ABSTRACT
Transplant recipients face an increased risk of cancer compared with the healthy population. Although several studies have examined the direct effects of immunosuppressive drugs on cancer cells, little is known about the interactions between pharmacological immunosuppression and cancer immunosurveillance. We investigated the different effects of rapamycin (Rapa) versus cyclosporine A (CsA) on tumor-reactive CD8(+) T cells. After adoptive transfer of CD8(+) T cell receptor-transgenic OTI T cells, recipient mice received either skin grafts expressing ovalbumin (OVA) or OVA-expressing B16F10 melanoma cells. Animals were treated daily with Rapa or CsA. Skin graft rejection and tumor growth as well as molecular and cellular analyses of skin- and tumor-infiltrating lymphocytes were performed. Both Rapa and CsA were equally efficient in prolonging skin graft survival when applied at clinically relevant doses. In contrast to Rapa-treated animals, CsA led to accelerated tumor growth in the presence of adoptively transferred tumor-reactive CD8(+) OTI T cells. Further analyses showed that T-bet was downregulated by CsA (but not Rapa) in CD8(+) T cells and that cancer cytotoxicity was profoundly inhibited in the absence of T-bet. CsA reduces T-bet-dependent cancer immunosurveillance by CD8(+) T cells. This may contribute to the increased cancer risk in transplant recipients receiving calcineurin inhibitors.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Cyclosporine/pharmacology , Graft Rejection/immunology , Immune Tolerance/immunology , Melanoma, Experimental/immunology , Skin Transplantation , T-Box Domain Proteins/physiology , Animals , Female , Graft Rejection/drug therapy , Graft Survival/drug effects , Graft Survival/immunology , Immune Tolerance/drug effects , Immunosuppressive Agents/pharmacology , Male , Melanoma, Experimental/drug therapy , Melanoma, Experimental/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Sirolimus/pharmacologyABSTRACT
The increased inhaled application of titanium dioxide nanoparticles (TiO(2) NPs) increases the potential pulmonary health risks. The present investigations were carried out to study the TiO(2) NPs-induced apoptosis, oxidative stress and genotoxicity in the human lung cancer cell line, A549, a widely used cell system for pulmonary toxicity studies. Tetrazolium bromide salt and lactate dehydrogenase release assays were used to study the cytotoxicity. The genotoxicity studies were carried out using cytokinesis block micronucleus assay. Apoptosis was confirmed by the formation of apoptotic bodies and altered expression (messenger RNA (mRNA) and protein) of markers such as P(53), P(21), Bax, Bcl(2) and cleaved caspase-3. Cells exposed to TiO(2) NPs (10 and 50 µg/ml) for 6-24 h shows significant induction in oxidative stress, that is, the production of reactive oxygen species and malondialdehyde and decrease in the activity of catalase and glutathione. TiO(2) NPs exposure also induces the formation of apoptotic bodies and micronucleus as marker of genotoxicity. A significant up-regulation in the expression of apoptosis markers such as P(53), P(21) and cleaved caspase-3 was observed, while the levels were down-regulated for Bcl(2) at both mRNA and protein levels. TiO(2) NPs exposure could not pose significant effects on Bax expression. Data indicate that nano-TiO(2) induces oxidative stress, genotoxicity and apoptosis in human lung cancer cell line, A549. Our result also identifies the mechanisms involved in TiO(2) NP-induced changes in A549 cells. Perhaps, reporting for the first time, the association of TiO(2) NPs-induced genotoxicity and apoptosis at transcriptional and translational level in the human lung cancer cell line, A549 cells.
Subject(s)
Mutagens/toxicity , Nanoparticles/toxicity , Titanium/toxicity , Apoptosis , Caspase 3/genetics , Caspase 3/metabolism , Catalase/metabolism , Cell Line, Tumor , Cyclin-Dependent Kinase Inhibitor p21/genetics , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Glutathione/metabolism , Humans , Lipid Peroxidation , Micronucleus Tests , Oxidative Stress , Reactive Oxygen Species/metabolism , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , bcl-2-Associated X Protein/genetics , bcl-2-Associated X Protein/metabolismABSTRACT
The possible harmful effects of radiofrequency electromagnetic fields (RF EMFs) are controversial. We have used human Mono Mac 6 cells to investigate the influence of RF EMFs in vitro on cell cycle alterations and BrdU uptake, as well as the induction of apoptosis and necrosis in human Mono Mac 6 cells, using flow cytometry after exposure to a 1,800 MHz, 2 W/kg specific absorption rate (SAR), GSM-DTX signal for 12 h. No statistically significant differences in the induction of apoptosis or necrosis, cell cycle kinetics, or BrdU uptake were detected after RF EMF exposure compared to sham or incubator controls. However, in the positive control cells treated with gliotoxin and PMA (phorbol 12 myristate-13 acetate), a significant increase in apoptotic and necrotic cells was seen. Cell cycle analysis or BrdU incorporation for 72 h showed no differences between RF EMF- or sham-exposed cells, whereas PMA treatment induced a significant accumulation of cells in G(0)/G(1)-phase and a reduction in S-phase cells. RF EMF radiation did not induce cell cycle alterations or changes in BrdU incorporation or induce apoptosis and necrosis in Mono Mac 6 cells under the exposure conditions used.
Subject(s)
Apoptosis/radiation effects , Cell Cycle/radiation effects , Microwaves/adverse effects , Monocytes/pathology , Monocytes/radiation effects , Necrosis/etiology , Necrosis/pathology , Cell Line, Tumor , Dose-Response Relationship, Radiation , Environmental Exposure/adverse effects , Humans , Kinetics , Radiation Dosage , Radio Waves/adverse effectsABSTRACT
The aim of this study is to investigate if 1,800 MHz radiofrequency electromagnetic fields (RF-EMF) can induce reactive oxygen species (ROS) release and/or changes in heat shock protein 70 (Hsp70) expression in human blood cells, using different exposure and co-exposure conditions. Human umbilical cord blood-derived monocytes and lymphocytes were used to examine ROS release after exposure to continuous wave or different GSM signals (GSM-DTX and GSM-Talk) at 2 W/kg for 30 or 45 min of continuous or intermittent (5 min ON/5 min OFF) exposure. The cells were exposed to incubator conditions, to sham, to RF-EMF, or to chemicals in parallel. Cell stimulation with the phorbol ester phorbol-12-myristate-13-acetate (PMA; 1 microM) was used as positive control for ROS release. To investigate the effects on Hsp70 expression, the human monocytes were exposed to the GSM-DTX signal at 2 W/kg for 45 min, or to heat treatment (42 degrees C) as positive control. ROS production and Hsp70 expression were determined by flow cytometric analysis. The data were compared to sham and/or to control values and the statistical analysis was performed by the Student's t-test (P<0.05). The PMA treatment induced a significant increase in ROS production in human monocytes and lymphocytes when the data were compared to sham or to incubator controls. After continuous or intermittent GSM-DTX signal exposure (2 W/kg), a significantly different ROS production was detected in human monocytes if the data were compared to sham. However, this significant difference appeared due to the lowered value of ROS release during sham exposure. In human lymphocytes, no differences could be detected if data were compared either to sham or to incubator control. The Hsp70 expression level after 0, 1, and 2 h post-exposure to GSM-DTX signal at 2 W/kg for 1 h did not show any differences compared to the incubator or to sham control.
Subject(s)
Electromagnetic Fields , HSP70 Heat-Shock Proteins/physiology , Monocytes/radiation effects , Reactive Oxygen Species , Antigens, CD/biosynthesis , Antigens, Differentiation, T-Lymphocyte/biosynthesis , Dose-Response Relationship, Radiation , Flow Cytometry , Humans , Immunophenotyping , Lectins, C-Type , Lymphocytes/metabolism , Monocytes/metabolism , Radio Waves , Tetradecanoylphorbol Acetate/pharmacology , Umbilical Cord/radiation effects , Umbilical Veins/cytologyABSTRACT
The goal of this study was to investigate whether radiofrequency (RF) electromagnetic-field (EMF) exposure at 1800 MHz causes production of free radicals and/or expression of heat-shock proteins (HSP70) in human immune-relevant cell systems. Human Mono Mac 6 and K562 cells were used to examine free radical release after exposure to incubator control, sham, RF EMFs, PMA, LPS, heat (40 degrees C) or co-exposure conditions. Several signals were used: continuous-wave, several typical modulations of the Global System for Mobile Communications (GSM): GSM-non DTX (speaking only), GSM-DTX (hearing only), GSM-Talk (34% speaking and 66% hearing) at specific absorption rates (SARs) of 0.5, 1.0, 1.5 and 2.0 W/kg. Heat and PMA treatment induced a significant increase in superoxide radical anions and in ROS production in the Mono Mac 6 cells when compared to sham and/or incubator conditions. No significant differences in free radical production were detected after RF EMF exposure or in the respective controls, and no additional effects on superoxide radical anion production were detected after co-exposure to RF EMFs+PMA or RF EMFs+LPS. The GSM-DTX signal at 2 W/kg produced a significant difference in free radical production when the data were compared to sham because of the decreasing sham value. This difference disappeared when data were compared to the incubator controls. To determine the involvement of heat-shock proteins as a possible inhibitor of free radical production, we investigated the HSP70 expression level after different RF EMF exposures; no significant effects were detected.
Subject(s)
Free Radicals/metabolism , HSP70 Heat-Shock Proteins/immunology , Microwaves , Monocytes/immunology , Monocytes/radiation effects , Radio Waves , Cell Line , Dose-Response Relationship, Radiation , Environmental Exposure , Free Radicals/immunology , Gene Expression Regulation/immunology , Gene Expression Regulation/radiation effects , HSP70 Heat-Shock Proteins/metabolism , Humans , K562 Cells , Radiation DosageABSTRACT
The contemporary urban environment has become increasingly complex in its composition, leading to discussions regarding possible novel health effects. Two factors that recently have received considerable attention are ultrafine particles (UFP; <0.1 microm) produced by combustion processes and emissions from wireless communication devices like mobile phones that emit in the radio-frequency (RF) part of the spectrum. Several studies have shown biological effects of both these exposures in various cell systems. Here we investigate if exposure to UFP (12-14 nm, 100 microg/ml) and RF-electromagnetic fields (EMF; 2 W/kg specific absorption rate (SAR); continuous wave (CW) or modulated (217Hz or GSM-nonDTX)), alone or in combination influences levels of the superoxide radical anion or the stress protein heat-shock protein (Hsp70) in the human monocyte cell line Mono Mac 6. Heat treatment (42-43 degrees C, 1h) was used as positive control for both stress reaction and for heat development in the RF exposure setup. Our results clearly show that Mono Mac 6 cells are capable to internalise UFP, and that this phagocytic activity is connected to an increased release of free radicals. This increase (40-45% above negative control) is stronger than the effect of heat treatment. On the other hand, none of the employed RF exposures showed any effects on free radical levels. Co-exposure of RF and UFP did not potentiate the UFP effect either. Our investigations showed a significantly increased Hsp70 expression level by heat treatment in a time-dependent manner, whereas UFP, RF, or UFP+RF were without any effect. Therefore, we conclude that in the investigated Mono Mac 6 cells, RF exposure alone or in combination with UFP cannot influence stress-related responses.