ryos because i) ionizing radiationinduced apoptosis has been extensively characterized in this specie, ii) 475110-96-4 pre-mid-blastula transition, but not post-MBT, irradiated embryos undergo apoptosis, iii) morphological and biochemical hallmarks of 6031788 apoptosis including the presence of membranebound apoptotic bodies, internucleosomal DNA fragmentation, pyknotic and condensed nuclei, loss of intracellular attachments, 8309351 caspase activation and cleavage of specific substrates have been established in this animal model, and iv) embryos are tractable to a number of manipulations. Our experiments focused on three specific scenarios. First, we evaluated the occurrence of apoptosis in embryos subjected to increasing incident energies during a fixed exposure time. Second, we monitored the response of the biological system when the exposure time was increased while a constant energy value was maintained. Lastly, we analyzed whether the embryos’ response to the same total dose of radiation varied as a result of increasing the energy while reducing the exposure time. A summary of the experimental conditions in each scenario is presented in Fig. Radiation Dosimetry Scenario A: Response of Embryos to Radiation Under Constant Exposure Time Our previous results showed that embryos irradiated at any stage before the MBT fail to repair the damage and irreversibly undergo cell death. Interestingly, irradiated embryos only exhibit apoptotic hallmarks after the MBT, suggesting that the MBT may be the first checkpoint monitoring developmental 
progression in early embryos. Initial experiments were devoted to determining the relevance of the incoming energy to cell fate in early embryos. Stage January Energy-Dependent Apoptosis Irradiated and control embryos were collected after irradiation at stage Scenario B: Response of Embryos to Radiation Under Constant Energy Values Next, we examined whether the length of the exposure time influenced the response of the embryo to a given radiation energy. Radiation from Energy-Dependent Apoptosis January Energy-Dependent Apoptosis January Energy-Dependent Apoptosis specifics are notable. First, embryos irradiated with energies between though caspase activity appeared to depend on the length of the exposure time. Cleavage of radiolabeled cyclin A January Energy-Dependent Apoptosis In agreement with the data summarized in Fig. Scenario C: Response of Embryos to Radiation Under Constant Total Dose Values Analysis of Cell Death Induced by Augmented Energy Levels We then employed an independent assay to test whether the apoptotic response that results from energy-dependent exposure correlates with DNA damage. With this in mind, we used a highly sensitive indicator of DNA fragmentation in situ, whole-mount TUNEL staining, a method that allows the detection of apoptotic cells at high frequency in early embryos. Accordingly, embryos were exposed to various energies and times for which the system is above or below a threshold response defined in our experimental sample. Treated embryos were analyzed for DNA fragmentation January Energy-Dependent Apoptosis January Energy-Dependent Apoptosis and mouse embryos as well. Whereas this event might be part of the normal developmental process in the embryo, the loss of a specific subset of cells as noticed by localized positive TUNEL-staining could well explain some of the phenotypic defects observed in a reduced number of tailbuds and revealed Discussion Our studies are the first direct demonstration
