Science.gov (United States)

    Jawaid, Paras; Rehman, Mati Ur; Hassan, Mariame Ali; Zhao, Qing Li; Li, Peng; Miyamoto, Yusei; Misawa, Masaki; Ogawa, Ryohei; Shimizu, Tadamichi; Kondo, Takashi

    2016-07-01

    In this study, we report on the potential use of platinum nanoparticles (Pt-NPs), a superoxide dismutase (SOD)/catalase mimetic antioxidant, in combination with 1MHz ultrasound (US) at an intensity of 0.4 W/cm(2), 10% duty factor, 100 Hz PRF, for 2 min. Apoptosis induction was assessed by DNA fragmentation assay, cell cycle analysis and Annexin V-FITC/PI staining. Cell killing was confirmed by cell counting and microscopic examination. The mitochondrial and Ca(2+)-dependent pathways were investigated. Caspase-8 expression and autophagy-related proteins were detected by spectrophotometry and western blot analysis, respectively. Intracellular reactive oxygen species (ROS) elevation was detected by flow cytometry, while extracellular free radical formation was assessed by electron paramagnetic resonance spin trapping spectrometry. The results showed that Pt-NPs exerted differential effects depending on their internalization. Pt-NPs functioned as potent free radical scavengers when added immediately before sonication while pre-treatment with Pt-NPs suppressed the induction of apoptosis as well as autophagy (AP), and resulted in enhanced cell killing. Dead cells displayed the features of pyknosis. The exact mode of cell death is still unclear. In conclusion, the results indicate that US-induced AP may contribute to cell survival post sonication. To our knowledge this is the first study to discuss autophagy as a pro-survival pathway in the context of US. The combination of Pt-NPs and US might be effective in cancer eradication.

  7. Effect of Magnesium on Inclusion Formation in Ti-Killed Steels and Microstructurai Evolution in Welding Induced Coarse-Grained Heat Affected Zone

    Institute of Scientific and Technical Information of China (English)

    CHAI Feng; YANG Cai-fu; SU Hang; ZHANG Yong-quan; XU Zhou

    2009-01-01

    Effects of Mg on the chemical component and size distribution of Ti-bearing inclusions favored grain refinement of the welding induced coarse-grained heat affected zone (CGHAZ),with enhanced impact toughness in Ti-killed steels,which were examined based on experimental observations and thermodynamic calculations.The results indicated that the chemical constituents of the inclusions gradually varied from the Ti-O+Ti-Mg-O compound oxide to the Ti-Mg-O+ MgO compound oxide and the single-phase MgO,as the Mg content increased from 0.002 3% to 0.006%.A trace addition of Mg (approximately 0.002 %) led to the refinement of Ti-bearing inclusions by creating the Ti-Mg-O compound oxide and provided favorable size distribution of the inclusions for acicular ferrite transformation with a high nucleation rate in the CGHAZ,and a high volume fraction of acicular ferrite was obtained in the CGHAZ with enhanced impact toughness.Otherwise,a high content of Mg (approximately 0.006%) produced a single-phase MgO,which was impotent to nucleate an acicular ferrite,and a microstructure comprised of a ferrite side plate and a grain boundary ferrite developed in the CGHAZ.The experimental results were confirmed by thermodynamic calculations.

  8. Selenium-platinum coordination compounds as novel anticancer drugs: selectively killing cancer cells via a reactive oxygen species (ROS)-mediated apoptosis route.

    Science.gov (United States)

    Zeng, Lingwu; Li, Yang; Li, Tianyu; Cao, Wei; Yi, Yu; Geng, Weijia; Sun, Zhiwei; Xu, Huaping

    2014-08-01

    We report the preparation of selenium-containing platinum-based anticancer drug EG-Se/Pt. EG-Se/Pt was obtained from the coordination of selenium-containing molecules (EG-Se) with cisplatin (CDDP). The structure of EG-Se/Pt was characterized by (1) H and (77) Se NMR spectroscopy, XPS, ESI-MS, and MALDI-TOF. In aqueous solution, EG-Se/Pt self-assembles to form spherical aggregates. EG-Se/Pt shows enhanced stability against dilution and high salt concentration compared with EG-Se. EG-Se/Pt induces cell apoptosis via reactive oxygen species (ROS), which leads to high selectivity between cancer cells and normal cells in cytotoxicity assays. More importantly, EG-Se/Pt effectively inhibits tumor growth in vivo in tumor-bearing mice. It is anticipated that tuning the ROS level through the assembly of selenium-containing molecules can be a general method to realize anticancer selectivity.

  9. CDK1 Inhibition Targets the p53-NOXA-MCL1 Axis, Selectively Kills Embryonic Stem Cells, and Prevents Teratoma Formation

    Directory of Open Access Journals (Sweden)

    Noelle E. Huskey

    2015-03-01

    Full Text Available Embryonic stem cells (ESCs have adopted an accelerated cell-cycle program with shortened gap phases and precocious expression of cell-cycle regulatory proteins, including cyclins and cyclin-dependent kinases (CDKs. We examined the effect of CDK inhibition on the pathways regulating proliferation and survival of ESCs. We found that inhibiting cyclin-dependent kinase 1 (CDK1 leads to activation of the DNA damage response, nuclear p53 stabilization, activation of a subset of p53 target genes including NOXA, and negative regulation of the anti-apoptotic protein MCL1 in human and mouse ESCs, but not differentiated cells. We demonstrate that MCL1 is highly expressed in ESCs and loss of MCL1 leads to ESC death. Finally, we show that clinically relevant CDK1 inhibitors prevent formation of ESC-derived tumors and induce necrosis in established ESC-derived tumors. Our data demonstrate that ES cells are uniquely sensitive to CDK1 inhibition via a p53/NOXA/MCL1 pathway.

  10. The effects of active dried and killed dried yeast on subacute ruminal acidosis, ruminal fermentation, and nutrient digestibility in beef heifers.

    Science.gov (United States)

    Vyas, D; Uwizeye, A; Mohammed, R; Yang, W Z; Walker, N D; Beauchemin, K A

    2014-02-01

    The study addressed the importance of yeast (Saccharomyces cerevisiae) viability for reducing the incidence of subacute ruminal acidosis (SARA) and improving total tract nutrient digestibility in beef heifers. Six ruminally cannulated beef heifers (680 ± 50 kg BW) were used in a replicated 3 × 3 Latin square design and were fed a diet consisting of 40% barley silage, 10% chopped grass hay, and 50% barley grain-based concentrate (DM basis). Treatments were 1) no yeast (Control), 2) active dried yeast (ADY; 4 g providing 10(10) cfu/g; AB Vista, Marlborough, UK), and 3) killed dried yeast (KDY; 4 g autoclaved ADY). The treatments were directly dosed via the ruminal cannula daily at the time of feeding. The periods consisted of 2 wk of adaptation (d 1 to 14) and 7 d of measurements (d 15 to 21). Ruminal pH was continuously measured (d 15 to 21) using an indwelling system. Ruminal contents were sampled on d 15 and 17 at 0, 3, 6, 9, and 12 h after feeding. Total tract nutrient digestibility was measured using an external marker (YbCl3) from d 15 to 19. No treatment difference was observed for DMI (P = 0.86). Yeast supplementation (ADY and KDY) tended to increase total tract digestibility of starch (P = 0.07) whereas no effects were observed on digestibility of other nutrients. Both ADY and KDY elevated minimum (P yeast supplementation was effective in reducing time that ruminal pH was below 5.8 (P 0.10); however, the proportion of Ruminococcus flavefaciens in solid fraction of digesta was greater with KDY (P = 0.05). The study demonstrates the positive effects of yeast, irrespective of its viability, in reducing the severity of SARA. However, further studies are required to evaluate the importance of yeast viability for other dietary conditions, particularly when the risk of acidosis is high.

  11. Minimal killing unit of the mitochondrial targeting domain of Noxa.

    Science.gov (United States)

    Kim, Ji-Young; Han, Ji Hye; Moon, Ae-Ran; Park, Jung Hee; Chang, Jeong Hwan; Bae, Jeehyeon; Kim, Tae-Hyoung

    2013-08-01

    Noxa is a key player in p53-induced cell death via mitochondrial dysfunction, and the mitochondrial-targeting domain (MTD) of Noxa is responsible for the translocation of Noxa to mitochondria and for the induction of necrotic cell death. The purpose of this study was to define the minimal killing unit of MTD in vitro and in vivo. It was found that the peptides R8:MTD(10), R8:MTD(9), and R8:MTD(8) can kill various human tumor cells (HCT116, HeLa, MCF-7, BJAB), but that R8:MTD(7) abolishes the killing activity of MTD mainly because of the loss of mitochondrial targeting activity. We find it interesting that R8:MTD(8) was found to kill tumor cells but showed a limited killing activity on normal peritoneal macrophages. Furthermore, R8:MTD(10), R8:MTD(9), and R8:MTD(8) limitedly suppressed tumor growth when injected i.v. into BalB/C mice bearing CT26 cell-derived tumors. These results indicate that MTD(8) is the minimal killing unit of MTD.

  12. MnTnBuOE-2-PyP protects normal colorectal fibroblasts from radiation damage and simultaneously enhances radio/chemotherapeutic killing of colorectal cancer cells

    Science.gov (United States)

    Kosmacek, Elizabeth A.; Chatterjee, Arpita; Tong, Qiang; Lin, Chi; Oberley, Rebecca E.

    2016-01-01

    Manganese porphyrins have been shown to be potent radioprotectors in a variety of cancer models. However, the mechanism as to how these porphyrins protect normal tissues from radiation damage still remains largely unknown. In the current study, we determine the effects of the manganese porphyrin, MnTnBuOE-2-PyP, on primary colorectal fibroblasts exposed to irradiation. We found that 2 Gy of radiation enhances the fibroblasts' ability to contract a collagen matrix, increases cell size and promotes cellular senesence. Treating fibroblasts with MnTnBuOE-2-PyP significantly inhibited radiation-induced collagen contraction, preserved cell morphology and also inhibited cellular senescence. We further showed that MnTnBuOE-2-PyP enhanced the overall viability of the fibroblasts following exposure to radiation but did not protect colorectal cancer cell viability. Specifically, MnTnBuOE-2-PyP in combination with irradiation, caused a significant decrease in tumor clonogenicity. Since locally advanced rectal cancers are treated with chemoradiation therapy followed by surgery and non-metastatic anal cancers are treated with chemoradiation therapy, we also investigated the effects of MnTnBuOE-2-PyP in combination with radiation, 5-fluorouracil with and without Mitomycin C. We found that MnTnBuOE-2-PyP in combination with Mitomycin C or 5-fluorouracil further enhances those compounds' ability to suppress tumor cell growth. When MnTnBuOE-2-PyP was combined with the two chemotherapeutics and radiation, we observed the greatest reduction in tumor cell growth. Therefore, these studies indicate that MnTnBuOE-2-PyP could be used as a potent radioprotector for normal tissue, while at the same time enhancing radiation and chemotherapy treatment for rectal and anal cancers. PMID:27119354

  13. Phantom metrics with Killing spinors

    Directory of Open Access Journals (Sweden)

    W.A. Sabra

    2015-11-01

    Full Text Available We study metric solutions of Einstein–anti-Maxwell theory admitting Killing spinors. The analogue of the IWP metric which admits a space-like Killing vector is found and is expressed in terms of a complex function satisfying the wave equation in flat (2+1-dimensional space–time. As examples, electric and magnetic Kasner spaces are constructed by allowing the solution to depend only on the time coordinate. Euclidean solutions are also presented.

  14. Human neutrophils dump Candida glabrata after intracellular killing.

    Science.gov (United States)

    Essig, Fabian; Hünniger, Kerstin; Dietrich, Stefanie; Figge, Marc Thilo; Kurzai, Oliver

    2015-11-01

    Interaction between fungal pathogens and human phagocytes can lead to remarkably variable outcomes, ranging from intracellular killing to prolonged survival and replication of the pathogen in the host cell. Using live cell imaging we observed primary human neutrophils that release phagocytosed Candida glabrata yeast cells after intracellular killing. This process, for which we propose the name "dumping", adds a new outcome to phagocyte-fungus interaction which may be of potential immunological importance as it allows professional antigen presenting cells to take up and process neutrophil-inactivated pathogens that in their viable state are able to evade intracellular degradation in these cells.

  15. Methionine oxidation contributes to bacterial killing by the myeloperoxidase system of neutrophils.

    Science.gov (United States)

    Rosen, Henry; Klebanoff, Seymour J; Wang, Yi; Brot, Nathan; Heinecke, Jay W; Fu, Xiaoyun

    2009-11-03

    Reactive oxygen intermediates generated by neutrophils kill bacteria and are implicated in inflammatory tissue injury, but precise molecular targets are undefined. We demonstrate that neutrophils use myeloperoxidase (MPO) to convert methionine residues of ingested Escherichia coli to methionine sulfoxide in high yield. Neutrophils deficient in individual components of the MPO system (MPO, H(2)O(2), chloride) exhibited impaired bactericidal activity and impaired capacity to oxidize methionine. HOCl, the principal physiologic product of the MPO system, is a highly efficient oxidant for methionine, and its microbicidal effects were found to correspond linearly with oxidation of methionine residues in bacterial cytosolic and inner membrane proteins. In contrast, outer envelope proteins were initially oxidized without associated microbicidal effect. Disruption of bacterial methionine sulfoxide repair systems rendered E. coli more susceptible to killing by HOCl, whereas over-expression of a repair enzyme, methionine sulfoxide reductase A, rendered them resistant, suggesting a direct role for methionine oxidation in bactericidal activity. Prominent among oxidized bacterial proteins were those engaged in synthesis and translocation of peptides to the cell envelope, an essential physiological function. Moreover, HOCl impaired protein translocation early in the course of bacterial killing. Together, our findings indicate that MPO-mediated methionine oxidation contributes to bacterial killing by neutrophils. The findings further suggest that protein translocation to the cell envelope is one important pathway targeted for damage.

  16. Effect of vaccinating breeder chickens with a killed Salmonella vaccine on Salmonella prevalences and loads in breeder and broiler chicken flocks.

    Science.gov (United States)

    Berghaus, R D; Thayer, S G; Maurer, J J; Hofacre, C L

    2011-05-01

    The objective of this study was to evaluate the effect of vaccination of breeder chickens on Salmonella prevalences and loads in breeder and broiler chicken flocks. Chickens housed on six commercial breeder farms were vaccinated with a killed Salmonella vaccine containing Salmonella Typhimurium, Salmonella Enteritidis, and Salmonella Kentucky. Unvaccinated breeders placed on six additional farms served as controls. Eggs from vaccinated and unvaccinated breeder flocks were kept separately in the hatchery, and the resulting chicks were used to populate 58 commercial broiler flock houses by using a pair-matched design. Vaccinated breeder flocks had significantly higher Salmonella-specific antibody titers than did the unvaccinated breeder flocks, although they did not differ significantly with respect to environmental Salmonella prevalences or loads. Broiler flocks that were the progeny of vaccinated breeders had significantly lower Salmonella prevalences and loads than broiler flocks that were the progeny of unvaccinated breeders. After adjusting for sample type and clustering at the farm level, the odds of detecting Salmonella in samples collected from broiler flocks originating from vaccinated breeders were 62% lower (odds ratio [95% confidence interval] = 0.38 [0.21, 0.68]) than in flocks from unvaccinated breeders. In addition, the mean load of culture-positive samples was lower in broilers from vaccinated breeders by 0.30 log most probable number per sample (95% confidence interval of -0.51, -0.09; P = 0.004), corresponding to a 50% decrease in Salmonella loads. In summary, vaccination of broiler breeder pullets increased humoral immunity in the breeders and reduced Salmonella prevalences and loads in their broiler progeny, but did not significantly decrease Salmonella in the breeder farm environment.

  17. Effects of AlMnCa and AlMnFe Alloys on Deoxidization of Low Carbon and Low Silicon Aluminum Killed Steels

    Institute of Scientific and Technical Information of China (English)

    ZHAN Dong-ping; ZHANG Hui-shu; JIANG Zhou-hua

    2008-01-01

    To confirm the effects of AlMnCa and AIMnFe alloys on the deoxidization and modification of Al2O3 inclu-sions, experiments of 4-heat low carbon and low silicon aluminum killed steels deoxidized by AlMnCa and AlMnFe alloys were done in a MoSi2 furnace at 1 873 K. It is found that the 1# A1MnCa alloy has the best ability of deoxidi-zation and modification of Al2 O3 inclusions than 2# A1MnCa and A1MnFe alloys. Steel A deoxidized by 1# AlMnCa alloy has the lowest total oxygen content in the terminal steel, which is 37 × 10-6. Most of the inclusions in the steel deoxidized by 1# AIMnCa alloy are spherical CaO-containing compound inclusions, and 89. 1% of them are smaller than 10 μm. The diameter of the inclusion bigger than 50 μm is not found in the final steels deoxidized by AlMnCa alloys. Whereas, for the steels deoxidized by AlMnFe alloys, most inclusions in the terminal steel are Al2O3 or Al2O3-MnO inclusions, and a few of them are spherical, and only 76. 8% of them are smaller than 10 μm. Some in-clusions bigger than 50 μm are found in the steel D deoxidized by AlMnFe alloy.

  18. The effect of iron-deficiency anemia on cytolytic activity of mice spleen and peritoneal cells against allogenic tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Kuvibidila, S.R.; Baliga, B.S.; Suskind, R.M.

    1983-08-01

    The capacity of spleen and peritoneal cells from iron deficient mice, ad libitum fed control mice, and pair-fed mice to kill allogenic tumor cells (mastocytoma tumor P815) has been investigated. In the first study, mice were sensitized in vivo with 10(7) viable tumor cells 51 and 56 days after weaning. The capacity of splenic cells and peritoneal cells from sensitized and nonsensitized mice to kill tumor cells was evaluated 5 days after the second dose of tumor cells. At ratios of 2.5:1 to 100:1 of attacker to target cells, the percentage /sup 51/Cr release after 4 h of incubation was significantly less in iron-deficient mice than control and/or pair-fed mice (p less than 0.05). Protein-energy undernutrition in pair-fed mice had no significant effect. In the second study, spleen cells and enriched T cell fractions were incubated in vitro for 5 days with uv irradiated Balb/C spleen cells in a 2:1 ratio. The cytotoxic capacity against the same allogenic tumor cells was again evaluated. The percentage chromium release at different attacker to target cells was less than 30% in the iron-deficient group compared to either control or pair-fed supporting the results of in vivo sensitized cells. The possible mode of impairment of the cytotoxic capacity is discussed.

  19. 等离子体臭氧对水中微生物的杀灭作用%Study on killing effect on microorganisms in water by plasmas ozone

    Institute of Scientific and Technical Information of China (English)

    顾春英; 薛广波; 居喜娟

    2001-01-01

    Objective To study killing effect of plasmas ozo ne water on microorganisms by plasmas ozone disinfection solution through plasma s ozone resulting from discharge along surface disinfection solution produced by machine and its effect factors. Methods Quantitative solut ion test was used to study killing action of plasmas ozone disinfection water to E. coli and staphyrococcus aureus, and to study its eff ect factors. Results Plasmas ozone water could kill 99.9% E. coli and staphyrococcus aureus after 15 min. The kill ing effect was affected by organism. With the increasing of organism concertrat ion, the killing efficacy increased. Conclusions Plasmas ozone can effectively kill microorgnisms in water and the efficacy was affected by org anism.%目的 通过沿面放电等离子体臭氧消毒液发生器 发生的等离子体臭氧消毒水来研究等离子体臭氧对水中微生物的杀灭效果及影响因素。方法 采用悬液定量消毒试验法来研究等离子体臭氧消毒水对金黄色葡萄球菌 和大肠杆菌的杀灭作用,并测试了有机物对其消毒效果的影响。结果 等 离子体臭氧对水中的大肠杆菌和金黄色葡萄球菌作用15 min,杀灭率达99.9%以上,而臭 氧浓度为1.0 mg*L-1,其杀菌作用受有机物的影响,有机物浓度越高对消 毒效果影响越大。结论 等离子体臭氧可以有效杀灭水中的细菌繁殖体, 其杀灭效果受有机物影响。

  20. Human Leukocytes Kill Brugia malayi Microfilariae Independently of DNA-Based Extracellular Trap Release

    Science.gov (United States)

    McCoy, Ciaran J.; Reaves, Barbara J.; Giguère, Steeve; Coates, Ruby; Rada, Balázs

    2017-01-01

    Background Wuchereria bancrofti, Brugia malayi and Brugia timori infect over 100 million people worldwide and are the causative agents of lymphatic filariasis. Some parasite carriers are amicrofilaremic whilst others facilitate mosquito-based disease transmission through blood-circulating microfilariae (Mf). Recent findings, obtained largely from animal model systems, suggest that polymorphonuclear leukocytes (PMNs) contribute to parasitic nematode-directed type 2 immune responses. When exposed to certain pathogens PMNs release extracellular traps (NETs) in the form of chromatin loaded with various antimicrobial molecules and proteases. Principal findings In vitro, PMNs expel large amounts of NETs that capture but do not kill B. malayi Mf. NET morphology was confirmed by fluorescence imaging of worm-NET aggregates labelled with DAPI and antibodies to human neutrophil elastase, myeloperoxidase and citrullinated histone H4. A fluorescent, extracellular DNA release assay was used to quantify and observe Mf induced NETosis over time. Blinded video analyses of PMN-to-worm attachment and worm survival during Mf-leukocyte co-culture demonstrated that DNase treatment eliminates PMN attachment in the absence of serum, autologous serum bolsters both PMN attachment and PMN plus peripheral blood mononuclear cell (PBMC) mediated Mf killing, and serum heat inactivation inhibits both PMN attachment and Mf killing. Despite the effects of heat inactivation, the complement inhibitor compstatin did not impede Mf killing and had little effect on PMN attachment. Both human PMNs and monocytes, but not lymphocytes, are able to kill B. malayi Mf in vitro and NETosis does not significantly contribute to this killing. Leukocytes derived from presumably parasite-naïve U.S. resident donors vary in their ability to kill Mf in vitro, which may reflect the pathological heterogeneity associated with filarial parasitic infections. Conclusions/Significance Human innate immune cells are able to

  1. Cytotoxic effects of curcumin in osteosarcoma cells

    Directory of Open Access Journals (Sweden)

    Moran JM

    2014-11-01

    Full Text Available JM Moran, FJ Rodriguez-Velasco, R Roncero-Martin, V Vera, JD Pedrera-Zamorano Metabolic Bone Diseases Research Group, University of Extremadura, Cáceres, SpainWe read with interest the results of the study from Chang et al1 in the International Journal of Nanomedicine. This article shows that curcumin (diferuloymethane, a phenolic compound extracted from the natural plant Curcuma longa L., exerts higher cytotoxicity in osteosarcoma MG-63 cells than in healthy human osteoblasts. Based on the dosages provided by the authors it is hypothesized that with the appropriate drug carriers, curcumin could be delivered to bone tumors and selectively kill osteosarcoma cells without harming healthy osteoblasts. This hypothesis is based in the data provided that claims the dose needed to exert a significant cytotoxicity on osteosarcoma cells was much lower than that needed to exert the same effect on healthy human osteoblasts. The topic of this study is of importance for both cancer and metabolic bone disease fields, we appreciate the methodology of the study, nevertheless we believe that the hypothesis formulated by the authors and their conclusion deserves further comment.View original paper by Chang and colleagues.

  2. The combination of BH3-mimetic ABT-737 with the alkylating agent temozolomide induces strong synergistic killing of melanoma cells independent of p53.

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    Steven N Reuland

    Full Text Available Metastatic melanoma has poor prognosis and is refractory to most conventional chemotherapies. The alkylating agent temozolomide (TMZ is commonly used in treating melanoma but has a disappointing response rate. Agents that can act cooperatively with TMZ and improve its efficacy are thus highly sought after. The BH3 mimetic ABT-737, which can induce apoptosis by targeting pro-survival Bcl-2 family members, has been found to enhance the efficacy of many conventional chemotherapeutic agents in multiple cancers. We found that combining TMZ and ABT-737 induced strong synergistic apoptosis in multiple human melanoma cell lines. When the drugs were used in combination in a mouse xenograft model, they drastically reduced tumor growth at concentrations where each individual drug had no significant effect. We found that TMZ treatment elevated p53 levels, and that the pro-apoptotic protein Noxa was elevated in TMZ/ABT-737 treated cells. Experiments with shRNA demonstrated that the synergistic effect of TMZ and ABT-737 was largely dependent on Noxa. Experiments with nutlin-3, a p53 inducer, demonstrated that p53 induction was sufficient for synergistic cell death with ABT-737 in a Noxa-dependent fashion. However, p53 was not necessary for TMZ/ABT-737 synergy as demonstrated by a p53-null line, indicating that TMZ and ABT-737 together induce Noxa in a p53-independent fashion. These results demonstrate that targeting anti-apoptotic Bcl-2 members is a promising method for treating metastatic melanoma, and that clinical trials with TMZ and Bcl-2 inhibitors are warranted.

  3. Killing, letting die and euthanasia.

    Science.gov (United States)

    Husak, D N

    1979-12-01

    Medical ethicists debate whether or not the moral assessment of cases of euthanasia should depend on whether the patient is 'killed' or 'allowed to die'. The usual presupposition is that a clear distinction between killing and letting die can be drawn so that this substantive question is not begged. I contend that the categorisation of cases of instances of killing rather than as instances of letting die depends in part on a prior moral assessment of the case. Hence is it trivially rather than substantively true that the distinction has moral significance. But even if a morally neutral (ie non-question begging) distinction could be drawn, its application to the euthanasia controversy is problematic. I illustrate the difficulties of employing this distinction to reach moral conclusions by critically discussing Philippa Foot's recent treatment of euthanasia. I conclude that even if an act of euthanasia is an instance of killing, and there exists a prima facie moral duty not to kill, and no more stringent duty overrides this duty, one still cannot determine such an act to be morally impermissible.

  4. Imaging and radiation effects of gold nanoparticles in tumour cells

    Science.gov (United States)

    McQuaid, Harold N.; Muir, Mark F.; Taggart, Laura E.; McMahon, Stephen J.; Coulter, Jonathan A.; Hyland, Wendy B.; Jain, Suneil; Butterworth, Karl T.; Schettino, Giuseppe; Prise, Kevin M.; Hirst, David G.; Botchway, Stanley W.; Currell, Fred J.

    2016-01-01

    Gold nanoparticle radiosensitization represents a novel technique in enhancement of ionising radiation dose and its effect on biological systems. Variation between theoretical predictions and experimental measurement is significant enough that the mechanism leading to an increase in cell killing and DNA damage is still not clear. We present the first experimental results that take into account both the measured biodistribution of gold nanoparticles at the cellular level and the range of the product electrons responsible for energy deposition. Combining synchrotron-generated monoenergetic X-rays, intracellular gold particle imaging and DNA damage assays, has enabled a DNA damage model to be generated that includes the production of intermediate electrons. We can therefore show for the first time good agreement between the prediction of biological outcomes from both the Local Effect Model and a DNA damage model with experimentally observed cell killing and DNA damage induction via the combination of X-rays and GNPs. However, the requirement of two distinct models as indicated by this mechanistic study, one for short-term DNA damage and another for cell survival, indicates that, at least for nanoparticle enhancement, it is not safe to equate the lethal lesions invoked in the local effect model with DNA damage events.

  5. Adenoviral delivery of pan-caspase inhibitor p35 enhances bystander killing by P450 gene-directed enzyme prodrug therapy using cyclophosphamide+

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    Doloff Joshua C

    2010-09-01

    Full Text Available Abstract Background Cytochrome P450-based suicide gene therapy for cancer using prodrugs such as cyclophosphamide (CPA increases anti-tumor activity, both directly and via a bystander killing mechanism. Bystander cell killing is essential for the clinical success of this treatment strategy, given the difficulty of achieving 100% efficient gene delivery in vivo using current technologies. Previous studies have shown that the pan-caspase inhibitor p35 significantly increases CPA-induced bystander killing by tumor cells that stably express P450 enzyme CYP2B6 (Schwartz et al, (2002 Cancer Res. 62: 6928-37. Methods To further develop this approach, we constructed and characterized a replication-defective adenovirus, Adeno-2B6/p35, which expresses p35 in combination with CYP2B6 and its electron transfer partner, P450 reductase. Results The expression of p35 in Adeno-2B6/p35-infected tumor cells inhibited caspase activation, delaying the death of the CYP2B6 "factory" cells that produce active CPA metabolites, and increased bystander tumor cell killing compared to that achieved in the absence of p35. Tumor cells infected with Adeno-2B6/p35 were readily killed by cisplatin and doxorubicin, indicating that p35 expression is not associated with acquisition of general drug resistance. Finally, p35 did not inhibit viral release when the replication-competent adenovirus ONYX-017 was used as a helper virus to facilitate co-replication and spread of Adeno-2B6/p35 and further increase CPA-induced bystander cell killing. Conclusions The introduction of p35 into gene therapeutic regimens constitutes an effective approach to increase bystander killing by cytochrome P450 gene therapy. This strategy may also be used to enhance other bystander cytotoxic therapies, including those involving the production of tumor cell toxic protein products.

  6. Extracellular Electron Transfer from Aerobic Bacteria to Au-Loaded TiO2 Semiconductor without Light: A New Bacteria-Killing Mechanism Other than Localized Surface Plasmon Resonance or Microbial Fuel Cells.

    Science.gov (United States)

    Wang, Guomin; Feng, Hongqing; Gao, Ang; Hao, Qi; Jin, Weihong; Peng, Xiang; Li, Wan; Wu, Guosong; Chu, Paul K

    2016-09-21

    Titania loaded with noble metal nanoparticles exhibits enhanced photocatalytic killing of bacteria under light illumination due to the localized surface plasmon resonance (LSPR) property. It has been shown recently that loading with Au or Ag can also endow TiO2 with the antibacterial ability in the absence of light. In this work, the antibacterial mechanism of Au-loaded TiO2 nanotubes (Au@TiO2-NT) in the dark environment is studied, and a novel type of extracellular electron transfer (EET) between the bacteria and the surface of the materials is observed to cause bacteria death. Although the EET-induced bacteria current is similar to the LSPR-related photocurrent, the former takes place without light, and no reactive oxygen species (ROS) are produced during the process. The EET is also different from that commonly attributed to microbial fuel cells (MFC) because it is dominated mainly by the materials' surface, but not the bacteria, and the environment is aerobic. EET on the Au@TiO2-NT surface kills Staphylococcus aureus, but if it is combined with special MFC bacteria, the efficiency of MFC may be improved significantly.

  7. In Vitro Photodynamic Effect of Phycocyanin against Breast Cancer Cells

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    Subramaniyan Bharathiraja

    2016-11-01

    Full Text Available C-phycocyanin, a natural blue-colored pigment-protein complex was explored as a novel photosensitizer for use in low-level laser therapy under 625-nm laser illumination. C-phycocyanin produced singlet oxygen radicals and the level of reactive oxygen species (ROS were raised in extended time of treatment. It did not exhibit any visible toxic effect in the absence of light. Under 625-nm laser irradiation, c-phycocyanin generated cytotoxic stress through ROS induction, which killed MDA-MB-231 breast cancer cells depending on concentrations. Different fluorescent staining of laser-treated cells explored apoptotic cell death characteristics like the shrinking of cells, cytoplasmic condensation, nuclei cleavage, and the formation of apoptotic bodies. In conclusion, phycocyanin is a non-toxic fluorescent pigment that can be used in low-level light therapy.

  8. HecA, a member of a class of adhesins produced by diverse pathogenic bacteria, contributes to the attachment, aggregation, epidermal cell killing, and virulence phenotypes of Erwinia chrysanthemi EC16 on Nicotiana clevelandii seedlings.

    Science.gov (United States)

    Rojas, Clemencia M; Ham, Jong Hyun; Deng, Wen-Ling; Doyle, Jeff J; Collmer, Alan

    2002-10-01

    Erwinia chrysanthemi is representative of a broad class of bacterial pathogens that are capable of inducing necrosis in plants. The E. chrysanthemi EC16 hecA gene predicts a 3,850-aa member of the Bordetella pertussis filamentous hemagglutinin family of adhesins. A hecATn7 mutant was reduced in virulence on Nicotiana clevelandii seedlings after inoculation without wounding. Epifluorescence and confocal laser-scanning microscopy observations of hecA and wild-type cells expressing the green fluorescent protein revealed that the mutant is reduced in its ability to attach and then form aggregates on leaves and to cause an aggregate-associated killing of epidermal cells. Cell killing also depended on production of the major pectate lyase isozymes and the type II, but not the type III, secretion pathway in E. chrysanthemi. HecA homologs were found in bacterial pathogens of plants and animals and appear to be unique to pathogens and universal in necrogenic plant pathogens. Phylogenetic comparison of the conserved two-partner secretion domains in the proteins and the 16S rRNA sequences in respective bacteria revealed the two datasets to be fundamentally incongruent, suggesting horizontal acquisition of these genes. Furthermore, hecA and its two homologs in Yersinia pestis had a G+C content that was 10% higher than that of their genomes and similar to that of plant pathogenic Ralstonia, Xylella, and Pseudomonas spp. Our data suggest that filamentous hemagglutinin-like adhesins are broadly important virulence factors in both plant and animal pathogens.

  9. Killing Effects of Ozone on Pathogens in Mineral Water%臭氧对矿泉水中致病菌的杀灭效果

    Institute of Scientific and Technical Information of China (English)

    张永清; 吴清平; 张菊梅; 郭伟鹏; 彭飞艇; 杨秀华

    2014-01-01

    采用悬液定量杀菌和流动杀菌的方法,对铜绿假单胞菌(Pseudomonas aeruginosa)、粪链球菌(Strep-tococcus faecium)和大肠杆菌(Escherichia coli)的臭氧消毒效果进行了研究。结果表明,悬液定量杀菌试验中,臭氧浓度0.36 mg/L,作用10 min后,铜绿假单胞菌存活率小于1.0%;臭氧浓度0.31 mg/L,作用3 min后,粪链球菌存活率小于0.1%;臭氧浓度在0.45 mg/L及以上,作用10 min后,大肠杆菌存活率在2.0%以下。流动杀菌试验中,臭氧浓度0.20~0.60 mg/L,作用3 min后,大肠杆菌、铜绿假单胞菌和粪链球菌的存活率在0.01%以下,即杀灭率在99.99%以上。%Both suspension quantitative germicidal test and flow germicidal test were used to study disinfection effects of ozone on the Pseudomonas aeruginosa, Streptococcus faecium and Escherichia coli. The suspension quantitative germicidal test found that the survival rate of Pseudomonas aeruginosa treated with 0.36 mg/L of ozone for 10 min was less than 1.0%, and that of Streptococcus faecium reacted with 0.31 mg/L of ozone for 10 min was less than 0.1%. When the ozone was more than 0.45 mg/L and the time was 10 min, the survival rates of Escherichia coli was under 2.0%. The results of the flow germicidal test showed that the probability of survivability of Escherichia coli, Pseudomonas aeruginosa and Streptococcus faecium treated with 0.20-0.60 mg/L of ozone for 3 min were under 0.01%, with the killing rates of more than 99.99%.

  10. Effect and Properties of Surface-Modified Copper Doped ZnO Nanoparticles (Cu:ZnO NPs on Killing Curves of Bacterial Pathogens

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    Enayatollah Kalantar

    2015-10-01

    Full Text Available Background: The current study aimed to determine the effect and properties of surface-modified copper doped Cu:ZnO NPs on killing curves of bacterial pathogens.Methods:  Preparation  of  in  situ  surface-modified  copper  doped  ZnOnanoparticles (Cu:ZnO NPs was done according to standard procedure.Assay for antimicrobial activity of Cu:ZnO NPs against bacterial pathogens was carried out based on disc diffusion method. Determination of shelf life, thermal and pH stability of antibacterial activity of Cu:ZnO NPs was done and residual activity was determined against the target cultures.Results: FTIR spectra indicate that the nanomaterials synthesized have higher peak intensity compared with reagent grade ZnO. According to the SEM image the nanoparticles synthesized have different size and heterogeneous morphology.  400  ppm  of  Cu:ZnO  NPs  gave  zones  of  inhibition  with diameters of 9.0 – 16 mm against the target cultures. Amongst the target cultures, Escherchia coli was the most sensitive to the Cu:ZnO NPs inhibition zone diameter 16 mm; whereas, 9 mm wide inhibition zone was obtained against  Staphylococcus  aureus. The  Cu:ZnO  NPs  was  fairly stable  for a period of 60 days at room temperature (RT showing lost of only 20% and30%     antibacterial  activity  as  tested  against  E.  coli  and  S.  aureus, respectively. The Cu:ZnO NPs was quite stable at this pH and temperaturerange tested against both E. coli and S. aureus.Conclusion: Surface-modified copper doped Cu:ZnO NPs have significant potential for their usefulness as antibacterial agents.

  11. Therapeutic relevance of penicillin-induced hypersensitivity of Staphylococcus aureus to killing by polymorphonuclear leukocytes.

    OpenAIRE

    Lam, C; Georgopoulos, A.; Laber, G.; Schütze, E

    1984-01-01

    There is an overwhelming body of evidence that certain Staphylococcus aureus strains become more sensitive to killing by polymorphonuclear leukocytes after their growth in media containing subinhibitory concentrations of penicillin. However, it is not clear to what extent this phenomenon contributes to the curative effect of penicillin in vivo. To explore its therapeutic relevance, we evaluated the interaction of staphylococci pretreated with penicillin in vitro with leukocytes in cell-proof ...

  12. CD8+ Granzyme B+–Mediated Tissue Injury vs. CD4+IFNγ+–Mediated Parasite Killing in Human Cutaneous Leishmaniasis

    Science.gov (United States)

    Santos, Claire da Silva; Boaventura, Viviane; Ribeiro Cardoso, Cristina; Tavares, Natalia; Lordelo, Morgana J; Noronha, Almério; Costa, Jackson; Borges, Valéria M.; de Oliveira, Camila I; Van Weyenbergh, Johan; Barral, Aldina; Barral-Netto, Manoel; Brodskyn, Cláudia Ida

    2013-01-01

    A protective or deleterious role of CD8+T cells in human cutaneous leishmaniasis (CL) has been debated. The present report explores the participation of CD8+T cells in disease pathogenesis as well as in parasite killing. CD8+T cells accumulated in CL lesions as suggested by a higher frequency of CD8+CD45RO+T cells and CD8+CLA+T cells compared with peripheral blood mononuclear cells. Upon Leishmania braziliensis restimulation, most of the CD8+T cells from the lesion expressed cytolytic markers, CD107a and granzyme B. Granzyme B expression in CL lesions positively correlated with lesion size and percentage of TUNEL-positive cells. We also observed a significantly higher percentage of TUNEL-positive cells and granzyme B expression in the biopsies of patients showing a more intense necrotic process. Furthermore, coculture of infected macrophages and CD8+T lymphocytes resulted in the release of granzyme B, and the use of granzyme B inhibitor, as well as z-VAD, Fas:Fc, or anti-IFN-γ, had no effect upon parasite killing. However, coculture of infected macrophages with CD4+T cells strongly increased parasite killing, which was completely reversed by anti-IFN-γ. Our results reveal a dichotomy in human CL: CD8+ granzyme B+T cells mediate tissue injury, whereas CD4+IFN-γ+T cells mediate parasite killing. PMID:23321919

  13. Gas Plasma Effects on Living Cells

    Science.gov (United States)

    Stoffels, E.; Sladek, R. E. J.; Kieft, I. E.

    This paper surveys the research activities at the Eindhoven University of Technology (The Netherlands) in the area of biomedical applications of gas discharge plasmas. A non-thermal atmospheric plasma source (the plasma needle) has been developed, and its interactions with living mammalian cells and bacteria are studied. It is concluded that plasma can efficiently kill bacteria without harming the cells, and also influence the cells without causing cell death (necrosis). In future it will lead to applications like skin (wound) and caries treatment.

  14. Protein chlorination in neutrophil phagosomes and correlation with bacterial killing.

    Science.gov (United States)

    Green, Jessie N; Kettle, Anthony J; Winterbourn, Christine C

    2014-12-01

    Neutrophils ingest and kill bacteria within phagocytic vacuoles. We investigated where they produce hypochlorous acid (HOCl) following phagocytosis by measuring conversion of protein tyrosine residues to 3-chlorotyrosine. We also examined how varying chloride availability affects the relationship between HOCl formation in the phagosome and bacterial killing. Phagosomal proteins, isolated following ingestion of opsonized magnetic beads, contained 11.4 Cl-Tyr per thousand tyrosine residues. This was 12 times higher than the level in proteins from the rest of the neutrophil and ~6 times higher than previously recorded for protein from ingested bacteria. These results indicate that HOCl production is largely localized to the phagosomes and a substantial proportion reacts with phagosomal protein before reaching the microbe. This will in part detoxify the oxidant but should also form chloramines which could contribute to the killing mechanism. Neutrophils were either suspended in chloride-free gluconate buffer or pretreated with formyl-Met-Leu-Phe, a procedure that has been reported to deplete intracellular chloride. These treatments, alone or in combination, decreased both chlorination in phagosomes and killing of Staphylococcus aureus by up to 50%. There was a strong positive correlation between the two effects. Killing was predominantly oxidant and myeloperoxidase dependent (88% inhibition by diphenylene iodonium and 78% by azide). These resul