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Large Field Alpha Irradiation Setup for Radiobiological Experiments.

ABSTRACT: The use of alpha particles irradiation in clinical practice has gained interest in the past years, for example with the advance of radionuclide therapy. The lack of affordable and easily accessible irradiation systems to study the cell biological impact of alpha particles hampers broad investigation. Here we present a novel alpha particle irradiation set-up for uniform irradiation of cell cultures. By combining a small alpha emitting source and a computer-directed movement stage, we established a new alpha particle irradiation method allowing more advanced biological assays, including large-field local alpha particle irradiation and cell survival assays. In addition, this protocol uses cell culture on glass cover-slips which allows more advanced microscopy, such as super-resolution imaging, for in-depth analysis of the DNA damage caused by alpha particles. This novel irradiation set-up provides the possibility to perform reproducible, uniform and directed alpha particle irradiation to investigate the impact of alpha radiation on the cellular level.

SUBMITTER: Roobol SJ

PROVIDER: S-EPMC6789741 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Large Field Alpha Irradiation Setup for Radiobiological Experiments.

Roobol Stefan J SJ Kouwenberg Jasper J M JJM Denkova Antonia G AG Kanaar Roland R Essers Jeroen J

Methods and protocols 20190828 3

The use of alpha particles irradiation in clinical practice has gained interest in the past years, for example with the advance of radionuclide therapy. The lack of affordable and easily accessible irradiation systems to study the cell biological impact of alpha particles hampers broad investigation. Here we present a novel alpha particle irradiation set-up for uniform irradiation of cell cultures. By combining a small alpha emitting source and a computer-directed movement stage, we established ...[more]

PMID: 31466405

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Project description:BackgroundAlthough two-dimensional (2-D) monolayer cell cultures provide important information on basic tumor biology and radiobiology, they are not representative of the complexity of three-dimensional (3-D) solid tumors. In particular, new models reproducing clinical conditions as closely as possible are needed for radiobiological studies to provide information that can be translated from bench to bedside.MethodsWe developed a novel system for the irradiation, under sterile conditions, of 3-D tumor spheroids, the in vitro model considered as a bridge between the complex architectural organization of in vivo tumors and the very simple one of in vitro monolayer cell cultures. The system exploits the same equipment as that used for patient treatments, without the need for dedicated and highly expensive instruments. To mimic the passage of radiation beams through human tissues before they reach the target tumor mass, 96-multiwell plates containing the multicellular tumor spheroids (MCTS) are inserted into a custom-built phantom made of plexiglass, the material most similar to water, the main component of human tissue.ResultsThe system was used to irradiate CAEP- and A549-derived MCTS, pre-treated or not with 20 μM cisplatin, with a dose of 20 Gy delivered in one session. We also tested the same treatment schemes on monolayer CAEP and A549 cells. Our preliminary results indicated a significant increment in radiotoxicity 20 days after the end of irradiation in the CAEP spheroids pre-treated with cisplatin compared to those treated with cisplatin or irradiation alone. Conversely, the effect of the radio- chemotherapy combination in A549-derived MCTS was similar to that induced by cisplatin or irradiation alone. Finally, the 20 Gy dose did not affect cell survival in monolayer CAEP and A549 cells, whereas cisplatin or cisplatin plus radiation caused 100% cell death, regardless of the type of cell line used.ConclusionsWe set up a system for the irradiation, under sterile conditions, of tumor cells grown in 3-D which allows for the use of the same dose intensities and schedules utilized in clinical practice. This irradiation system, coupled with 3-D cell cultures, has the potential to generate information that could be used to individually tailor radiotherapy.

Project description:Targeted alpha therapy is an emerging innovative approach for the treatment of advanced cancers, in which targeting agents deliver radionuclides directly to tumors and metastases. The biological effects of α-radiation are still not fully understood - partly due to the lack of sufficiently accurate research methods. The range of α-particles is <100 μm, and therefore, standard in vitro assays may underestimate α-radiation-specific radiation effects. In this report we focus on α-radiation-induced DNA lesions, DNA repair as well as cellular responses to DNA damage. Herein, we used Ra-223 to deliver α-particles to various tumor cells in a Transwell system. We evaluated the time and dose-dependent biological effects of α-radiation on several tumor cell lines by biological endpoints such as clonogenic survival, cell cycle distribution, comet assay, foci analysis for DNA damage, and calculated the absorbed dose by Monte-Carlo simulations. The radiobiological effects of Ra-223 in various tumor cell lines were evaluated using a novel in vitro assay designed to assess α-radiation-mediated effects. The α-radiation induced increasing levels of DNA double-strand breaks (DSBs) as detected by the formation of 53BP1 foci in a time- and dose-dependent manner in tumor cells. Short-term exposure (1-8 h) of different tumor cells to α-radiation was sufficient to double the number of cells in G2/M phase, reduced cell survival to 11-20% and also increased DNA fragmentation measured by tail intensity (from 1.4 to 3.9) dose-dependently. The α-particle component of Ra-223 radiation caused most of the Ra-223 radiation-induced biological effects such as DNA DSBs, cell cycle arrest and micronuclei formation, leading ultimately to cell death. The variable effects of α-radiation onto the different tumor cells demonstrated that tumor cells show diverse sensitivity towards damage caused by α-radiation. If these differences are caused by genetic alterations and if the sensitivity could be modulated by the use of DNA damage repair inhibitors remains a wide field for further investigations.

Project description:To pursue a more mechanistic understanding of the neural control of behavior, many neuroethologists study animal behavior in controlled laboratory environments. One popular approach is to measure the movements of restrained animals while presenting controlled sensory stimulation. This approach is especially powerful when applied to genetic model organisms, such as Drosophila melanogaster, where modern genetic tools enable unprecedented access to the nervous system for activity monitoring or targeted manipulation. While there is a long history of measuring the behavior of body- and head-fixed insects walking on an air-supported ball, the methods typically require complex setups with many custom components. Here we present a compact, simplified setup for these experiments that achieves high-performance at low cost. The simplified setup integrates existing hardware and software solutions with new component designs. We replaced expensive optomechanical and custom machined components with off-the-shelf and 3D-printed parts, and built the system around a low-cost camera that achieves 180 Hz imaging and an inexpensive tablet computer to present view-angle-corrected stimuli updated through a local network. We quantify the performance of the integrated system and characterize the visually guided behavior of flies in response to a range of visual stimuli. In this paper, we thoroughly document the improved system; the accompanying repository incorporates CAD files, parts lists, source code, and detailed instructions. We detail a complete ~$300 system, including a cold-anesthesia tethering stage, that is ideal for hands-on teaching laboratories. This represents a nearly 50-fold cost reduction as compared to a typical system used in research laboratories, yet is fully featured and yields excellent performance. We report the current state of this system, which started with a 1-day teaching lab for which we built seven parallel setups and continues toward a setup in our lab for larger-scale analysis of visual-motor behavior in flies. Because of the simplicity, compactness, and low cost of this system, we believe that high-performance measurements of tethered insect behavior should now be widely accessible and suitable for integration into many systems. This access enables broad opportunities for comparative work across labs, species, and behavioral paradigms.

Dataset Information

Large Field Alpha Irradiation Setup for Radiobiological Experiments.

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Large Field Alpha Irradiation Setup for Radiobiological Experiments.

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