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Report No.

Autophagy activated by Auger effect induced by soft X-ray microbeam

Noguchi, Miho; Yokoya, Akinari; Kaminaga, Kiichi; Fujii, Kentaro; Suzuki, Keiji*; Usami, Noriko*

In this study, to clarify activation of autophagy on energy deposition in the cell nucleus or cytoplasm by the photoelectric effect, we investigated change of autophagic activity in human fibroblast cells irradiated with soft X-ray microbeam (5.35keV).Using a synchrotron X-ray microbeam, we irradiated 25 to 61 cells by targeting nuclei with square X-ray microbeam (10 $$mu$$m $$times$$ 10 $$mu$$m). For irradiating cytoplasm, 60 $$mu$$m $$times$$ 60 $$mu$$m square X-ray microbeam was used with a metal mask of 10$$mu$$m $$times$$ 10 $$mu$$m central area not to irradiate the cell nucleus. Induction of autophagy was measured using the fluorescent probe, Cyto-ID Green, which stains specifically autophagic vacuoles. Irradiated cells were treated with the dye 15 min before observation. The cells observed by a fluorescent microscope were quantified as mean values of the fluorescent intensity per cell. We observed autophagic fluorescence in nucleus- or cytoplasm-irradiated cells at 1 day to 4 days after irradiation. Some of these cells showed highly localized strong fluorescence. Such localized fluorescence was rarely observed when irradiated with a wide beam from a conventional higher energy X-ray machine (150 kVp). A lot of nucleic irradiated cells, rather than cytoplasm-irradiated cell, showed cell death especially at 4 days after irradiation. When irradiated with the wide beam X-rays, on the other hand, the cells sustained irreversible growth arrest, and maintained their viability. These results indicate that the soft X-ray microbeam exposure is a powerful probe to provide us an aspect of autophagic activation by photoelectric (Auger) effect in a particular part of cells.



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