RESUMEN
Although radiation related research has been conducted extensively, the molecular toxicology and cellular mechanisms affected by proton radiation remain poorly understood. We recently reported that the high energy protons induce cell death through activation of apoptotic signaling genes; caspase 3 and 8 (Baluchamy et al. J Biol Chem 285:24769-24774, 2010). In this study, we investigated the effect of different doses of protons in in vivo mouse system, particularly, brain tissues. A significant dose-dependent induction of reactive oxygen species and lipid peroxidation and reduction of antioxidants; glutathione and superoxide dismutase were observed in proton irradiated mouse brain as compared to control brain. Furthermore, histopathology studies on proton irradiated mouse brain showed significant tissue damage as compared to control brain. Together, our in vitro and in vivo results suggest that proton irradiation alters oxidant and antioxidant levels in the cells to cause proton mediated DNA/tissue damage followed by apoptotic cell death.
Asunto(s)
Encéfalo/efectos de la radiación , Protones , Traumatismos por Radiación/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Animales , Antioxidantes/metabolismo , Encéfalo/metabolismo , Encéfalo/patología , Glutatión/metabolismo , Peroxidación de Lípido/efectos de la radiación , Masculino , Ratones , Ratones Endogámicos BALB C , Protones/efectos adversos , Superóxido Dismutasa/metabolismo , Superóxido Dismutasa-1RESUMEN
There is an ongoing concern regarding the biocompatibility of nanoparticles with sizes less than 100 nm as compared to larger particles of the same nominal substance. In this study, we investigated the toxic properties of magnetite stabilized with polyacrylate sodium. The magnetite was characterized by X-ray powder diffraction analysis, and the mean particle diameter was calculated using the Scherrer formula and was found to be 9.3 nm. In this study, we treated lung epithelial cells with different concentrations of magnetite and investigated their effects on oxidative stress and cell proliferation. Our data showed an inhibition of cell proliferation in magnetite-treated cells with a significant dose-dependent activation and induction of reactive oxygen species. Also, we observed a depletion of antioxidants, glutathione, and superoxide dismutase, respectively, as compared with control cells. In addition, apoptotic-related protease/enzyme such as caspase-3 and -8 activities, were increased in a dose-dependent manner with corresponding increased levels of DNA fragmentation in magnetite-treated cells compared to than control cells. Together, the present study reveals that magnetite exposure induces oxidative stress and depletes antioxidant levels in the cells to stimulate apoptotic pathway for cell death.
Asunto(s)
Apoptosis/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Pulmón/efectos de los fármacos , Nanopartículas de Magnetita/toxicidad , Estrés Oxidativo/efectos de los fármacos , Animales , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Línea Celular , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Epiteliales/metabolismo , Células Epiteliales/patología , Glutatión/metabolismo , Peroxidación de Lípido/efectos de los fármacos , Pulmón/metabolismo , Pulmón/patología , Tamaño de la Partícula , Difracción de Polvo , Ratas , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa/metabolismo , Difracción de Rayos XRESUMEN
The synthesis of nanoparticles and their functionalization to effectively utilize them in biological applications including drug delivery is currently a challenge. Calcium carbonate among many other inorganic nanosized particles offers promising results for such applications. We have synthesized calcium carbonate nanoparticles using polymer mediated growth technique, where one of the ions bound within polymer matrix and the other diffuses and reacts to form desired compound. The synthesized nanoparticles are characterized using X-ray diffraction, Scanning Electron Microscopy and spectroscopic techniques such as Fourier-Transform Infra-red spectroscopy and UV-Vis spectroscopy. The diameter of the calcium carbonate nanoparticles is estimated to be 39.8 nm and their biocompatibility studies showed no significant induction of oxidative stress or cell death even at higher concentrations (50 microg) upon exposure to HeLa and LE cells. Here, we report that the synthesized calcium carbonate nanosized particles using polymer mediated growth technique are biocompatible and can be safely used for biomedical applications.
Asunto(s)
Materiales Biocompatibles , Carbonato de Calcio/síntesis química , Carbonato de Calcio/química , Células HeLa , Humanos , Microscopía Electrónica de Rastreo , Espectrofotometría Ultravioleta , Espectroscopía Infrarroja por Transformada de Fourier , Difracción de Rayos XRESUMEN
Here, we have synthesized Zinc Oxide (ZnO) nanorods at room temperature using zinc acetate and hexamethylenetetramine as precursors followed by characterization using X-ray diffraction (XRD), fourier transform infra red spectroscopy, scanning electron microscopy (SEM) and transmission electron microscopy. The growth of the synthesized ZnO was found to be very close to its hexagonal nature, which is confirmed by XRD. The nanorods were grown perpendicular to the long-axis and grew along the [001] direction, which is the nature of ZnO growth. The morphology of the synthesized ZnO nanorods was also confirmed by SEM. The size of the nanorod was estimated to be around 20-25 nm in diameter and approximately 50-60 nm in length. Our biocompatibility studies using synthesized ZnO showed no significant dose- or time-dependent increase in the formation of free radicals, accumulation of peroxidative products, antioxidant depletion or loss of cell viability on lung epithelial cells.
Asunto(s)
Nanopartículas/efectos adversos , Nanopartículas/química , Óxido de Zinc/efectos adversos , Óxido de Zinc/química , Animales , Antioxidantes/metabolismo , Materiales Biocompatibles/efectos adversos , Materiales Biocompatibles/química , Línea Celular , Supervivencia Celular , Células Epiteliales/efectos de los fármacos , Glutatión/metabolismo , Microscopía Electrónica de Rastreo , Estrés Oxidativo/efectos de los fármacos , RatasRESUMEN
With the widespread application of carbon nanotubes (CNTs) in diverse commercial processes, scientists are now concerned about the potential health risk of occupational exposures. In this study, CNT-induced pulmonary toxicity was investigated by exposing BALB/c mice to aerosolized single-wall (SW) CNT and multiwall (MW) CNT (5 µg/g of mice) for 7 consecutive days in a nose-only exposure system. Microscopic studies showed that inhaled CNTs were homogeneously distributed in the mouse lung. The total number of bronchoalveolar lavage polymorphonuclear leukocytes recovered from the mice exposed to SWCNT and MWCNT (1.2 × 10(6) ± 0.52 and 9.87 × 10(5) ± 1.45; respectively) was significantly greater than control mice (5.46 × 10(5) ± 0.78). Rapid development of pulmonary fibrosis in mice that inhaled CNT was also confirmed by significant increases in the collagen level. The lactate dehydrogenase levels were increased nearly 2- and 2.4-fold in mice that inhaled SWCNT and MWCNT, respectively, as compared with control mice. In addition, exposure of CNTs to mice showed a significant (p < 0.05) reduction of antioxidants (glutathione, superoxide dismutase, and catalase) and induction of oxidants (myloperoxidase, oxidative stress, and lipid peroxidation) compared with control. Apoptosis-related proteins such as caspase-3 and -8 activities were also significantly increased in mice that inhaled CNT than in control mice. Together, this study shows that inhaled CNTs induce inflammation, fibrosis, alteration of oxidant and antioxidant levels, and induction of apoptosis-related proteins in the lung tissues to trigger cell death.
Asunto(s)
Pulmón/metabolismo , Ensayo de Materiales , Nanotubos de Carbono/efectos adversos , Neumonía/inducido químicamente , Neumonía/metabolismo , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/metabolismo , Aerosoles , Animales , Antioxidantes/metabolismo , Apoptosis/efectos de los fármacos , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Pulmón/patología , Ratones , Ratones Endogámicos BALB C , Estrés Oxidativo/efectos de los fármacos , Oxidorreductasas/metabolismo , Neumonía/patología , Fibrosis Pulmonar/patologíaRESUMEN
Our previous report on multiwall carbon nanotubes (MWCNT) has demonstrated the generation of reactive radicals and depletion of intracellular antioxidants which in turn cause cell death through activation of caspases. The molecular mechanism of cellular death due to MWCNT is not clear yet. In this study, we investigated the signaling pathways implicated in MWCNT-induced apoptosis in rat lung epithelial cells. First, we assessed the DNA damage in response to MWCNT treatment and showed the significant DNA damage as compared to control. The collapse of the mitochondrial membrane integrity, release of cytochrome c into the cytosol, reduction in cellular ATP content, increased levels of mitochondrial apoptogenic factor and activation and nuclear translocation of NF-κB were observed in MWCNT treated cells. In addition, a time-dependent induction of phosphorylated IκBα and its degradation were detected in cells exposed to MWCNT. Furthermore, MWCNT activated several death related proteins including apoptosis inducing factor, p53, p21 and bax. Together, our results suggest that signaling pathways such as NF-κB and AP-1 are activated upon MWCNT treatment for cellular cytotoxicity.
Asunto(s)
Adenosina Trifosfato/análisis , Factor Inductor de la Apoptosis/metabolismo , Apoptosis , Citocromos c/análisis , Daño del ADN/efectos de los fármacos , Daño del ADN/fisiología , Complejo IV de Transporte de Electrones/metabolismo , Activación Enzimática/efectos de los fármacos , Activación Enzimática/fisiología , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , FN-kappa B/metabolismo , Nanotubos de Carbono , Mucosa Respiratoria/efectos de los fármacos , Mucosa Respiratoria/fisiología , Factor de Transcripción AP-1/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Línea Celular , Pulmón/fisiología , Potencial de la Membrana Mitocondrial/fisiología , Nanotecnología , Nanotubos de Carbono/toxicidad , Ratas , Mucosa Respiratoria/ultraestructura , Transducción de SeñalRESUMEN
Radiation is known to potentially interfere with cellular functions at all levels of cell organization. The radiation-induced stress response is very complex and involves altered expression of many genes. Identification of specific genes may allow the determination of pathways important in radiation responses. Although several radiation-related research have been studied extensively, the molecular and cellular processes affected by proton exposure remain poorly understood. Our earlier reports have shown that proton radiation induces reactive oxygen species (ROS) formation and lipid peroxidation and inhibits antioxidants, superoxide dismutase, and glutathione. Therefore, in this present study, we used quantitative real-time reverse transcription polymerase chain reaction approach and showed the modulation of several genes including oxidative stress, antioxidants defense mechanism, ROS metabolism, and oxygen transporters related genes expression in 2-Gy proton-exposed mouse brain. Literature evidences suggest that change in oxidants and antioxidants levels induce DNA damage, followed by cell death. In conclusion, changes in the gene profile of mouse brain after proton irradiation are complex and the exposed cells might undergo programmed cell death through alteration of genes responsible for oxidative stress signaling mechanism.
Asunto(s)
Encéfalo/efectos de la radiación , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de la radiación , Estrés Oxidativo/genética , Protones , Animales , Antioxidantes/metabolismo , Apoptosis/genética , Encéfalo/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Radiation affects several cellular and molecular processes, including double strand breakage and modifications of sugar moieties and bases. In outer space, protons are the primary radiation source that poses a range of potential health risks to astronauts. On the other hand, the use of proton irradiation for tumor radiation therapy is increasing, as it largely spares healthy tissues while killing tumor tissues. Although radiation-related research has been conducted extensively, the molecular toxicology and cellular mechanisms affected by proton irradiation remain poorly understood. Therefore, in this study, we irradiated rat lung epithelial cells with different doses of protons and investigated their effects on cell proliferation and death. Our data show an inhibition of cell proliferation in proton-irradiated cells with a significant dose-dependent activation and repression of reactive oxygen species and antioxidants glutathione and superoxide dismutase, respectively, compared with control cells. In addition, the activities of apoptosis-related genes such as caspase-3 and -8 were induced in a dose-dependent manner with corresponding increased levels of DNA fragmentation in proton-irradiated cells compared with control cells. Together, our results show that proton irradiation alters oxidant and antioxidant levels in cells to activate the apoptotic pathway for cell death.
Asunto(s)
Antioxidantes/química , Células Epiteliales/citología , Pulmón/citología , Oxidantes/química , Animales , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Muerte Celular , Supervivencia Celular , Fragmentación del ADN , Relación Dosis-Respuesta a Droga , Glutatión/química , Protones , Ratas , Especies Reactivas de OxígenoRESUMEN
Exposure of living systems to radiation results in a wide assortment of lesions, the most significant of is damage to genomic DNA which alter specific cell functions including cell proliferation. The radiation induced DNA damage investigation is one of the important area in biology, but still the information available regarding the effects of proton is very limited. In this report, we investigated the differential gene expression pattern of DNA damage signaling genes such as damaged DNA binding, repair, cell cycle arrest, checkpoints and apoptosis using quantitative real-time RT-PCR in proton exposed mouse brain tissues. The expression profiles showed significant changes in DNA damage related genes in 2 Gy proton exposed mouse brain tissues as compared to control brain tissues. Furthermore, we also show that significantly increased levels of apoptotic related genes, caspase-3 and 8 activities in these cells, suggesting that in addition to differential expression of DNA damage genes, the alteration of apoptosis related genes may also contribute to the radiation induced DNA damage followed by programmed cell death. In summary, our findings suggest that proton exposed cells undergo severe DNA damage which in turn destabilize the chromatin stability.
Asunto(s)
Apoptosis/genética , Encéfalo/efectos de la radiación , Daño del ADN/genética , Perfilación de la Expresión Génica , Animales , Apoptosis/efectos de la radiación , Daño del ADN/efectos de la radiación , Fragmentación del ADN , Relación Dosis-Respuesta en la Radiación , Masculino , Ratones , Ratones Endogámicos BALB C , Protones/efectos adversos , Transducción de Señal/genéticaRESUMEN
Carbon nanotubes (CNTs), the most promising material with unique characteristics, find its application in different fields ranging from composite materials to medicine and from electronics to energy storage. However, little is known about the mechanism behind the interaction of these particles with cells and their toxicity. So, here we investigated the adverse effects of multiwalled CNTs (MWCNTs) in rat lung epithelial (LE) cells. The results showed that the incubation of LE cells with 0.5-10 microg/mL of MWCNTs caused a dose- and time-dependent increase in the formation of free radicals, the accumulation of peroxidative products, the loss of cell viability, and antioxidant depletion. The significant amount of incorporation of dUTPs in the nucleus after 24 h confirms the induction of apoptosis. It was also observed that there is an increase in the activity of both caspases-3 and caspase-8 in cells, with increases in time and the concentration of MWCNTs. No significant incorporation of dUTPs was observed in cells, incubated with z-VAD-fmk, which confirmed the role of caspases in DNA fragmentation. The present study reveals that MWCNTs induced oxidative stress and stimulated apoptosis signaling pathway through caspase activation in rat LE cell lines.
Asunto(s)
Apoptosis/efectos de los fármacos , Pulmón/citología , Nanotubos de Carbono/toxicidad , Animales , Antioxidantes/metabolismo , Apoptosis/fisiología , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Línea Celular , Núcleo Celular/metabolismo , Supervivencia Celular/efectos de los fármacos , Fragmentación del ADN/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Activación Enzimática/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Formazáns/metabolismo , Radicales Libres/metabolismo , Glutatión/análisis , Glutatión/metabolismo , Etiquetado Corte-Fin in Situ , Peroxidación de Lípido/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Ratas , Especies Reactivas de Oxígeno/análisis , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa/metabolismo , Sales de Tetrazolio/metabolismo , Factores de TiempoRESUMEN
Uranium is a naturally occurring radioactive material present everywhere in the environment. It is toxic because of its chemical or radioactive properties. Uranium enters environment mainly from mines and industry and cause threat to human health by accumulating in lungs as a result of inhalation. In our previous study, we have shown the effectiveness of antioxidant system response to the oxidative stress induced by uranyl acetate (UA) in rat lung epithelial (LE) cells. As part of our continuing studies; here, we investigated the mechanism underlying when LE cells are exposed to different concentration of UA. Oxidative stress may lead to apoptotic signaling pathways. LE cells treated with 0.25, 0.5 and 1 mM of UA results in dose and time-dependent increase in activity of both caspases-3 and -8. Increase in the concentration of cytochrome-c oxidase in cytosol was seen in LE cells treated with 1 mM UA as a result of mitochondria membrane permeability. The cytochrome-c leakage may trigger the apoptotic pathway. TUNEL assay performed in LE cells treated with 1 mM of UA showed significant incorporation of dNTPs in the nucleus after 24 h. In the presence of the caspase inhibitors, we observed the significant decrease in the activity of caspases-8 and -3 in 0.5 and 1 mM UA-treated LE cells.
Asunto(s)
Apoptosis/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Compuestos Organometálicos/toxicidad , Animales , Caspasa 3/efectos de los fármacos , Caspasa 3/metabolismo , Caspasa 8/efectos de los fármacos , Caspasa 8/metabolismo , Línea Celular , Relación Dosis-Respuesta a Droga , Complejo IV de Transporte de Electrones/metabolismo , Células Epiteliales/metabolismo , Etiquetado Corte-Fin in Situ , Pulmón/citología , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Membranas Mitocondriales/efectos de los fármacos , Membranas Mitocondriales/metabolismo , Compuestos Organometálicos/administración & dosificación , Estrés Oxidativo/efectos de los fármacos , Permeabilidad , Ratas , Transducción de Señal/efectos de los fármacos , Factores de TiempoRESUMEN
Microgravity is known to have significant effect on all aspects of reproductive function in animal models. Recent studies have also shown that microgravity induces changes at the cellular level, including apoptosis. Our effort here was to study the effect of simulated microgravity on caspase-8 and the caspase-3 activities, the effectors of the apoptotic pathway and on the transcription factor NF-kappaB a signaling molecule in mouse testis. Morey-Holton hind limb suspension model was used to simulate microgravity. Caspase-8 and 3 fluorometric assays were carried out and HLS mice testis exhibited a 51% increase in caspase-8 and caspase-3 compared to the controls. A sandwich ELISA-based immunoassay was carried out for detection of NF-kappaB which again significantly increased in the test mice. Testosterone levels were measured using an ELISA kit and in HLS mice the decrease was significant. There was also a significant decrease in testis weight in the test mice. Simulated microgravity activates caspase 8, 3 and NF-kappaB necessary to stimulate the apoptotic pathway in mice testis. This may account for the drop in testis weight and testosterone level further affecting testicular physiology and function.