RESUMEN
Centrosomes are important organelles for cell division and genome stability. Ionizing radiation exposure efficiently induces centrosome overduplication via the disconnection of the cell and centrosome duplication cycles. Over duplicated centrosomes cause mitotic catastrophe or chromosome aberrations, leading to cell death or tumorigenesis. Pluripotent stem cells, including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), can differentiate into all organs. To maintain pluripotency, PSCs show specific cellular dynamics, such as a short G1 phase and silenced cell-cycle checkpoints for high cellular proliferation. However, how exogenous DNA damage affects cell cycle-dependent centrosome number regulation in PSCs remains unknown. This study used human iPSCs (hiPSCs) derived from primary skin fibroblasts as a PSC model to address this question. hiPSCs derived from somatic cells could be a useful tool for addressing the radiation response in cell lineage differentiation. After radiation exposure, the hiPSCs showed a higher frequency of centrosome overduplication and multipolar cell division than the differentiated cells. To suppress the indirect effect of radiation exposure, we used the radical scavenger dimethyl sulfoxide (DMSO). Combined treatment with radiation and DMSO efficiently suppressed DNA damage and centrosome overduplication in hiPSCs. Our results will contribute to the understanding of the dynamics of stem cells and the assessment of the risk of genome instability for regenerative medicine.
Asunto(s)
División Celular , Centrosoma , Dimetilsulfóxido , Células Madre Pluripotentes Inducidas , Humanos , Centrosoma/efectos de la radiación , Centrosoma/efectos de los fármacos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Células Madre Pluripotentes Inducidas/efectos de la radiación , Dimetilsulfóxido/farmacología , División Celular/efectos de la radiación , División Celular/efectos de los fármacos , Daño del ADN , Radiación Ionizante , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/efectos de la radiaciónRESUMEN
5-Aminolevulinic acid (5-ALA) photodynamic therapy (PDT) is a treatment for actinic keratosis (AK) and has been studied as a treatment for noninvasive cutaneous squamous cell carcinoma (cSCC). PDT induces apoptosis and necrosis in AKs and cSCC. 5-ALA blue light PDT may modulate gene expression and pathways in surviving cells. In this study, differential gene expression and pathway analysis of cSCC and human dermal fibroblasts were compared before and after 5-ALA blue light PDT using RNA sequencing. No genes were differentially expressed after correcting for multiple testing (false discovery rate < 0.05). As a result, transcription factor, gene enrichment, and pathway analysis were performed with genes identified before multiple testing (p < 0.05). Pathways associated with proliferation and carcinogenesis were downregulated. These findings using 5-ALA blue light PDT are similar to previously published studies using methyl-aminolevulinic and red light protocols, indicating that surviving residual cells may undergo changes consistent with a less aggressive cancerous phenotype.
Asunto(s)
Ácido Aminolevulínico , Carcinoma de Células Escamosas , Regulación hacia Abajo , Fotoquimioterapia , Neoplasias Cutáneas , Humanos , Ácido Aminolevulínico/farmacología , Luz Azul , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/patología , División Celular/efectos de los fármacos , División Celular/efectos de la radiación , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Regulación hacia Abajo/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Fármacos Fotosensibilizantes/farmacología , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/patologíaRESUMEN
In this study we demonstrated that exposure of Escherichia coli (E. coli) to terahertz (THz) radiation resulted in a change in the activities of the tdcABCDEFGR and matA-F genes (signs of cell aggregation), gene yjjQ (signs of suppression of cell motility), dicABCF, FtsZ, and minCDE genes (signs of suppression of cell division), sfmACDHF genes (signs of adhesin synthesis), yjbEFGH and gfcA genes (signs of cell envelope stabilization). Moreover, THz radiation induced E. coli csg operon genes of amyloid biosynthesis. Electron microscopy revealed that the irradiated bacteria underwent increased aggregation; 20% of them formed bundle-like structures consisting of two to four pili clumped together. This could be the result of changes in the adhesive properties of the pili. We also found aberrations in cell wall structure in the middle part of the bacterial cell; these aberrations impaired the cell at the initial stages of division and resulted in accumulation of long rod-like cells. Overall, THz radiation was shown to have adverse effects on bacterial populations resulting in cells with abnormal morphology.
Asunto(s)
Agregación Celular/efectos de la radiación , División Celular/efectos de la radiación , Escherichia coli/efectos de la radiación , Radiación Terahertz , Pared Celular/efectos de la radiación , Escherichia coli/citología , Escherichia coli/metabolismo , Regulación Bacteriana de la Expresión Génica/efectos de la radiación , Microscopía Electrónica , Operón/genéticaRESUMEN
Biological phenomena induced by terahertz (THz) irradiation are described in recent reports, but underlying mechanisms, structural and dynamical change of specific molecules are still unclear. In this paper, we performed time-lapse morphological analysis of human cells and found that THz irradiation halts cell division at cytokinesis. At the end of cytokinesis, the contractile ring, which consists of filamentous actin (F-actin), needs to disappear; however, it remained for 1 hour under THz irradiation. Induction of the functional structures of F-actin was also observed in interphase cells. Similar phenomena were also observed under chemical treatment (jasplakinolide), indicating that THz irradiation assists actin polymerization. We previously reported that THz irradiation enhances the polymerization of purified actin in vitro; our current work shows that it increases cytoplasmic F-actin in vivo. Thus, we identified one of the key biomechanisms affected by THz waves.
Asunto(s)
Actinas/efectos de la radiación , División Celular/efectos de la radiación , Radiación Terahertz , Actinas/metabolismo , Citocinesis/efectos de la radiación , Células HeLa/efectos de la radiación , Humanos , Interfase/efectos de la radiación , Microscopía Fluorescente , Análisis de la Célula IndividualRESUMEN
Ciprofloxacin hydrochloride and Norfloxacin are second-generation fluoroquinolone antibiotic against bacterial DNA gyrase, which reduces DNA strain throughout replication. As DNA gyrase is essential through DNA replication, subsequent DNA synthesis and cell division are inhibited. Direct photolysis of fluoroquinolones was studied by using UV irradiation in the presence or absence of other substances that generate free radicals. This study aimed to assess the effect of Ultraviolet B (UVB) irradiation in removing ciprofloxacin and norfloxacin by using a simulating model of wastewater contained urea at pH 4. A known concentration of ciprofloxacin and norfloxacin were prepared in an appropriate aqueous solution in presence or absence 0.2M urea and adjusted at pH 4. The dis-solved drugs were irradiated with UVB-lamp in a dark place for 60 minutes. The percent of removal and the rate of elimination (k) of each drug were calculated. The direct photolysis effect of UVB irradiation was observed with ciprofloxacin which amounted to 24.4% removal compared with12.4% removal of norfloxacin after 60 minutes of irradiation. The effect of UVB irradiation was enhanced by urea to reach 38.9% and 15% for ciprofloxacin and norfloxacin. The calculated k of ciprofloxacin has amounted to three folds of that of norfloxacin. Direct photolysis of ciprofloxacin and norfloxacin can be achieved simply by using a simulation model of 0.2 M urea and UVB irradiation at pH 4. UVB is highly effective in removing ciprofloxacin compared with norfloxacin by 2-3 folds.
Asunto(s)
División Celular/efectos de los fármacos , Ciprofloxacina/farmacología , Replicación del ADN/efectos de los fármacos , ADN Bacteriano/efectos de los fármacos , Norfloxacino/farmacología , Rayos Ultravioleta , Urea/química , División Celular/efectos de la radiación , Ciprofloxacina/efectos de la radiación , Medios de Cultivo , Replicación del ADN/efectos de la radiación , ADN Bacteriano/efectos de la radiación , Norfloxacino/efectos de la radiación , Análisis de RegresiónRESUMEN
The effect of low-dose-rate exposure to ionizing radiation on cancer risk is a major issue associated with radiation protection. Tissue stem cells are regarded as one of the targets of radiation-induced carcinogenesis. However, it is hypothesized that the effect of radiation may be reduced if damaged stem cells are eliminated via stem cell competition between damaged and intact stem cells. This would be particularly effective under very low-dose-rate conditions, in which only a few stem cells in a stem cell pool may be affected by radiation. Following this hypothesis, we constructed a simple mathematical model to discuss the influence of stem cell competition attenuating the accumulation of damaged cells under very low-dose-rate conditions. In this model, a constant number of cells were introduced into a cell pool, and the numbers of intact and damaged cells were calculated via transition and turnover events. A transition event emulates radiation dose, whereby an intact cell is changed into a damaged cell with a given probability. On the other hand, a turnover event expresses cell competition, where reproduction and elimination of cells occur depending on the properties of cells. Under very low-dose-rate conditions, this model showed that radiation damage to the stem cell pool was strongly suppressed when the damaged cells were less reproductive and tended to be eliminated compared to the intact cells. Furthermore, the size of the stem cell pool was positively correlated with reduction in radiation damage.
Asunto(s)
Simulación por Computador , Modelos Biológicos , Células Madre/efectos de la radiación , División Celular/efectos de la radiación , Relación Dosis-Respuesta en la RadiaciónRESUMEN
Both SIRT1 and UVA radiation are involved in cellular damage processes such as apoptosis, senescence and ageing. MicroRNAs (miRNAs) have been reported to be closely related to UV radiation, as well as to SIRT1. In this study, we investigated the connections among SIRT1, UVA and miRNA in human skin primary fibroblasts. Our results showed that UVA altered the protein level of SIRT1 in a time point-dependent manner. Using miRNA microarray, bioinformatics analysis, we found that knocking down SIRT1 could cause up-regulation of miR-27a-5p and the latter could down-regulate SMAD2, and these results were verified by qRT-PCR or Western blot. Furthermore, UVA radiation (5 J/cm2 ), knocking down SIRT1 or overexpression of miR-27a-5p led to increased expression of MMP1, and decreased expressions of COL1 and BCL2. We also found additive impacts on MMP1, COL1 and BCL2 under the combination of UVA radiation + Sirtinol (SIRT1 inhibitor), or UVA radiation + miR-27a-5p mimic. SIRT1 activator resveratrol could reverse damage changes caused by UVA radiation. Besides, absent of SIRT1 or overexpression of miR-27a-5p increased cell apoptosis and induced cell arrest in G2/M phase. Taken together, these results demonstrated that UVA could influence a novel SIRT1-miR-27a-5p-SMAD2-MMP1/COL1/BCL2 axis in skin primary fibroblasts, and may provide potential therapeutic targets for UVA-induced skin damage.
Asunto(s)
Fibroblastos/metabolismo , Fibroblastos/efectos de la radiación , Proteínas/metabolismo , Transducción de Señal/efectos de la radiación , Piel/metabolismo , Piel/efectos de la radiación , Rayos Ultravioleta/efectos adversos , Adolescente , Adulto , Apoptosis/efectos de la radiación , Puntos de Control del Ciclo Celular/efectos de la radiación , División Celular/efectos de la radiación , Células Cultivadas , Regulación hacia Abajo/efectos de la radiación , Fase G2/efectos de la radiación , Humanos , Regulación hacia Arriba/efectos de la radiación , Adulto JovenRESUMEN
Cell division can perturb the metabolic performance of industrial microbes. The C period of cell division starts from the initiation to the termination of DNA replication, whereas the D period is the bacterial division process. Here, we first shorten the C and D periods of E. coli by controlling the expression of the ribonucleotide reductase NrdAB and division proteins FtsZA through blue light and near-infrared light activation, respectively. It increases the specific surface area to 3.7 µm-1 and acetoin titer to 67.2 g·L-1. Next, we prolong the C and D periods of E. coli by regulating the expression of the ribonucleotide reductase NrdA and division protein inhibitor SulA through blue light activation-repression and near-infrared (NIR) light activation, respectively. It improves the cell volume to 52.6 µm3 and poly(lactate-co-3-hydroxybutyrate) titer to 14.31 g·L-1. Thus, the optogenetic-based cell division regulation strategy can improve the efficiency of microbial cell factories.
Asunto(s)
División Celular/efectos de la radiación , Escherichia coli/citología , Escherichia coli/efectos de la radiación , Luz , Acetoína/metabolismo , Reactores Biológicos/microbiología , Escherichia coli/genética , Escherichia coli/ultraestructura , Genes Bacterianos , Poliésteres/metabolismoRESUMEN
The cell fusion is a widespread process, which takes place in many systems in vivo and in vitro. Fusion of cells is frequently related to tetraploidy, which can be found within natural physiological conditions, e.g., placentation, and in pathophysiological conditions, such as cancer and early pregnancy failure in humans. Here we investigate the mechanism of tetraploidization with help of femtosecond laser-induced mouse blastomere fusion by the means of Hoechst staining, GFP, BODIPY dyes and fluorescent species generated intracellularly by a femtosecond laser. We establish diffusive mixing of cytosol, whereas the large components of a cytoplasm (organelles, cytoskeleton) are poorly diffusible and are not completely mixed after cell fusion and a subsequent division. We show that mechanisms which are responsible for the formation of a common metaphase plate triggered tetraploidization in fused mouse embryos and could be a significant factor in polyploidy formation in vivo. Thus, our results suggest that microtubules play a critical role in tetraploidization.
Asunto(s)
Blastómeros/fisiología , Blastómeros/efectos de la radiación , Rayos Láser , Tetraploidía , Animales , Blastómeros/citología , División Celular/efectos de la radiación , Fusión Celular/métodos , Embrión de Mamíferos/citología , Embrión de Mamíferos/efectos de la radiación , Femenino , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Masculino , Metafase/fisiología , Metafase/efectos de la radiación , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , EmbarazoRESUMEN
Light-regulated modules offer unprecedented new ways to control cellular behaviour with precise spatial and temporal resolution. Among a variety of bacterial light-switchable gene expression systems, single-component systems consisting of single transcription factors would be more useful due to the advantages of speed, simplicity, and versatility. In the present study, we developed a single-component light-activated bacterial gene expression system (eLightOn) based on a novel LOV domain from Rhodobacter sphaeroides (RsLOV). The eLightOn system showed significant improvements over the existing single-component bacterial light-activated expression systems, with benefits including a high ON/OFF ratio of >500-fold, a high activation level, fast activation kinetics, and/or good adaptability. Additionally, the induction characteristics, including regulatory windows, activation kinetics and light sensitivities, were highly tunable by altering the expression level of LexRO. We demonstrated the usefulness of the eLightOn system in regulating cell division and swimming by controlling the expression of the FtsZ and CheZ genes, respectively, as well as constructing synthetic Boolean logic gates using light and arabinose as the two inputs. Taken together, our data indicate that the eLightOn system is a robust and highly tunable tool for quantitative and spatiotemporal control of bacterial gene expression.
Asunto(s)
Regulación Bacteriana de la Expresión Génica/efectos de la radiación , Luz , Rhodobacter sphaeroides/citología , Rhodobacter sphaeroides/efectos de la radiación , Proteínas Bacterianas/metabolismo , División Celular/efectos de la radiación , Cinética , Lógica , Factores de Transcripción/metabolismoRESUMEN
All organisms regulate cell cycle progression by coordinating cell division with DNA replication status. In eukaryotes, DNA damage or problems with replication fork progression induce the DNA damage response (DDR), causing cyclin-dependent kinases to remain active, preventing further cell cycle progression until replication and repair are complete. In bacteria, cell division is coordinated with chromosome segregation, preventing cell division ring formation over the nucleoid in a process termed nucleoid occlusion. In addition to nucleoid occlusion, bacteria induce the SOS response after replication forks encounter DNA damage or impediments that slow or block their progression. During SOS induction, Escherichia coli expresses a cytoplasmic protein, SulA, that inhibits cell division by directly binding FtsZ. After the SOS response is turned off, SulA is degraded by Lon protease, allowing for cell division to resume. Recently, it has become clear that SulA is restricted to bacteria closely related to E. coli and that most bacteria enforce the DNA damage checkpoint by expressing a small integral membrane protein. Resumption of cell division is then mediated by membrane-bound proteases that cleave the cell division inhibitor. Further, many bacterial cells have mechanisms to inhibit cell division that are regulated independently from the canonical LexA-mediated SOS response. In this review, we discuss several pathways used by bacteria to prevent cell division from occurring when genome instability is detected or before the chromosome has been fully replicated and segregated.
Asunto(s)
División Celular/efectos de los fármacos , División Celular/efectos de la radiación , Replicación del ADN/efectos de los fármacos , Replicación del ADN/efectos de la radiación , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Bleomicina/farmacología , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Ciclo Celular/efectos de la radiación , División Celular/genética , Daño del ADN/efectos de los fármacos , Daño del ADN/efectos de la radiación , Replicación del ADN/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/efectos de la radiación , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Proteasa La/genética , Proteasa La/metabolismo , Radiación Ionizante , Respuesta SOS en Genética/efectos de los fármacos , Respuesta SOS en Genética/genética , Respuesta SOS en Genética/efectos de la radiaciónRESUMEN
Purpose: Accumulated damage in neural stem cells (NSCs) during brain tumor radiotherapy causes cognitive dysfunction to the patients. Carbon-ion radiotherapy can reduce undesired irradiation of normal tissues more efficiently than conventional photon radiotherapy. This study elucidates the responses of NSCs to carbon-ion radiation.Methods: Human NSCs and glioblastoma A-172 cells were irradiated with carbon-ion radiation and γ-rays, which have different linear-energy-transfer (LET) values of 108 and 0.2 keV/µm, respectively. After irradiation, growth rates were measured, apoptotic cells were detected by flow cytometry, and DNA synthesizing cells were immunocytochemically visualized.Results: Growth rates of NSCs and A-172 cells were decreased after irradiation. The percentages of apoptotic cells were remarkably increased in NSCs but not in A-172 cells. In contrast, the fractions of DNA synthesizing A-172 cells were decreased in a dose-dependent manner. These results indicate that apoptosis induction and DNA synthesis inhibition contribute to the growth inhibition of NSCs and glioblastoma cells, respectively. In addition, high-LET carbon ions induced more profound effects than low-LET γ-rays.Conclusions: Apoptosis is an important clinical target to protect NSCs during brain tumor radiotherapy using carbon-ion radiation as well as conventional X-rays.
Asunto(s)
Apoptosis/efectos de la radiación , Neoplasias Encefálicas/radioterapia , Rayos gamma , Glioblastoma/radioterapia , Radioterapia de Iones Pesados/métodos , Células-Madre Neurales/efectos de la radiación , Biomarcadores/metabolismo , Carbono , División Celular/efectos de la radiación , Línea Celular Tumoral , Supervivencia Celular/efectos de la radiación , ADN/efectos de la radiación , Daño del ADN , Relación Dosis-Respuesta en la Radiación , Humanos , Inmunohistoquímica , Iones , Transferencia Lineal de Energía , Nestina/metabolismo , Fotones , Factores de Transcripción SOXB1/metabolismoRESUMEN
Temporal programs synchronised with the daily cycle are of adaptive importance for organisms exposed to periodic fluctuations. This study deepens into several aspects of the exogenous and endogenous nature of microbial grazers. We investigated the diel rhythms of cell division and feeding activity of four marine protists under different light regimes. In particular, we tested if the feeding cycle of protistan grazers could be mediated by a light-aided enhancement of prey digestion, and also explored the consequences of cell division on diel feeding rhythms. Cell division occurred at night for the heterotrophic dinoflagellates Gyrodinium dominans and Oxyrrhis marina. In contrast, the mixotrophic dinoflagellate Karlodinium armiger and the ciliate Strombidium sp. mostly divided during the day. Additionally, a significant diurnal feeding rhythm was observed in all species. When exposed to continuous darkness, nearly all species maintained the cell division rhythm, but lost the feeding cycle within several hours/days (with the exception of O. marina that kept the rhythm for 9.5 days). Additional feeding experiments under continuous light also showed the same pattern. We conclude that the feeding rhythms of protistan grazers are generally regulated not by cell division nor by the enhancement of digestion by light. Our study, moreover, indicates that the cell division cycle is under endogenous control, whereas an external trigger is required to maintain the feeding rhythm, at least for most of the species studied here.
Asunto(s)
Cilióforos/fisiología , Dinoflagelados/fisiología , División Celular/efectos de la radiación , Cilióforos/efectos de la radiación , Dinoflagelados/efectos de la radiación , Procesos Heterotróficos , LuzRESUMEN
The factors and signals driving T cell activation and polarisation during immune responses have been studied mainly at the level of cells and chemical mediators. Here we describe a physical driver of these processes in the form of physiological-strength electric fields (EFs). EFs are generated at sites where epithelium is disrupted (e.g. wounded skin/bronchial epithelia) and where T cells frequently are present. Using live-cell imaging, we show human primary T cells migrate directionally to the cathode in low strength (50/150 mV/mm) EFs. Strikingly, we show for the first time that EFs significantly downregulate T cell activation following stimulation with antigen-activated APCs or anti-CD3/CD28 antibodies, as demonstrated by decreased IL-2 secretion and proliferation. These EF-induced functional changes were accompanied by a significant dampening of CD4+ T cell polarisation. Expression of critical markers of the Th17 lineage, RORγt and IL-17, and the Th17 polarisation mediator phospho-STAT3 were reduced significantly, while STAT1, ERK and c-Jun phosphorylation were comparatively unaffected suggesting STAT3 modulation by EFs as one mechanism driving effects. Overall, we identify electrical signals as important contributors to the co-ordination and regulation of human T cell functions, paving the way for a new research area into effects of naturally occurring and clinically-applied EFs in conditions where control of T cell activity is paramount.
Asunto(s)
Campos Electromagnéticos , Activación de Linfocitos/efectos de la radiación , Subgrupos de Linfocitos T/efectos de la radiación , Células Presentadoras de Antígenos/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/efectos de la radiación , División Celular/efectos de la radiación , Movimiento Celular , Polaridad Celular/efectos de la radiación , Células Cultivadas , Citocinas/biosíntesis , Electrodos , Endotoxinas/farmacología , Humanos , Interleucina-2/biosíntesis , Activación de Linfocitos/efectos de los fármacos , Fosforilación , Procesamiento Proteico-Postraduccional , Factor de Transcripción STAT3/metabolismo , Subgrupos de Linfocitos T/efectos de los fármacos , Subgrupos de Linfocitos T/inmunología , Células Th17/inmunología , Células Th17/efectos de la radiaciónRESUMEN
Studies on UV-B-induced plant photomorphogenesis mainly focus on Arabidopsis shoots (hypocotyl, leaf, petiole, and stem) but less on roots. In the present research, the low-level UV-B (0.2 W·m-2) induced a decrease in the number of root cells in the meristem zone and an inhibition of the cell length in the maturation zone of roots in Arabidopsis thaliana L.Heynh (Col-0). UV-B-induced root growth inhibition was recovered by the addition of GA3 to culture media. GA3 played an important role in UV-B-induced inhibition of root growth. The cop1-4 mutant with more meristem cell and longer mature cells exhibited longer root length under low-level UV-B. COP1 acted as a positive regulator of root growth under UV-B, through regulation of cell division and elongation. The sto mutant exhibited a shorter root length under UV-B with similar cell length but fewer meristem cells compared with wild type (Col-0). STO only regulated cell division, but cell expansion was not affected. UV-B radiation also inhibited the root growth of uvr8 mutant, and the degree of inhibition was greater than for wild type (Ler). UV-B inhibited the growth of Arabidopsis root, possibly because it changes the GA signal and inhibited cell division and cell elongation, which be related to COP1 and STO genes.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Arabidopsis/efectos de la radiación , Giberelinas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/efectos de la radiación , Rayos Ultravioleta , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , División Celular/efectos de los fármacos , División Celular/efectos de la radiación , Proteínas Cromosómicas no Histona/metabolismo , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Reguladores del Crecimiento de las Plantas/farmacología , Raíces de Plantas/citología , Raíces de Plantas/efectos de los fármacos , Plantones/efectos de los fármacos , Plantones/crecimiento & desarrollo , Plantones/efectos de la radiación , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Charged-particle microbeams (CPMs) provide a unique opportunity to investigate the effects of ionizing radiation on living biological specimens with a precise control of the delivered dose, i.e. the number of particles per cell. We describe a methodology to manipulate and micro-irradiate early stage C. elegans embryos at a specific phase of the cell division and with a controlled dose using a CPM. To validate this approach, we observe the radiation-induced damage, such as reduced cell mobility, incomplete cell division and the appearance of chromatin bridges during embryo development, in different strains expressing GFP-tagged proteins in situ after irradiation. In addition, as the dosimetry of such experiments cannot be extrapolated from random irradiations of cell populations, realistic three-dimensional models of 2 cell-stage embryo were imported into the Geant4 Monte-Carlo simulation toolkit. Using this method, we investigate the energy deposit in various chromatin condensation states during the cell division phases. The experimental approach coupled to Monte-Carlo simulations provides a way to selectively irradiate a single cell in a rapidly dividing multicellular model with a reproducible dose. This method opens the way to dose-effect investigations following targeted irradiation.
Asunto(s)
Caenorhabditis elegans/efectos de la radiación , Embrión no Mamífero/efectos de la radiación , Animales , Caenorhabditis elegans/embriología , Caenorhabditis elegans/ultraestructura , División Celular/efectos de la radiación , Cromatina/efectos de la radiación , Cromosomas/efectos de la radiación , Embrión no Mamífero/ultraestructura , Desarrollo Embrionario/efectos de la radiación , Microscopía Confocal/métodos , Método de Montecarlo , RadiometríaRESUMEN
Bone provides supportive microenvironments for hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) and is a frequent site of metastasis. While incidences of bone metastases increase with age, the properties of the bone marrow microenvironment that regulate dormancy and reactivation of disseminated tumor cells (DTCs) remain poorly understood. Here, we elucidate the age-associated changes in the bone secretome that trigger proliferation of HSCs, MSCs, and DTCs in the aging bone marrow microenvironment. Remarkably, a bone-specific mechanism involving expansion of pericytes and induction of quiescence-promoting secretome rendered this proliferative microenvironment resistant to radiation and chemotherapy. This bone-specific expansion of pericytes was triggered by an increase in PDGF signaling via remodeling of specialized type H blood vessels in response to therapy. The decline in bone marrow pericytes upon aging provides an explanation for loss of quiescence and expansion of cancer cells in the aged bone marrow microenvironment. Manipulation of blood flow - specifically, reduced blood flow - inhibited pericyte expansion, regulated endothelial PDGF-B expression, and rendered bone metastatic cancer cells susceptible to radiation and chemotherapy. Thus, our study provides a framework to recognize bone marrow vascular niches in age-associated increases in metastasis and to target angiocrine signals in therapeutic strategies to manage bone metastasis.
Asunto(s)
Envejecimiento/patología , Médula Ósea/patología , Neoplasias Óseas/terapia , Microambiente Tumoral/fisiología , Antagonistas de Receptores Adrenérgicos alfa 1/administración & dosificación , Animales , Antineoplásicos/administración & dosificación , Médula Ósea/irrigación sanguínea , Médula Ósea/efectos de los fármacos , Médula Ósea/efectos de la radiación , Neoplasias Óseas/irrigación sanguínea , Neoplasias Óseas/secundario , División Celular/efectos de los fármacos , División Celular/efectos de la radiación , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Resistencia a Antineoplásicos/fisiología , Células Madre Hematopoyéticas/efectos de los fármacos , Células Madre Hematopoyéticas/patología , Células Madre Hematopoyéticas/efectos de la radiación , Humanos , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/patología , Células Madre Mesenquimatosas/efectos de la radiación , Ratones , Pericitos/efectos de los fármacos , Pericitos/patología , Pericitos/efectos de la radiación , Prazosina/administración & dosificación , Tolerancia a Radiación/fisiología , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/efectos de la radiación , Irradiación Corporal Total , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Using data generated with cells exposed to ionizing-radiation (IR) in G2-phase of the cell cycle, we describe dose-dependent interactions between ATM, ATR and DNA-PKcs revealing unknown mechanistic underpinnings for two key facets of the DNA damage response: DSB end-resection and G2-checkpoint activation. At low IR-doses that induce low DSB-numbers in the genome, ATM and ATR regulate epistatically the G2-checkpoint, with ATR at the output-node, interfacing with the cell-cycle predominantly through Chk1. Strikingly, at low IR-doses, ATM and ATR epistatically regulate also resection, and inhibition of either activity fully suppresses resection. At high IR-doses that induce high DSB-numbers in the genome, the tight ATM/ATR coupling relaxes and independent outputs to G2-checkpoint and resection occur. Consequently, both kinases must be inhibited to fully suppress checkpoint activation and resection. DNA-PKcs integrates to the ATM/ATR module by regulating resection at all IR-doses, with defects in DNA-PKcs causing hyper-resection and G2-checkpoint hyper-activation. Notably, hyper-resection is absent from other c-NHEJ mutants. Thus, DNA-PKcs specifically regulates resection and adjusts the activation of the ATM/ATR module. We propose that selected DSBs are shepherd by DNA-PKcs from c-NHEJ to resection-dependent pathways for processing under the regulatory supervision of the ATM/ATR module.
Asunto(s)
Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteína Quinasa Activada por ADN/genética , Puntos de Control de la Fase G2 del Ciclo Celular/efectos de la radiación , Radiación Ionizante , Ciclo Celular/genética , Ciclo Celular/efectos de la radiación , División Celular/genética , División Celular/efectos de la radiación , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/genética , Roturas del ADN de Doble Cadena/efectos de la radiación , Daño del ADN/genética , Daño del ADN/efectos de la radiación , Reparación del ADN por Unión de Extremidades/efectos de la radiación , Proteínas de Unión al ADN/genética , Puntos de Control de la Fase G2 del Ciclo Celular/genética , Humanos , Fosforilación/efectos de la radiación , Transducción de Señal/efectos de la radiaciónRESUMEN
Gastrointestinal toxicity limits the clinical application of abdominal and pelvic radiotherapy and currently has no effective treatment. Intestinal leucine-rich-repeat-containing GPCR 5 (Lgr5)-positive stem cell depletion and loss of proliferative ability due to radiation may be the primary factors causing intestinal injury following radiation. Here, we report the critical role of ß-arrestin1 (ßarr1) in radiation-induced intestinal injury. Intestinal ßarr1 was highly expressed in radiation enteritis and in a radiation model. ßarr1 knockout (KO) or knockdown mice exhibited increased proliferation in intestinal Lgr5+ stem cell, crypt reproduction, and survival following radiation. Unexpectedly, the beneficial effects of ßarr1 deficiency on intestinal stem cells in response to radiation were compromised when the endoplasmic reticulum stress-related protein kinase RNA-like ER kinase (PERK)/eukaryotic initiation factor-2α (eIF2α) pathway was inhibited, and this result was further supported in vitro. Furthermore, we found that ßarr1 knockdown with small interfering RNA significantly enhanced intestinal Lgr5+ stem cell proliferation after radiation via directly targeting PERK. ßarr1 offers a promising target for mitigating radiation-induced intestinal injury.-Liu, Z., Jiang, J., He, Q., Liu, Z., Yang, Z., Xu, J., Huang, Z., Wu, B. ß-Arrestin1-mediated decrease in endoplasmic reticulum stress impairs intestinal stem cell proliferation following radiation.
Asunto(s)
Estrés del Retículo Endoplásmico/fisiología , Enteritis/patología , Intestinos/efectos de la radiación , Traumatismos Experimentales por Radiación/patología , Traumatismos por Radiación/patología , Células Madre/efectos de la radiación , beta-Arrestina 1/fisiología , eIF-2 Quinasa/fisiología , Anciano , Animales , División Celular/efectos de la radiación , Ensayo de Unidades Formadoras de Colonias , Enteritis/etiología , Enteritis/fisiopatología , Factor 2 Eucariótico de Iniciación/fisiología , Femenino , Técnicas de Silenciamiento del Gen , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , Interferencia de ARN , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/farmacología , Quimera por Radiación , Traumatismos por Radiación/fisiopatología , Traumatismos Experimentales por Radiación/fisiopatología , Radioterapia/efectos adversos , Receptores Acoplados a Proteínas G/análisis , Regeneración , Transducción de Señal/fisiología , Células Madre/patología , beta-Arrestina 1/deficiencia , beta-Arrestina 1/genéticaRESUMEN
Telomeres are essential for genome stability. Oxidative stress caused by excess reactive oxygen species (ROS) accelerates telomere shortening. Although telomeres are hypersensitive to ROS-mediated 8-oxoguanine (8-oxoG) formation, the biological effect of this common lesion at telomeres is poorly understood because ROS have pleiotropic effects. Here we developed a chemoptogenetic tool that selectively produces 8-oxoG only at telomeres. Acute telomeric 8-oxoG formation increased telomere fragility in cells lacking OGG1, the enzyme that removes 8-oxoG, but did not compromise cell survival. However, chronic telomeric 8-oxoG induction over time shortens telomeres and impairs cell growth. Accumulation of telomeric 8-oxoG in chronically exposed OGG1-deficient cells triggers replication stress, as evidenced by mitotic DNA synthesis at telomeres, and significantly increases telomere losses. These losses generate chromosome fusions, leading to chromatin bridges and micronucleus formation upon cell division. By confining base damage to the telomeres, we show that telomeric 8-oxoG accumulation directly drives telomere crisis.