RESUMO
BACKGROUND: Glioma is the most prevalent malignant tumor in human central nervous systems. Recently, the development of resistance to radiotherapy in glioma patients markedly vitiates the therapy outcome. MiR-153-3p has been reported to be closely correlated with tumor progression, but its effect and molecular mechanism underlying radioresistance remains unclear in glioma. METHODS: The expression of miR-153-3p was determined in radioresistant glioma clinical specimens as well as glioma cell lines exposed to irradiation (IR) using quantitative real-time PCR. Cell viability, proliferation and apoptosis were then evaluated by MTT assay, colony formation assay, Flow cytometry analysis and caspase-3 activity assay in glioma cells (U87 and U251). Tumor forming was evaluated by nude mice model in vivo. TUNEL staining was used to detect cell apoptosis in nude mice model. The target genes of miR-153-3p were predicted and validated using integrated bioinformatics analysis and a luciferase reporter assay. RESULTS: Here, we found that miR-153-3p was down-regulated in radioresistant glioma clinical specimens as well as glioma cell lines (U87 and U251) exposed to IR. Enhanced expression of miR-153-3p promoted the radiosensitivity, promoted apoptosis and elevated caspase-3 activity in glioma cells in vitro, as well as the radiosensitivity in U251 cell mouse xenografs in vivo. Mechanically, B cell lymphoma-2 gene (BCL2) was identified as the direct and functional target of miR-153-3p. Moreover, restoration of BCL2 expression reversed miR-153-3p-induced increase of radiosensitivity, apoptosis and caspase-3 activity in U251 cells in vitro. In addition, clinical data indicated that the expression of miR-153-3p was significantly negatively associated with BCL2 in radioresistance of glioma samples. CONCLUSIONS: Our findings suggest that miR-153-3p is a potential target to enhance the effect of radiosensitivity on glioma cells, thus representing a new potential therapeutic target for glioma.
Assuntos
Genes bcl-2/fisiologia , Glioma/genética , MicroRNAs/fisiologia , MicroRNAs/efeitos da radiação , Tolerância a Radiação/genética , Adulto , Análise de Variância , Western Blotting , Caspase 3/análise , Linhagem Celular Tumoral , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Regulação para Baixo , Feminino , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica , Marcação de Genes/métodos , Genes bcl-2/efeitos da radiação , Glioma/radioterapia , Humanos , Marcação In Situ das Extremidades Cortadas , Masculino , MicroRNAs/análise , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Fatores de TempoRESUMO
Acute respiratory distress syndrome (ARDS) and acute lung injury (ALI) are defined as pulmonary inflammation that could occur from sepsis and lead to pulmonary permeability and alveolar edema making them life-threatening diseases. Photobiomodulation (PBM) properties have been widely described in the literature in several inflammatory diseases; although the mechanisms of action are not always clear, this could be a possible treatment for ARDS/ALI. Thus, the aim of this study was to evaluate the mRNA levels from caspase-3 and BCL-2 genes and DNA fragmentation in lung tissue from Wistar rats affected by ALI and subjected to photobiomodulation by exposure to a low power infrared laser (808 nm; 100 mW; 3.571 W cm-2; four points per lung). Adult male Wistar rats were randomized into 6 groups (n = 5, for each group): control, PBM10 (10 J cm-2, 2 J and 2 seconds), PBM20 (20 J cm-2, 5 J and 5 seconds), ALI, ALI + PBM10 and ALI + PBM20. ALI was induced by intraperitoneal Escherichia coli lipopolysaccharide injection. Lung samples were collected and divided for mRNA expression of caspase-3 and Bcl-2 and DNA fragmentation quantifications. Data show that caspase-3 mRNA levels are reduced and Bcl-2 mRNA levels increased in ALI after low power infrared laser exposure when compared to the non-exposed ALI group. DNA fragmentation increased in inflammatory infiltrate cells and reduced in alveolar cells. Our research shows that photobiomodulation can alter relative mRNA levels in genes involved in the apoptotic process and DNA fragmentation in inflammatory and alveolar cells after lipopolysaccharide-induced acute lung injury. Also, inflammatory cell apoptosis is part of the photobiomodulation effects induced by exposure to a low power infrared laser.
Assuntos
Lesão Pulmonar Aguda/terapia , Caspase 3/genética , Fragmentação do DNA/efeitos da radiação , Genes bcl-2/efeitos da radiação , Terapia com Luz de Baixa Intensidade , Pulmão/patologia , RNA Mensageiro/genética , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/patologia , Animais , Apoptose/efeitos da radiação , Regulação da Expressão Gênica/efeitos da radiação , Raios Infravermelhos/uso terapêutico , Pulmão/metabolismo , Pulmão/efeitos da radiação , Masculino , Ratos WistarRESUMO
BACKGROUND: Glioma is the most prevalent malignant tumor in human central nervous systems. Recently, the development of resistance to radiotherapy in glioma patients markedly vitiates the therapy outcome. MiR-153-3p has been reported to be closely correlated with tumor progression, but its effect and molecular mechanism underlying radioresistance remains unclear in glioma. METHODS: The expression of miR-153-3p was determined in radioresistant glioma clinical specimens as well as glioma cell lines exposed to irradiation (IR) using quantitative real-time PCR. Cell viability, proliferation and apoptosis were then evaluated by MTT assay, colony formation assay, Flow cytometry analysis and caspase-3 activity assay in glioma cells (U87 and U251). Tumor forming was evaluated by nude mice model in vivo. TUNEL staining was used to detect cell apoptosis in nude mice model. The target genes of miR-153-3p were predicted and validated using integrated bioinformatics analysis and a luciferase reporter assay. RESULTS: Here, we found that miR-153-3p was down-regulated in radioresistant glioma clinical specimens as well as glioma cell lines (U87 and U251) exposed to IR. Enhanced expression of miR-153-3p promoted the radiosensitivity, promoted apoptosis and elevated caspase-3 activity in glioma cells in vitro, as well as the radiosensitivity in U251 cell mouse xenografs in vivo. Mechanically, B cell lymphoma-2 gene (BCL2) was identified as the direct and functional target of miR-153-3p. Moreover, restoration of BCL2 expression reversed miR-153-3p-induced increase of radiosensitivity, apoptosis and caspase-3 activity in U251 cells in vitro. In addition, clinical data indicated that the expression of miR-153-3p was significantly negatively associated with BCL2 in radioresistance of glioma samples. CONCLUSIONS: Our findings suggest that miR-153-3p is a potential target to enhance the effect of radiosensitivity on glioma cells, thus representing a new potential therapeutic target for glioma.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Tolerância a Radiação/genética , Genes bcl-2/fisiologia , MicroRNAs/efeitos da radiação , MicroRNAs/fisiologia , Glioma/genética , Fatores de Tempo , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Sobrevivência Celular/efeitos da radiação , Western Blotting , Análise de Variância , Marcação de Genes/métodos , Genes bcl-2/efeitos da radiação , Marcação In Situ das Extremidades Cortadas , MicroRNAs/análise , Linhagem Celular Tumoral , Proliferação de Células/efeitos da radiação , Caspase 3/análise , Reação em Cadeia da Polimerase em Tempo Real , Citometria de Fluxo , Glioma/radioterapiaRESUMO
Pathological alterations to the retinal pigment epithelium underlie several eye diseases, which lead to visual impairment and even blindness. Exposure to ultraviolet (UV) radiation is associated with some skin and ocular pathologies; UV radiation may induce DNA breakdown and cause cellular damage through the production of reactive oxygen species (ROS), thus leading to programmed cell death. The present study aimed to investigate the production of ROS and the gene expression levels of anti and proapoptotic proteins [Bcell lymphoma 2 (Bcl2), Bcl2associated X protein (Bax) and caspase3] in human retinal pigment epithelial cells (ARPE19) treated with UVA for 5 h consecutively. The results demonstrated that prolonged exposure to UVA induced: i) Cell death, the decrease in cell viability was timedependent and reached statistical significance after 3 h; ii) a significant and substantial increase in ROS levels that remained constant for the duration of the experiment, the levels were significantly increased after 1 h of exposure; iii) an activation of apoptotic genes (Bax and caspase3) after 1 h of treatment, which was accompanied by a decrease in the antiapoptotic gene Bcl2; and iv) a loss of apoptotic signals and a rapid decrease in cellular viability after 3 h of consecutive treatment. These processes may trigger necrosis, which was observed in the cells following treatment with UVA for 5 consecutive hours. In conclusion, the present study is the first, to the best of our knowledge, to provide in vitro evidence regarding the sequence of events that underlie the cellular damage induced by prolonged UVA radiation, starting from the first 30 min of treatment. UVA radiation resulted in the activation of apoptotic events, and subsequently led to irreversible cell necrosis.
Assuntos
Caspase 3/genética , Sobrevivência Celular/efeitos da radiação , Regulação da Expressão Gênica/efeitos da radiação , Genes bcl-2/efeitos da radiação , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/efeitos da radiação , Proteína X Associada a bcl-2/genética , Apoptose/efeitos da radiação , Linhagem Celular , Humanos , Estresse Oxidativo/efeitos da radiação , Proteínas Proto-Oncogênicas c-bcl-2/genética , RNA Mensageiro/genética , Espécies Reativas de Oxigênio/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Raios UltravioletaRESUMO
Radiation exerts direct as well as indirect effects on DNA through the generation of reactive oxygen species (ROS). Irradiated hematopoietic progenitor cells (HPCs) experience DNA strand breaks, favoring genetic instability, due to ROS generation. Our aim was to study the effect of a range of radiation doses in HPCs and the possible protective mechanisms activated by insulin-like growth factor-1 (IGF-1). ROS generation was evaluated, in the presence or absence of IGF-1 in liquid cultures of human HPCs-CD34(+) irradiated with 1-, 2- and 5-Gy X-rays, using a flow cytometry assay. Manganese superoxide dismutase (MnSOD) expression was studied by western blot analysis and visualized by an immunofluorescence assay. Apoptosis was estimated using the following assays: Annexin-V assay, DNA degradation assay, BCL-2/BAX mRNA and protein levels and caspase-9 protein immunofluorescence visualization. Viability and clonogenic potential were studied in irradiated HPCs. The generation of superoxide anion radicals at an early and a late time point was increased, while the hydrogen peroxide generation at a late time point was stable. IGF-1 presence further enhanced the radiation-induced increase of MnSOD at 24 h post irradiation. IGF-1 inhibited the mitochondria-mediated pathway of apoptosis by regulating the m-RNA and protein expression of BAX, BCL-2 and the BCL-2/BAX ratio and by decreasing caspase-9 protein expression. IGF-1 presence in culture media of irradiated cells restored the clonogenic capacity and the viability of HPCs as well. In conclusion, IGF-1 protects HPCs-CD34(+) from radiation effects, by eliminating the oxidative microenvironment through the enhancement of MnSOD activation and by regulating the mitochondria-mediated pathway of apoptosis.
Assuntos
Células-Tronco Hematopoéticas/efeitos da radiação , Antígenos CD34/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Fragmentação do DNA/efeitos dos fármacos , Fragmentação do DNA/efeitos da radiação , Relação Dose-Resposta à Radiação , Genes bcl-2/efeitos dos fármacos , Genes bcl-2/efeitos da radiação , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/metabolismo , Humanos , Técnicas In Vitro , Fator de Crescimento Insulin-Like I/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/efeitos da radiação , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Superóxido Dismutase/metabolismo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
BACKGROUND: With the objective of developing a locally-produced radioactive stent, the present study used in vivo animal experiments to explore apoptosis of proliferative smooth muscle cells resulting from facilitation of the expression of genes caused by gamma-radiation in order to prevent bile duct restenosis. We therefore explored the effects and significance of gamma-radiation on the activity of caspase-3, Fas and Bcl-2 genes in apoptosis of proliferative smooth muscle cells in the bile duct walls of dogs. METHODS: Twelve dogs were randomly divided into 2 groups (6 in each group). A postinjury bile duct stenosis model was established and radioactive (103)Pd ((103)palladium) or ordinary bile duct stents were implanted into the bile ducts. HE staining, RT-PCR and immunohistochemistry were used to detect the proliferation and apoptosis of bile duct smooth muscle cells in proliferative endomembrane and the expression of related caspase-3, Bcl-2 and Fas genes. RESULTS: The expression of caspase-3 and Fas genes in the bile duct tissues of dogs with radioactive stents was higher than that of dogs with ordinary stents. There was significant apoptosis of proliferative smooth muscle cells in the bile ducts. The expression of the Bcl-2 gene in the bile duct tissues of dogs with radioactive stents was lower than that in those with ordinary stents. There was significant apoptosis of proliferative smooth muscle cells in the dogs with low Bcl-2 gene expression. CONCLUSIONS: Radiation increases the activity of caspase-3 and Fas genes and is associated with apoptosis. The radioactive (103)Pd stent may facilitate apoptosis of proliferative smooth muscle cells in the bile ducts of dogs by activating these genes. The Bcl-2 gene expression level is correlated with the occurrence of apoptosis and the radiosusceptibility of cells.
Assuntos
Apoptose/fisiologia , Ductos Biliares/ultraestrutura , Músculo Liso/ultraestrutura , Paládio/farmacologia , RNA Mensageiro/genética , Animais , Ductos Biliares/efeitos da radiação , Ductos Biliares/cirurgia , Caspase 3/genética , Caspase 3/efeitos da radiação , Proliferação de Células , Materiais Revestidos Biocompatíveis , Modelos Animais de Doenças , Cães , Feminino , Expressão Gênica/efeitos da radiação , Genes bcl-2/genética , Genes bcl-2/efeitos da radiação , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Isótopos , Masculino , Microscopia Eletrônica , Músculo Liso/efeitos da radiação , Implantação de Prótese/instrumentação , RNA Mensageiro/efeitos da radiação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Stents , Receptor fas/genética , Receptor fas/efeitos da radiaçãoRESUMO
Radiation-induced communication of stress signals between rainbow trout (Oncorhynchus mykiss W) have recently been described by this group and linked to the bystander effect. This paper addresses the question of whether another totally unrelated fish species (Danio rerio L) can demonstrate the effect and also looks at attenuation of both the bystander signal, from irradiated fish, and the bystander effect, in naive fish. The data show that zebrafish produce bystander signals, and that, as with rainbow trout these can affect naïve (i.e., non-irradiated) fish placed in water with X-rayed fish or in water previously occupied by X-rayed fish. Skin explants from directly X-rayed fish still reduce HPV-G reporter cell growth 6 h after X-ray, but the bystander signal to naïve fish is lost. Twelve h after X-ray the signal is lost in X-rayed fish. The bystander effect is also attenuated if induction was by placing naïve fish in water which previously held the X-rayed fish. However, the effect is retained if induction was by placing X-rayed and naïve fish together. This suggests the signal is not retained by water for long periods of time. Individual fish data reveal unique responses by bystander fish which could indicate varying levels of sensitivity to signal strength among individuals.
Assuntos
Efeito Espectador/efeitos da radiação , Raios X , Peixe-Zebra/fisiologia , Animais , Efeito Espectador/fisiologia , Feminino , Genes bcl-2/efeitos da radiação , Genes myc/efeitos da radiação , Brânquias/metabolismo , Brânquias/efeitos da radiação , Masculino , Pele/metabolismo , Pele/efeitos da radiação , Estresse FisiológicoRESUMO
The present study investigates the relationship between the subcellular localisation of Foscan and intrinsic apoptotic pathway post Foscan-based photodynamic therapy (PDT). With this purpose, mammary carcinoma MCF-7 cells were incubated with Foscan for 3 or 24 h and then subjected to equitoxic light doses. Fluorescence microscopy revealed very good Foscan co-localization to endoplasmic reticulum (ER) and Golgi apparatus after 3 h incubation with MCF-7 cells. Progressive increase in incubation time shows leakage of Foscan from Golgi apparatus. Twenty-four hours incubation yielded a fluence-dependent enhanced induction of the ER-resident glucose-regulated protein 78 (Bip/GRP78), along with a weak mitochondrial damage, thus underscoring the ER as the main site of photodamage after prolonged incubation. Analysis of events implicated in apoptotic pathway after 24 h incubation demonstrated photodamage to Bcl-2 protein in total cellular extract, but not in the mitochondrial fraction. We further determined an increase in caspases-7 and -6 activation, which was strongly related to the expression of GRP78. The above findings demonstrate that Foscan localisation in ER improves the photoactivation of the caspase-7 apoptotic pathway, which is poorly related to mitochondrial damage.
Assuntos
Neoplasias da Mama/metabolismo , Caspases/metabolismo , Mesoporfirinas/farmacocinética , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/farmacocinética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/enzimologia , Linhagem Celular Tumoral , Chaperona BiP do Retículo Endoplasmático , Ativação Enzimática , Fluorometria , Genes bcl-2/efeitos da radiação , Proteínas de Choque Térmico/biossíntese , Humanos , Mesoporfirinas/farmacologia , Mitocôndrias/enzimologia , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Chaperonas Moleculares/biossíntese , Fármacos Fotossensibilizantes/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Frações Subcelulares/metabolismoRESUMO
BACKGROUND: When dissection of porcine eyes from a living body results in the cessation of aqueous humor production and blood flow, programmed cell death regulated by TPR53BP2 and BCL2 genes may occur in the pigmented epithelium (PE) and non-pigmented epithelium (NPE) of the ciliary body. Blood products are subjected to gamma irradiation in order to prevent cellular damage resulting from transfusion-associated graft-versus-host disease. We investigated whether gamma irradiation influenced BCL2 or TPR53BP2 genes as well as the morphology of the porcine ciliary body. METHODS: We irradiated the anterior segments of porcine eyes by using (60)Co gamma-rays (20 Gy). To study BCL2 and TPR53BP2 expression, the irradiated specimens were fixed in formalin and embedded in paraffin and then incubated with mouse monoclonal anti-human BCL2 or TPR53BP2 antibody. RESULTS: Following dissection, an imbalance in homeostasis began with positive BCL2 and TPR53BP2 expression in the edematous ciliary processes, and resulted in atrophy of the NPE. Increased BCL2 and TPR53BP2 expression were evident just after gamma irradiation. Decreased TPR53BP2 expression occurred after 8 h of incubation, and thereby suppressed apoptosis in the NPE; hence, the structure of the ciliary body that was incubated for 8 h after gamma irradiation was well preserved. CONCLUSIONS: Irradiation renders the ciliary body in enucleated porcine eyes less vulnerable to apoptosis, and thereby exerts a profound preservative effect.
Assuntos
Apoptose/genética , Corpo Ciliar/efeitos da radiação , Raios gama , Genes bcl-2/efeitos da radiação , Proteínas Proto-Oncogênicas/genética , Animais , Morte Celular/genética , Corpo Ciliar/patologia , Epitélio/efeitos da radiação , Expressão Gênica/efeitos da radiação , Técnicas In Vitro , Preservação de Órgãos/métodos , SuínosRESUMO
We have quantified the emergence of early chromatin breaks during the signal transduction phase of apoptosis in mouse thymocytes after treatment with either ionizing radiation or dexamethasone. Dexamethasone at 1 microM can induce significant levels of DNA breaks (equivalent to the amount induced directly by 7.5 Gy ionizing radiation) within 0.5 h of treatment. The execution phase of apoptosis was not observed until 4-6 h after the same treatment. The presence of the Bcl2 transgene under the control of the p56lck promoter almost completely inhibited apoptosis up to 24 h after treatment, but it had virtually no effect on the early chromatin cleavage occurring in the first 6 h. Ionizing radiation induced chromatin cleavage both directly by damaging DNA and indirectly with kinetics similar to the induction of chromatin cleavage by dexamethasone. The presence of the Bcl2 transgene had no effect on the direct or indirect radiation-induced cleavage in the first 6 h, but after the first 6 h, the Bcl2 gene inhibited further radiation-induced chromatin cleavage. These results suggest that endonucleases are activated within minutes of treatment with either dexamethasone or ionizing radiation as part of the very early signal transduction phase of apoptosis, and prior to the irreversible commitment to cell death.
Assuntos
Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Dano ao DNA , Dexametasona/farmacologia , Proteínas Proto-Oncogênicas/metabolismo , Timo/efeitos dos fármacos , Timo/efeitos da radiação , Animais , Apoptose/genética , Divisão Celular , Células Cultivadas , Cromatina/efeitos dos fármacos , Cromatina/efeitos da radiação , Feminino , Genes bcl-2/efeitos dos fármacos , Genes bcl-2/efeitos da radiação , Masculino , Camundongos , Camundongos Transgênicos , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-bcl-2 , Radiação Ionizante , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Timo/patologia , TransgenesAssuntos
Carcinoma de Células Escamosas/terapia , Neoplasias Esofágicas/terapia , Apoptose/efeitos da radiação , Braquiterapia , Carcinoma de Células Escamosas/epidemiologia , Carcinoma de Células Escamosas/genética , Neoplasias Esofágicas/epidemiologia , Neoplasias Esofágicas/genética , Esofagectomia , Genes bcl-2/genética , Genes bcl-2/efeitos da radiação , Genes p53/genética , Genes p53/efeitos da radiação , Humanos , Incidência , Prognóstico , Radioterapia Adjuvante , África do Sul/epidemiologiaRESUMO
Pre-brachytherapy biopsies and post-brachytherapy oesophagectomy specimens of 10 patients with early squamous cell carcinoma of the middle third of the oesophagus were examined for the expression of p53, bcl-2 and apoptosis using immunohistochemical markers. There was no expression of p53 in one patient in both pre- and post-brachytherapy specimens. In 8 patients, p53 staining was strongly positive (3+) with approximately 50% or more cells, and with diffuse and no specific pattern in the pre-brachytherapy biopsies. The tumour areas of the post-brachytherapy specimens of this group showed strong 3+ positivity with p53 (10-50% positive cell count), with the pattern being focal and peripheral in the tumour islands. The centre of the tumour islands showed necrosis and/or keratinisation. In one patient, the pre-brachytherapy biopsy showed expression of p53 while the post-brachytherapy specimen was negative. bcl-2 expression in both pre- and post-brachytherapy was equivocal and inconclusive in both the pre- and post-brachytherapy specimens. Apoptosis was negative in all the pre- and post-brachytherapy tissue sections in the presence of positive controls. Brachytherapy does not cause cell death by apoptosis but by necrosis and maturation of the cells into better differentiated cells, which is caused by OH free radical, and induction of the keratin gene respectively. It is possible that brachytherapy may cause destruction of cells containing wild-type p53, while mutant p53 in cells located at the tumour periphery escape the effect of brachytherapy. This may be responsible for the high incidence of local recurrence and distant metastasis in oesophageal cancer treated with radiotherapy. There is no effect of brachytherapy on bcl-2 expression in oesophageal cancer.
Assuntos
Apoptose/efeitos da radiação , Braquiterapia/métodos , Carcinoma de Células Escamosas/terapia , Neoplasias Esofágicas/terapia , Expressão Gênica/efeitos da radiação , Genes bcl-2/efeitos da radiação , Genes p53/efeitos da radiação , Adulto , Idoso , Biópsia , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/genética , Neoplasias Esofágicas/diagnóstico , Neoplasias Esofágicas/genética , Esofagectomia , Feminino , Expressão Gênica/genética , Genes bcl-2/genética , Genes p53/genética , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Mutação/genética , Radioterapia Adjuvante , Resultado do TratamentoRESUMO
BACKGROUND: Recently, apoptosis has been identified in treated lymphocytes, prior to their re-infusion, when tested ex vivo. Previous work has demonstrated a close association between the genes p53, Bcl-2 and Bax and apoptosis induced by UV irradiation. OBJECTIVES: We wanted to establish whether the expression of the protein product of these genes was altered in lymphocytes treated with extracorporeal photopheresis (ECP) prior to re-infusion and therefore possibly implicated in the early apoptosis observed. METHOD: Lymphocytes were isolated immediately before treatment and immediately prior to re-infusion and tested for intracellular levels of p53, Bcl-2 and Bax proteins. RESULTS: No increase in p53 expression was observed at re-infusion; however, the mean fluorescent intensity ratio of the apoptotic inhibitor protein Bcl-2 to the apoptosis-inducing protein Bax dropped significantly. CONCLUSION: The early apoptosis observed in ECP-treated lymphocytes at re-infusion might be attributed to dysregulation in the expression of the apoptotic genes Bcl-2 and Bax.
Assuntos
Apoptose/efeitos da radiação , Genes bcl-2/efeitos da radiação , Linfócitos/metabolismo , Linfoma de Células T/genética , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Proto-Oncogênicas/genética , Proteína Supressora de Tumor p53/genética , Apoptose/fisiologia , Citometria de Fluxo , Expressão Gênica/efeitos da radiação , Humanos , Linfócitos/efeitos da radiação , Linfoma de Células T/imunologia , Linfoma de Células T/metabolismo , Fotoferese , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas/efeitos da radiação , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/efeitos da radiação , Proteína X Associada a bcl-2RESUMO
Radiation therapy plays an important role as part of multimodality treatment for a number of childhood malignancies. The damaging effects of radiation on bone formation in children have been well documented. Recent work suggests that the postirradiation increase in cytosolic calcium is probably responsible for the deleterious effects of radiation on growth plate chondrocytes because it causes a specific suppression of the mitogen PTHrP. Using an in vitro model of avian growth plate chondrocytes, this study demonstrates that pentoxifylline is effective in increasing basal PTHrP mRNA levels and partially preventing the radiation-induced decrease in PTHrP mRNA. This effect of pentoxifylline is probably due to its ability to lower basal levels of cytosolic calcium and the radiation-induced increase in cytosolic calcium in chondrocytes. Pentoxifylline also prevented the radiation-induced decreases in [3H]thymidine uptake and BCL2 and PTHrP receptor mRNA levels in chondrocytes. The effects of pentoxifylline appear to be specific for the PTHrP signaling pathway because it did not alter basal TGFB mRNA levels or TGFB mRNA expression in irradiated chondrocytes. The results of the current study suggest that by decreasing basal cytosolic calcium levels and curtailing the radiation-induced increase in cytosolic calcium levels in chondrocytes, pentoxifylline is able to sustain PTHrP signaling in chondrocytes and maintains the proliferative signal that is necessary to prevent chondrocytes from undergoing apoptosis.
Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Condrócitos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Lâmina de Crescimento/efeitos dos fármacos , Pentoxifilina/farmacologia , Protetores contra Radiação/farmacologia , Animais , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Embrião de Galinha , Condrócitos/efeitos da radiação , Citosol/metabolismo , Avaliação Pré-Clínica de Medicamentos , Genes bcl-2/efeitos da radiação , Lâmina de Crescimento/efeitos da radiação , Proteína Relacionada ao Hormônio Paratireóideo , Biossíntese de Proteínas , Proteínas/genética , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , RNA Mensageiro/biossíntese , Tolerância a Radiação/efeitos dos fármacos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Cepharanthin (CE) is a medicine that contains several biscoclaurine alkaloids. We examined the effects of CE on radiation-induced T cell apoptosis. Radiation induced apoptosis on T cells in a dose-dependent manner, while CE inhibited radiation-induced apoptosis. CE also attenuated the cytotoxic effects of radiation on the proliferative response of T cells. CE inhibited not only the loss of mitochondrial transmembrane potential but also the activation of caspase 3 in irradiated T cells. Radiation plus CE induced the up-regulation of Bax and the down-regulation of Bcl-2 in T cells in comparison with radiation alone. These results suggest that CE inhibits the signal transduction pathway of apoptosis induced by radiation, regardless of the expression of Bcl-2 or Bax.
Assuntos
Alcaloides/farmacologia , Apoptose/efeitos dos fármacos , Benzilisoquinolinas , Protetores contra Radiação/farmacologia , Linfócitos T/efeitos dos fármacos , Adulto , Apoptose/efeitos da radiação , Caspase 3 , Caspases/metabolismo , Células Cultivadas/citologia , Células Cultivadas/efeitos dos fármacos , Células Cultivadas/metabolismo , Células Cultivadas/efeitos da radiação , Relação Dose-Resposta à Radiação , Combinação de Medicamentos , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/efeitos da radiação , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Genes bcl-2/efeitos dos fármacos , Genes bcl-2/efeitos da radiação , Genes p53/efeitos dos fármacos , Genes p53/efeitos da radiação , Humanos , Membranas Intracelulares/efeitos dos fármacos , Membranas Intracelulares/efeitos da radiação , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/efeitos da radiação , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/efeitos da radiação , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/efeitos da radiação , Fito-Hemaglutininas/farmacologia , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Linfócitos T/efeitos da radiação , Proteína Supressora de Tumor p53/biossíntese , Proteína X Associada a bcl-2RESUMO
In this paper we present some preliminary results on alteration of gene expression caused by radiation on human endometrial cells. To this purpose, we have studied the modulation of the expression of the bcl-2 gene family in two cell lines following irradiations with low energy protons and gamma-rays from a 60Co. The two epithelial cell strains, namely AN3Ca and HEC1B cells, both obtained from human neoplastic endometrial tissues, grow in culture and continue to maintain some differentiated functions typical of the original tissue. Indeed, these cells, that can be considered as representative of different stages of cellular transformation of endometrium. Because their epithelial nature and rapid growth, the expression of genes related to the maintenance of the cellular homeostasis (correction of omeostasis), as the pro and anti-apoptotic ones, is expected to be susceptible to changes in environment, including radiation. The effects have been evaluated in terms of both cell survival and changes in the expression of pro- and anti apoptotic proteins. Even though the data reported above can not be considered complete and/or definitive, nevertheless, in whole, they confirm that these cells may constitute a suitable model system to study, at molecular level, the effects of cosmic radiation on endometrium. Further observation, ensuing from these preliminary data, is that endometrial cells present different sensitivity to radiation in regard to its 'quality' and 'dosage', in accord to the original stage of differentiation.
Assuntos
Apoptose/efeitos da radiação , Radiação Cósmica , Endométrio/efeitos da radiação , Regulação da Expressão Gênica/efeitos da radiação , Genes bcl-2/efeitos da radiação , Ciclo Celular/efeitos da radiação , Linhagem Celular , Transformação Celular Neoplásica/genética , Relação Dose-Resposta à Radiação , Endométrio/citologia , Feminino , Raios gama , Humanos , Prótons , Células Tumorais Cultivadas/efeitos da radiaçãoRESUMO
Several protocols for the adjuvant treatment of glioblastoma multiforme (GBM) are currently being evaluated. In this context, little is known about the influence of radiochemotherapy on apoptosis and the expression of apoptosis-related proteins in vivo. We have analyzed the incidence of apoptosis using in situ nick translation (ISNT) and expression of Ki-67 (MIB- 1), p53 (DO-1 and DO-7), Bcl-2 and transglutaminase II (TGase II) by immunohistochemistry in 41 patients with GBM and their matched relapses. Sixteen patients received radiochemotherapy, 18 irradiation and 7 no treatment. Radiochemotherapy resulted in an increase in Bcl-2+ cells (p = 0.013). Irradiation caused the reduction of MIB-1+ (p = 0.0015), DO-7+ (p = 0.0043) and the increase of Bcl-2+ cells (p = 0.016). We calculated a positive correlation between high TGase II scores in patients preceding radiochemotherapy (p = 0.0186) and no treatment (p = 0.0158), low ISNT scores (p = 0.0018) and high DO-1 scores (p = 0.0233) in patients preceding irradiation and short time to progression. These data show that distinct postsurgical radiochemotherapy protocols differentially alter cellular proliferation and expression of p53 and Bcl-2 in GBM relapses. Furthermore, we show that ISNT, DO-I and TGase II labeling scores are therapy-specific predictors of time to progression in GBM patients.
Assuntos
Neoplasias Encefálicas/patologia , Quimioterapia Adjuvante , Irradiação Craniana , Regulação Neoplásica da Expressão Gênica , Genes bcl-2 , Genes , Glioblastoma/patologia , Recidiva Local de Neoplasia/patologia , Radioterapia Adjuvante , Adulto , Idoso , Antineoplásicos Alquilantes/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Biomarcadores , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/radioterapia , Neoplasias Encefálicas/cirurgia , Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Terapia Combinada , Ciclofosfamida/análogos & derivados , Ciclofosfamida/uso terapêutico , Citarabina/administração & dosagem , Progressão da Doença , Feminino , Proteínas de Ligação ao GTP/biossíntese , Proteínas de Ligação ao GTP/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Genes/efeitos dos fármacos , Genes/efeitos da radiação , Genes bcl-2/efeitos dos fármacos , Genes bcl-2/efeitos da radiação , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Glioblastoma/mortalidade , Glioblastoma/radioterapia , Glioblastoma/cirurgia , Humanos , Antígeno Ki-67/biossíntese , Antígeno Ki-67/genética , Tábuas de Vida , Lomustina/uso terapêutico , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/tratamento farmacológico , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/mortalidade , Recidiva Local de Neoplasia/radioterapia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos da radiação , Nimustina/administração & dosagem , Nimustina/uso terapêutico , Proteína 2 Glutamina gama-Glutamiltransferase , Proteínas Proto-Oncogênicas c-bcl-2/biossínteseRESUMO
Studies of the mechanism of actions of estrogen, antiestrogen and physical factors may provide clues to an understanding of breast cancer growth and/or regression regulation and thus identify novel targets for therapeutic intervention. Defective control of apoptosis appears to play a central role in the pathogenesis of neoplasia. Conversely, cancer therapy and ionizing radiation can induce cancer cell death by apoptosis and/or necrosis. bcl-2 gene and p-53 gene products have been both linked to programmed cell death pathways. We have analyzed the effect of estradiol, tamoxifen and UV exposure on the induction of apoptosis, expression of p53 and bcl-2 gene products as well as the proliferative activity (expressed as [3H]thymidine incorporation and PCNA and MPM2 antigens involvement) in MCF7. It has been found that estradiol increases the speed of cell cycle in MCF7 and acts as antiapoptotic factor. Tamoxifen has multiple influence on the rate of growth of cancer cells: depends on estrogen receptor (ER), conducts reduction of proliferation rate; depends on ER and other mechanisms conducts to suppressions of Bcl-2 protein expression and induction of cell death through apoptotic pathway. Estradiol prevents the apoptotic influence of tamoxifen probably by enhancement of Bcl-2 protein expression and does not prevent the inhibition of proliferation rate. The irradiation with UV induces apoptosis by over-expression of p53 and down-regulation of bcl-2 gene.
Assuntos
Adenocarcinoma/patologia , Neoplasias da Mama/patologia , Proteínas de Ciclo Celular , Estradiol/farmacologia , Antagonistas de Estrogênios/farmacologia , Tamoxifeno/farmacologia , Raios Ultravioleta , Adenocarcinoma/metabolismo , Anticorpos Monoclonais/efeitos dos fármacos , Anticorpos Monoclonais/efeitos da radiação , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Neoplasias da Mama/metabolismo , Divisão Celular/efeitos dos fármacos , Divisão Celular/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Relação Dose-Resposta a Droga , Feminino , Regulação Neoplásica da Expressão Gênica , Genes bcl-2/efeitos dos fármacos , Genes bcl-2/efeitos da radiação , Humanos , Imuno-Histoquímica , Cinesinas , Necrose , Fosfoproteínas/efeitos dos fármacos , Fosfoproteínas/efeitos da radiação , Antígeno Nuclear de Célula em Proliferação/efeitos dos fármacos , Antígeno Nuclear de Célula em Proliferação/efeitos da radiação , Fatores de Tempo , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/efeitos da radiação , Proteína Supressora de Tumor p53/efeitos dos fármacos , Proteína Supressora de Tumor p53/efeitos da radiaçãoRESUMO
Between 1983 and 1995 we have monitored human populations for evidence of exposure to environmental mutagens, taking blood samples to measure hprt(-) mutant frequency in T cells and more recently bcl-2 t(14:18) translocation frequency in B cells. We have now analysed data from 785 assays on 448 blood samples from 308 normal subjects and find that there is a highly significant statistical correlation between hprt(-) mutant frequency and the sunlight record for the 3 weeks prior to taking the blood sample. We discuss the weaknesses in retrospective studies of this nature and the possibility of spurious epidemiological correlations that may result. More controlled experiments can be envisaged that would give a firmer basis to the statistical associations observed. hprt(-) mutations in T cells show little evidence of a UV fingerprint, so that the correlation may be due to immunomodulation rather than mutation. We also find a correlation between the sunlight record and bcl-2 translocation. This translocation is found at a low frequency in the B cells of many normal subjects and is the commonest translocation observed in non-Hodgkin's lymphoma. Our results strengthen the case for a link between sunlight and this increasingly common cancer.
Assuntos
Genes bcl-2/efeitos da radiação , Hipoxantina Fosforribosiltransferase/genética , Linfócitos/efeitos da radiação , Mutação , Luz Solar , Translocação Genética/efeitos da radiação , Bases de Dados Factuais , Genes bcl-2/genética , Humanos , Linfoma não Hodgkin/metabolismo , Mutação/genética , Mutação/efeitos da radiação , Vigilância da População/métodos , Luz Solar/efeitos adversos , Translocação Genética/genética , Raios Ultravioleta/efeitos adversos , Reino Unido/epidemiologiaRESUMO
BACKGROUND: Apoptosis plays a crucial role in radiation therapy (RT) in various carcinomas. This study was designed to investigate the relation between apoptosis and RT in invasive squamous cell carcinoma (ISCC) of the uterine cervix METHODS: Thirty-five specimens were obtained from 7 patients with ISCC before and during a fractionated RT. The occurrence of apoptosis was examined by end labeling of DNA gel fractionation and in situ 3' end labeling of DNA. The expression of Bax and Bcl-2 proteins was investigated by immunohistochemical staining. RESULTS: Autoradiographic analysis revealed that high molecular weight DNA was predominant in the untreated ISCC specimens. However, a ladder-like pattern, characteristic of the apoptotic breakdown of DNA, was identified at doses of 900 cGy and 1980 cGy. At doses >1980 cGy, DNA laddering disappeared without any extensive smearing. Quantitative analysis of low molecular weight fragments of DNA revealed significant increases at doses of 900 cGy and 1980 cGy compared with those before RT and at doses of >1980 cGy. Labeling of DNA in situ indicated that cells undergoing apoptosis increased dramatically at a dose of 900 cGy. However, apoptotic cells decreased at a dose of 3960 cGy. In addition, a large fraction of tumor cells was immunonegative for Bcl-2 before and during RT. By contrast, immunoreactive Bax was observed intensely in many neoplastic cells at doses of 900 cGy and 1980 cGy. CONCLUSIONS: The current investigation indicates that low doses of RT result in apoptotic cell death in ISCC in association with the increased expression of Bax but not with increased Bcl-2 expression.