RESUMO
Extracellular vesicles (EVs), through their cargo, are important mediators of bystander responses in the irradiated bone marrow (BM). MiRNAs carried by EVs can potentially alter cellular pathways in EV-recipient cells by regulating their protein content. Using the CBA/Ca mouse model, we characterised the miRNA content of BM-derived EVs from mice irradiated with 0.1 Gy or 3 Gy using an nCounter analysis system. We also analysed proteomic changes in BM cells either directly irradiated or treated with EVs derived from the BM of irradiated mice. Our aim was to identify key cellular processes in the EV-acceptor cells regulated by miRNAs. The irradiation of BM cells with 0.1 Gy led to protein alterations involved in oxidative stress and immune and inflammatory processes. Oxidative stress-related pathways were also present in BM cells treated with EVs isolated from 0.1 Gy-irradiated mice, indicating the propagation of oxidative stress in a bystander manner. The irradiation of BM cells with 3 Gy led to protein pathway alterations involved in the DNA damage response, metabolism, cell death and immune and inflammatory processes. The majority of these pathways were also altered in BM cells treated with EVs from mice irradiated with 3 Gy. Certain pathways (cell cycle, acute and chronic myeloid leukaemia) regulated by miRNAs differentially expressed in EVs isolated from mice irradiated with 3 Gy overlapped with protein pathway alterations in BM cells treated with 3 Gy EVs. Six miRNAs were involved in these common pathways interacting with 11 proteins, suggesting the involvement of miRNAs in the EV-mediated bystander processes. In conclusion, we characterised proteomic changes in directly irradiated and EV-treated BM cells, identified processes transmitted in a bystander manner and suggested miRNA and protein candidates potentially involved in the regulation of these bystander processes.
Assuntos
Vesículas Extracelulares , MicroRNAs , Camundongos , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Medula Óssea/metabolismo , Proteômica , Camundongos Endogâmicos CBA , Vesículas Extracelulares/metabolismo , Radiação IonizanteRESUMO
Cell communication via exosomes is capable of influencing cell fate in stress situations such as exposure to ionizing radiation. In vitro and in vivo studies have shown that exosomes might play a role in out-of-target radiation effects by carrying molecular signaling mediators of radiation damage, as well as opposite protective functions resulting in resistance to radiotherapy. However, a global understanding of exosomes and their radiation-induced regulation, especially within the context of an intact mammalian organism, has been lacking. In this in vivo study, we demonstrate that, compared to sham-irradiated (SI) mice, a distinct pattern of proteins and miRNAs is found packaged into circulating plasma exosomes after whole-body and partial-body irradiation (WBI and PBI) with 2 Gy X-rays. A high number of deregulated proteins (59% of WBI and 67% of PBI) was found in the exosomes of irradiated mice. In total, 57 and 13 miRNAs were deregulated in WBI and PBI groups, respectively, suggesting that the miRNA cargo is influenced by the tissue volume exposed to radiation. In addition, five miRNAs (miR-99b-3p, miR-200a-3p, miR-200a, miR-182-5p, miR-182) were commonly overexpressed in the exosomes from the WBI and PBI groups. In this study, particular emphasis was also given to the determination of the in vivo effect of exosome transfer by intracranial injection in the highly radiosensitive neonatal cerebellum at postnatal day 3. In accordance with a major overall anti-apoptotic function of the commonly deregulated miRNAs, here, we report that exosomes from the plasma of irradiated mice, especially in the case of WBI, prevent radiation-induced apoptosis, thus holding promise for exosome-based future therapeutic applications against radiation injury.
Assuntos
Exossomos , MicroRNAs , Lesões por Radiação , Animais , Apoptose , Cerebelo/metabolismo , Exossomos/metabolismo , Mamíferos/metabolismo , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Proteômica , Lesões por Radiação/metabolismoRESUMO
The brain undergoes ionizing radiation exposure in many clinical situations, particularly during radiotherapy for brain tumors. The critical role of the hippocampus in the pathogenesis of radiation-induced neurocognitive dysfunction is well recognized. The goal of this study is to test the potential contribution of non-targeted effects in the detrimental response of the hippocampus to irradiation and to elucidate the mechanisms involved. C57Bl/6 mice were whole body (WBI) or partial body (PBI) irradiated with 0.1 or 2.0 Gy of X-rays or sham irradiated. PBI consisted of the exposure of the lower third of the mouse body, whilst the upper two thirds were shielded. Hippocampi were collected 15 days or 6 months post-irradiation and a multi-omics approach was adopted to assess the molecular changes in non-coding RNAs, proteins and metabolic levels, as well as histological changes in the rate of hippocampal neurogenesis. Notably, at 2.0 Gy the pattern of early molecular and histopathological changes induced in the hippocampus at 15 days following PBI were similar in quality and quantity to the effects induced by WBI, thus providing a proof of principle of the existence of out-of-target radiation response in the hippocampus of conventional mice. We detected major alterations in DAG/IP3 and TGF-ß signaling pathways as well as in the expression of proteins involved in the regulation of long-term neuronal synaptic plasticity and synapse organization, coupled with defects in neural stem cells self-renewal in the hippocampal dentate gyrus. However, compared to the persistence of the WBI effects, most of the PBI effects were only transient and tended to decrease at 6 months post-irradiation, indicating important mechanistic difference. On the contrary, at low dose we identified a progressive accumulation of molecular defects that tended to manifest at later post-irradiation times. These data, indicating that both targeted and non-targeted radiation effects might contribute to the pathogenesis of hippocampal radiation-damage, have general implications for human health.
Assuntos
Irradiação Craniana , Hipocampo/metabolismo , Hipocampo/efeitos da radiação , Metaboloma , Neurogênese/genética , Neurogênese/efeitos da radiação , Proteoma , Transcriptoma , Animais , Biologia Computacional/métodos , Irradiação Craniana/efeitos adversos , Feminino , Regulação da Expressão Gênica , Imuno-Histoquímica , Camundongos , Doses de Radiação , Transdução de SinaisRESUMO
The impact of low-dose ionizing radiation (IR) on the human brain has recently attracted attention due to the increased use of IR for diagnostic purposes. The aim of this study was to investigate low-dose radiation response in the hippocampus. Female B6C3F1 mice were exposed to total body irradiation with 0 (control), 0.063, 0.125, or 0.5 Gy. Quantitative label-free proteomic analysis of the hippocampus was performed after 24 months. CREB signaling and CREB-associated pathways were affected at all doses. The lower doses (0.063 and 0.125 Gy) induced the CREB pathway, whereas the exposure to 0.5 Gy deactivated CREB. Similarly, the lowest dose (0.063 Gy) was anti-inflammatory, reducing the number of activated microglia. In contrast, induction of activated microglia and reactive astroglia was found at 0.5 Gy, suggesting increased inflammation and astrogliosis, respectively. The apoptotic markers BAX and cleaved CASP-3 and oxidative stress markers were increased only at the highest dose. Since the activated CREB pathway plays a central role in learning and memory, these data suggest neuroprotection at the lowest dose (0.063 Gy) but neurodegeneration at 0.5 Gy. The response to 0.5 Gy resembles alterations found in healthy aging and thus may represent radiation-induced accelerated aging of the brain.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Hipocampo/metabolismo , Hipocampo/efeitos da radiação , Animais , Apoptose/efeitos da radiação , Relação Dose-Resposta à Radiação , Feminino , Inflamação/etiologia , Camundongos Endogâmicos , Plasticidade Neuronal/efeitos da radiação , Estresse Oxidativo/efeitos da radiação , Carbonilação Proteica/efeitos da radiação , Radiação Ionizante , Transdução de Sinais/efeitos da radiação , Fatores de Tempo , Irradiação Corporal TotalRESUMO
In the current dismal situation of the COVID-19 pandemic, effective management of patients with pneumonia and acute respiratory distress syndrome is of vital importance. Due to the current lack of effective pharmacological concepts, this situation has caused interest in (re)considering historical reports on the treatment of patients with low-dose radiation therapy for pneumonia. Although these historical reports are of low-level evidence per se, hampering recommendations for decision-making in the clinical setting, they indicate effectiveness in the dose range between 0.3 and 1â¯Gy, similar to more recent dose concepts in the treatment of acute and chronic inflammatory/degenerative benign diseases with, e.g., a single dose per fraction of 0.5â¯Gy. This concise review aims to critically review the evidence for low-dose radiation treatment of COVID-19 pneumopathy and discuss whether it is worth investigating in the present clinical situation.
Assuntos
Infecções por Coronavirus/radioterapia , Pneumonia Viral/radioterapia , Síndrome Respiratória Aguda Grave/radioterapia , COVID-19 , Medicina Baseada em Evidências , Humanos , Pandemias , Dosagem Radioterapêutica , Resultado do TratamentoRESUMO
Molecular communication between irradiated and unirradiated neighbouring cells initiates radiation-induced bystander effects (RIBE) and out-of-field (abscopal) effects which are both an example of the non-targeted effects (NTE) of ionising radiation (IR). Exosomes are small membrane vesicles of endosomal origin and newly identified mediators of NTE. Although exosome-mediated changes are well documented in radiation therapy and oncology, there is a lack of knowledge regarding the role of exosomes derived from inside and outside the radiation field in the early and delayed induction of NTE following IR. Therefore, here we investigated the changes in exosome profile and the role of exosomes as possible molecular signalling mediators of radiation damage. Exosomes derived from organs of whole body irradiated (WBI) or partial body irradiated (PBI) mice after 24 h and 15 days post-irradiation were transferred to recipient mouse embryonic fibroblast (MEF) cells and changes in cellular viability, DNA damage and calcium, reactive oxygen species and nitric oxide signalling were evaluated compared to that of MEF cells treated with exosomes derived from unirradiated mice. Taken together, our results show that whole and partial-body irradiation increases the number of exosomes, instigating changes in exosome-treated MEF cells, depending on the source organ and time after exposure.
Assuntos
Exossomos/efeitos da radiação , Lesões por Radiação/patologia , Animais , Efeito Espectador/efeitos da radiação , Cálcio/metabolismo , Sobrevivência Celular/efeitos da radiação , Células Cultivadas , Dano ao DNA/efeitos da radiação , Exossomos/metabolismo , Feminino , Fibroblastos/metabolismo , Fibroblastos/efeitos da radiação , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico/metabolismo , Lesões por Radiação/metabolismo , Radiação Ionizante , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos da radiaçãoRESUMO
Normal tissue toxicity is a dose-limiting factor in radiation therapy. Therefore, a detailed understanding of the normal tissue response to radiation is necessary to predict the risk of normal tissue toxicity and to development strategies for tissue protection. One component of normal tissue that is continuously exposed during therapeutic irradiation is the circulating population of peripheral blood mononuclear cells (PBMC). PBMCs are highly sensitive to ionizing radiation (IR); however, little is known about how IR affects the PBMC response on a systemic level. It was the aim of this study to investigate whether IR was capable to induce changes in the composition and function of extracellular vesicles (EVs) secreted from PBMCs after radiation exposure to different doses. Therefore, whole blood samples from healthy donors were exposed to X-ray radiation in the clinically relevant doses of 0, 0.1, 2 or 6 Gy and PBMC-secreted EVs were isolated 72 h later. Proteome and miRNome analysis of EVs as well as functional studies were performed. Secreted EVs showed a dose-dependent increase in the number of significantly deregulated proteins and microRNAs. For both, proteome and microRNA data, principal component analysis showed a dose-dependent separation of control and exposed groups. Integrated pathway analysis of the radiation-regulated EV proteins and microRNAs consistently predicted an association of deregulated molecules with apoptosis, cell death and survival. Functional studies identified endothelial cells as an efficient EV recipient system, in which irradiation of recipient cells further increased the uptake. Furthermore an apoptosis suppressive effect of EVs from irradiated PBMCs in endothelial recipient cells was detected. In summary, this study demonstrates that IR modifies the communication between PBMCs and endothelial cells. EVs from irradiated PBMC donors were identified as transmitters of protective signals to irradiated endothelial cells. Thus, these data may lead to the discovery of biomarker candidates for radiation dosimetry and even more importantly, they suggest EVs as a novel systemic communication pathway between irradiated normal, non-cancer tissues.
Assuntos
Vesículas Extracelulares/metabolismo , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/efeitos da radiação , Exposição à Radiação , Vesículas Secretórias/metabolismo , Apoptose/efeitos da radiação , Células Endoteliais/metabolismo , Células Endoteliais/efeitos da radiação , Humanos , MicroRNAs/genética , Proteoma/metabolismo , Radiação Ionizante , Radioterapia/métodosRESUMO
Epidemiological studies on workers employed at the Mayak plutonium enrichment plant have demonstrated an association between external gamma ray exposure and an elevated risk of ischemic heart disease (IHD). In a previous study using fresh-frozen post mortem samples of the cardiac left ventricle of Mayak workers and non-irradiated controls, we observed radiation-induced alterations in the heart proteome, mainly downregulation of mitochondrial and structural proteins. As the control group available at that time was younger than the irradiated group, we could not exclude age as a confounding factor. To address this issue, we have now expanded our study to investigate additional samples using archival formalin-fixed paraffin-embedded (FFPE) tissue. Importantly, the control group studied here is older than the occupationally exposed (>500 mGy) group. Label-free quantitative proteomics analysis showed that proteins involved in the lipid metabolism, sirtuin signaling, mitochondrial function, cytoskeletal organization, and antioxidant defense were the most affected. A histopathological analysis elucidated large foci of fibrotic tissue, myocardial lipomatosis and lymphocytic infiltrations in the irradiated samples. These data highlight the suitability of FFPE material for proteomics analysis. The study confirms the previous results emphasizing the role of adverse metabolic changes in the radiation-associated IHD. Most importantly, it excludes age at the time of death as a confounding factor.
Assuntos
Isquemia Miocárdica/metabolismo , Plutônio/efeitos adversos , Proteoma/metabolismo , Proteoma/efeitos da radiação , Cromatografia Líquida , Citoesqueleto/metabolismo , Citoesqueleto/efeitos da radiação , Formaldeído/química , Humanos , Metabolismo dos Lipídeos/efeitos da radiação , Masculino , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Isquemia Miocárdica/epidemiologia , Isquemia Miocárdica/etiologia , Isquemia Miocárdica/patologia , Exposição Ocupacional , Inclusão em Parafina , Análise de Componente Principal , Mapas de Interação de Proteínas , Proteômica/métodos , Radiação Ionizante , Transdução de Sinais/efeitos da radiação , Sirtuínas/metabolismo , Espectrometria de Massas em Tandem , Fixação de TecidosRESUMO
Extracellular vesicles (EVs) are cell-derived membrane-bound organelles that have generated interest as they reflect the physiological condition of their source. Mass spectrometric (MS) analyses of protein cargo of EVs may lead to the discovery of biomarkers for diseases. However, for a comprehensive MS-based proteomics analysis, an optimal lysis of the EVs is required. Six methods for the protein extraction from EVs secreted by the head and neck cell line BHY were compared. Commercial radioimmunoprecipitation assay (RIPA) buffer outperformed the other buffers investigated in this study (Tris-SDS, Tris-Triton, GuHCl, urea-thiourea, and commercial Cell-lysis buffer). Following lysis with RIPA buffer, 310 proteins and 1469 peptides were identified using LTQ OrbitrapXL mass spectrometer. Among these, 86% of proteins and 72% of peptides were identified in all three replicates. In the case of other buffers, Tris-Triton identified on average 277 proteins, Cell-lysis buffer 257 proteins, and Tris-SDS, GuHCl and urea-thiourea each 267 proteins. In total, 399 proteins including 74 of the top EV markers (Exocarta) were identified, the most of the latter (73) using RIPA. The proteins exclusively identified using RIPA represented all Gene Ontology cell compartments. This study suggests that RIPA is an optimal lysis buffer for EVs in combination with MS.
Assuntos
Fracionamento Químico/métodos , Vesículas Extracelulares/metabolismo , Espectrometria de Massas , Proteoma/isolamento & purificação , Proteoma/metabolismo , Proteômica , Soluções Tampão , Linhagem Celular Tumoral , HumanosRESUMO
Chronic exposure to low-dose ionizing radiation is associated with an increased risk of cardiovascular disease. Alteration in energy metabolism has been suggested to contribute to radiation-induced heart pathology, mitochondrial dysfunction being a hallmark of this disease. The goal of this study was to investigate the regulatory role of acetylation in heart mitochondria in the long-term response to chronic radiation. ApoE-deficient C57Bl/6J mice were exposed to low-dose-rate (20 mGy/day) gamma radiation for 300 days, resulting in a cumulative total body dose of 6.0 Gy. Heart mitochondria were isolated and analyzed using quantitative proteomics. Radiation-induced proteome and acetylome alterations were further validated using immunoblotting, enzyme activity assays, and ELISA. In total, 71 proteins showed peptides with a changed acetylation status following irradiation. The great majority (94%) of the hyperacetylated proteins were involved in the TCA cycle, fatty acid oxidation, oxidative stress response and sirtuin pathway. The elevated acetylation patterns coincided with reduced activity of mitochondrial sirtuins, increased the level of Acetyl-CoA, and were accompanied by inactivation of major cardiac metabolic regulators PGC-1 alpha and PPAR alpha. These observations suggest that the changes in mitochondrial acetylation after irradiation is associated with impairment of heart metabolism. We propose a novel mechanism involved in the development of late cardiac damage following chronic irradiation.
Assuntos
Proteínas Mitocondriais/metabolismo , Miócitos Cardíacos/metabolismo , Processamento de Proteína Pós-Traducional , Sirtuínas/genética , Irradiação Corporal Total/efeitos adversos , Acetilação , Animais , Apolipoproteínas E/deficiência , Apolipoproteínas E/genética , Regulação para Baixo , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias Cardíacas/metabolismo , Mitocôndrias Cardíacas/efeitos da radiação , Proteínas Mitocondriais/efeitos da radiação , Miócitos Cardíacos/efeitos da radiação , PPAR alfa/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismoRESUMO
In children, ketamine sedation is often used during radiological procedures. Combined exposure of ketamine and radiation at doses that alone did not affect learning and memory induced permanent cognitive impairment in mice. The aim of this study was to elucidate the mechanism behind this adverse outcome. Neonatal male NMRI mice were administered ketamine (7.5 mg kg-1) and irradiated (whole-body, 100 mGy or 200 mGy, 137Cs) one hour after ketamine exposure on postnatal day 10. The control mice were injected with saline and sham-irradiated. The hippocampi were analyzed using label-free proteomics, immunoblotting, and Golgi staining of CA1 neurons six months after treatment. Mice co-exposed to ketamine and low-dose radiation showed alterations in hippocampal proteins related to neuronal shaping and synaptic plasticity. The expression of brain-derived neurotrophic factor, activity-regulated cytoskeleton-associated protein, and postsynaptic density protein 95 were significantly altered only after the combined treatment (100 mGy or 200 mGy combined with ketamine, respectively). Increased numbers of basal dendrites and branching were observed only after the co-exposure, thereby constituting a possible reason for the displayed alterations in behavior. These data suggest that the risk of radiation-induced neurotoxic effects in the pediatric population may be underestimated if based only on the radiation dose.
Assuntos
Região CA1 Hipocampal/patologia , Ketamina/toxicidade , Neurônios/patologia , Neurônios/efeitos da radiação , Radiação Ionizante , Animais , Animais Recém-Nascidos , Forma Celular/efeitos dos fármacos , Forma Celular/efeitos da radiação , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Citoesqueleto/efeitos da radiação , Masculino , Camundongos , Plasticidade Neuronal/efeitos dos fármacos , Plasticidade Neuronal/efeitos da radiação , Neurônios/efeitos dos fármacos , Proteoma/metabolismoRESUMO
High-dose ionizing radiation is known to induce adverse effects such as inflammation and fibrosis in the heart. Transcriptional regulators PPARα and TGFß are known to be involved in this radiation response. PPARα, an anti-inflammatory transcription factor controlling cardiac energy metabolism, is inactivated by irradiation. The pro-inflammatory and pro-fibrotic TGFß is activated by irradiation via SMAD-dependent and SMAD-independent pathways. The goal of this study was to investigate how altering the level of PPARα influences the radiation response of these signaling pathways. For this purpose, we used genetically modified C57Bl/6 mice with wild type (+/+), heterozygous (+/-) or homozygous (-/-) PPARα genotype. Mice were locally irradiated to the heart using doses of 8 or 16 Gy; the controls were sham-irradiated. The heart tissue was investigated using label-free proteomics 20 weeks after the irradiation and the predicted pathways were validated using immunoblotting, ELISA, and immunohistochemistry. The heterozygous PPARα mice showed most radiation-induced changes in the cardiac proteome, whereas the homozygous PPARα mice showed the least changes. Irradiation induced SMAD-dependent TGFß signaling independently of the PPARα status, but the presence of PPARα was necessary for the activation of the SMAD-independent pathway. These data indicate a central role of PPARα in cardiac response to ionizing radiation.
Assuntos
Coração/efeitos da radiação , Miocárdio/metabolismo , PPAR alfa/fisiologia , Fator de Crescimento Transformador beta/metabolismo , Animais , Genótipo , Heterozigoto , Camundongos , Camundongos Endogâmicos C57BL , Miocárdio/química , PPAR alfa/genética , Proteômica , Transdução de Sinais , Proteínas Smad/metabolismoRESUMO
Because of the increasing application of ionizing radiation in medicine, quantitative data on effects of low-dose radiation are needed to optimize radiation protection, particularly with respect to cataract development. Using mice as mammalian animal model, we applied a single dose of 0, 0.063, 0.125 and 0.5 Gy at 10 weeks of age, determined lens opacities for up to 2 years and compared it with overall survival, cytogenetic alterations and cancer development. The highest dose was significantly associated with increased body weight and reduced survival rate. Chromosomal aberrations in bone marrow cells showed a dose-dependent increase 12 months after irradiation. Pathological screening indicated a dose-dependent risk for several types of tumors. Scheimpflug imaging of the lens revealed a significant dose-dependent effect of 1% of lens opacity. Comparison of different biological end points demonstrated long-term effects of low-dose irradiation for several biological end points.
Assuntos
Catarata/genética , Lesões Experimentais por Radiação/genética , Animais , Catarata/etiologia , Aberrações Cromossômicas/efeitos da radiação , Relação Dose-Resposta à Radiação , Feminino , Estimativa de Kaplan-Meier , Masculino , Camundongos , Lesões Experimentais por Radiação/etiologia , Proteção Radiológica , Medição de Risco , Telômero/efeitos da radiação , Fatores de TempoRESUMO
Epidemiological data from patients undergoing radiotherapy for thoracic tumors clearly show the damaging effect of ionizing radiation on cardiovascular system. The long-term impairment of heart function and structure after local high-dose irradiation is associated with systemic inflammatory response, contraction impairment, microvascular damage, and cardiac fibrosis. The goal of the present study was to investigate molecular mechanisms involved in this process. C57BL/6J mice received a single X-ray dose of 16 Gy given locally to the heart at the age of 8 weeks. Radiation-induced changes in the heart transcriptome and proteome were investigated 40 weeks after the exposure. The omics data were analyzed by bioinformatics tools and validated by immunoblotting. Integrated network analysis of transcriptomics and proteomics data elucidated the signaling pathways that were similarly affected at gene and protein level. Analysis showed induction of transforming growth factor (TGF) beta signaling but inactivation of peroxisome proliferator-activated receptor (PPAR) alpha signaling in irradiated heart. The putative mediator role of mitogen-activated protein kinase cascade linking PPAR alpha and TGF beta signaling was supported by data from immunoblotting and ELISA. This study indicates that both signaling pathways are involved in radiation-induced heart fibrosis, metabolic disordering, and impaired contractility, a pathophysiological condition that is often observed in patients that received high radiation doses in thorax.
Assuntos
Fibrose Endomiocárdica/genética , Coração/efeitos da radiação , PPAR alfa/genética , Proteoma/genética , Transcriptoma , Fator de Crescimento Transformador beta/genética , Animais , Biologia Computacional , Fibrose Endomiocárdica/etiologia , Fibrose Endomiocárdica/metabolismo , Fibrose Endomiocárdica/patologia , Raios gama/efeitos adversos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Coração/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , PPAR alfa/metabolismo , Proteoma/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismoRESUMO
Radiation is the most common treatment of cancer. Minimizing the normal tissue injury, especially the damage to vascular endothelium, remains a challenge. This study aimed to analyze direct and indirect radiation effects on the endothelium by investigating mechanisms of signal transfer from irradiated to nonirradiated endothelial cells by means of secreted proteins. Human coronary artery endothelial cells (HCECest2) undergo radiation-induced senescence in vitro 14 days after exposure to 10 Gy X-rays. Proteomics analysis was performed on HCECest2 14 days after irradiation with X-ray doses of 0 Gy (control) or 10 Gy using label-free technology. Additionally, the proteomes of control and radiation-induced secretomes, and those of nonirradiated HCECest2 exposed for 24 h to secreted proteins of either condition were measured. Key changes identified by proteomics and bioinformatics were validated by immunoblotting, ELISA, bead-based multiplex assays, and targeted transcriptomics. The irradiated cells, their secretome, and the nonirradiated recipient cells showed similar inflammatory response, characterized by induction of interferon type I-related proteins and activation of the STAT3 pathway. These data indicate that irradiated endothelial cells may adversely affect nonirradiated surrounding cells via senescence-associated secretory phenotype. This study adds to our knowledge of the pathological background of radiation-induced cardiovascular disease.
Assuntos
Inflamação/genética , Neoplasias/radioterapia , Proteoma/genética , Radioterapia/efeitos adversos , Fator de Transcrição STAT3/genética , Senescência Celular/genética , Senescência Celular/efeitos da radiação , Vasos Coronários/metabolismo , Vasos Coronários/patologia , Vasos Coronários/efeitos da radiação , Relação Dose-Resposta à Radiação , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Células Endoteliais/efeitos da radiação , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Humanos , Inflamação/etiologia , Inflamação/patologia , Masculino , Neoplasias/complicações , Neoplasias/genética , Proteoma/efeitos da radiação , Proteômica/métodos , Transdução de Sinais/efeitos da radiaçãoRESUMO
INTRODUCTION: Epidemiological studies clearly show that thoracic or whole body exposure to ionizing radiation increases the risk of cardiac morbidity and mortality. Radiation-induced cardiovascular disease (CVD) has been intensively studied during the last ten years but the underlying molecular mechanisms are still poorly understood. Areas covered: Heart proteomics is a powerful tool holding promise for the future research. The central focus of this review is to compare proteomics data on radiation-induced CVD with data arising from proteomics of healthy and diseased cardiac tissue in general. In this context we highlight common and unique features of radiation-related and other heart pathologies. Future prospects and challenges of the field are discussed. Expert commentary: Data from comprehensive cardiac proteomics have deepened the knowledge of molecular mechanisms involved in radiation-induced cardiac dysfunction. State-of-the-art proteomics has the potential to identify novel diagnostic and therapeutic markers of this disease.
Assuntos
Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/metabolismo , Proteoma , Lesões por Radiação , Animais , Biomarcadores , Humanos , Proteômica , Radiação IonizanteRESUMO
BACKGROUND: The underlying molecular processes representing stress responses to low-dose ionising radiation (LDIR) in mammals are just beginning to be understood. In particular, LDIR effects on the brain and their possible association with neurodegenerative disease are currently being explored using omics technologies. RESULTS: We describe a light-weight approach for the storage, analysis and distribution of relevant LDIR omics datasets. The data integration platform, called BRIDE, contains information from the literature as well as experimental information from transcriptomics and proteomics studies. It deploys a hybrid, distributed solution using both local storage and cloud technology. CONCLUSIONS: BRIDE can act as a knowledge broker for LDIR researchers, to facilitate molecular research on the systems biology of LDIR response in mammals. Its flexible design can capture a range of experimental information for genomics, epigenomics, transcriptomics, and proteomics. The data collection is available at:
Assuntos
Encéfalo/efeitos da radiação , Radiação Ionizante , Pesquisa , Software , Relação Dose-Resposta à Radiação , HumanosRESUMO
Recent epidemiological data indicate that radiation doses as low as those used in computer tomography may result in long-term neurocognitive side effects. The aim of this study was to elucidate long-term molecular alterations related to memory formation in the brain after low and moderate doses of γ radiation. Female C57BL/6J mice were irradiated on postnatal day 10 with total body doses of 0.1, 0.5, or 2.0 Gy; the control group was sham-irradiated. The proteome analysis of hippocampus, cortex, and synaptosomes isolated from these brain regions indicated changes in ephrin-related, RhoGDI, and axonal guidance signaling. Immunoblotting and miRNA-quantification demonstrated an imbalance in the synapse morphology-related Rac1-Cofilin pathway and long-term potentiation-related cAMP response element-binding protein (CREB) signaling. Proteome profiling also showed impaired oxidative phosphorylation, especially in the synaptic mitochondria. This was accompanied by an early (4 weeks) reduction of mitochondrial respiration capacity in the hippocampus. Although the respiratory capacity was restored by 24 weeks, the number of deregulated mitochondrial complex proteins was increased at this time. All observed changes were significant at doses of 0.5 and 2.0 Gy but not at 0.1 Gy. This study strongly suggests that ionizing radiation at the neonatal state triggers persistent proteomic alterations associated with synaptic impairment.
Assuntos
Córtex Cerebral/efeitos da radiação , Raios gama/efeitos adversos , Hipocampo/efeitos da radiação , Potenciação de Longa Duração/efeitos da radiação , Proteoma/genética , Transmissão Sináptica/efeitos da radiação , Fatores de Despolimerização de Actina/genética , Fatores de Despolimerização de Actina/metabolismo , Animais , Animais Recém-Nascidos , Axônios/metabolismo , Axônios/efeitos da radiação , Axônios/ultraestrutura , Córtex Cerebral/metabolismo , Córtex Cerebral/fisiopatologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Complexo de Proteínas da Cadeia de Transporte de Elétrons/genética , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Efrinas/genética , Efrinas/metabolismo , Feminino , Hipocampo/metabolismo , Hipocampo/fisiopatologia , Memória/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Fosforilação Oxidativa/efeitos da radiação , Proteoma/metabolismo , Sinaptossomos/metabolismo , Sinaptossomos/efeitos da radiação , Irradiação Corporal Total , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo , Inibidores da Dissociação do Nucleotídeo Guanina rho-Específico/genética , Inibidores da Dissociação do Nucleotídeo Guanina rho-Específico/metabolismoRESUMO
The increased use of radiation-based medical imaging methods such as computer tomography is a matter of concern due to potential radiation-induced adverse effects. Efficient protection against such detrimental effects has not been possible due to inadequate understanding of radiation-induced alterations in signaling pathways. The aim of this study was to elucidate the molecular mechanisms behind learning and memory deficits after acute low and moderate doses of ionizing radiation. Female C57BL/6J mice were irradiated on postnatal day 10 (PND10) with gamma doses of 0.1 or 0.5 Gy. This was followed by evaluation of the cellular proteome, pathway-focused transcriptome, and neurological development/disease-focused miRNAome of hippocampus and cortex 24 h postirradiation. Our analysis showed that signaling pathways related to mitochondrial and synaptic functions were changed by acute irradiation. This may lead to reduced mitochondrial function paralleled by enhanced number of dendritic spines and neurite outgrowth due to elevated long-term potentiation, triggered by increased phosphorylated CREB. This was predominately observed in the cortex at 0.1 and 0.5 Gy and in the hippocampus only at 0.5 Gy. Moreover, a radiation-induced increase in the expression of several neural miRNAs associated with synaptic plasticity was found. The early changes in signaling pathways related to memory formation may be associated with the acute neurocognitive side effects in patients after brain radiotherapy but might also contribute to late radiation-induced cognitive injury.
Assuntos
Córtex Cerebral/efeitos da radiação , Hipocampo/efeitos da radiação , Potenciação de Longa Duração/efeitos da radiação , Memória/efeitos da radiação , Mitocôndrias/efeitos da radiação , Sinapses/efeitos da radiação , Animais , Proteína de Ligação a CREB/genética , Proteína de Ligação a CREB/metabolismo , Córtex Cerebral/fisiologia , Relação Dose-Resposta à Radiação , Feminino , Raios gama , Expressão Gênica , Hipocampo/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Fosforilação , Proteoma/metabolismo , Transdução de Sinais/efeitos da radiação , Sinapses/fisiologia , TranscriptomaRESUMO
Epidemiological data from radiotherapy patients show the damaging effect of ionizing radiation on heart and vasculature. The endothelium is the main target of radiation damage and contributes essentially to the development of cardiac injury. However, the molecular mechanisms behind the radiation-induced endothelial dysfunction are not fully understood. In the present study, 10-week-old C57Bl/6 mice received local X-ray heart doses of 8 or 16 Gy and were sacrificed after 16 weeks; the controls were sham-irradiated. The cardiac microvascular endothelial cells were isolated from the heart tissue using streptavidin-CD31-coated microbeads. The cells were lysed and proteins were labeled with duplex isotope-coded protein label methodology for quantification. All samples were analyzed by LC-ESI-MS/MS and Proteome Discoverer software. The proteomics data were further studied by bioinformatics tools and validated by targeted transcriptomics, immunoblotting, immunohistochemistry, and serum profiling. Radiation-induced endothelial dysfunction was characterized by impaired energy metabolism and perturbation of the insulin/IGF-PI3K-Akt signaling pathway. The data also strongly suggested premature endothelial senescence, increased oxidative stress, decreased NO availability, and enhanced inflammation as main causes of radiation-induced long-term vascular dysfunction. Detailed data on molecular mechanisms of radiation-induced vascular injury as compiled here are essential in developing radiotherapy strategies that minimize cardiovascular complications.