Quantitative volumetric analysis of the Golgi apparatus following X-ray irradiation by super-resolution 3D-SIM microscopy

摘要

To obtain quantitative volumetric data for the Golgi apparatus after ionizing radiation (IR) using super-resolution three-dimensional structured illumination (3D-SIM) microscopy. Normal human retinal pigment epithelial (RPE) cells were irradiated with X-rays (10 Gy), followed by immunofluorescence staining of the Golgi marker RCAS1. 3D-SIM imaging was performed using Delta Vision OMX version 4 and SoftWoRx 6.1. Polygon rendering and spot signal identification were performed using Imaris 8.1.2. Differences between groups were assessed by Welch's t test. RCAS1 signals in untreated cells were located adjacent to nuclei and showed a reticular morphology. Upon IR, the area of RCAS1 signals expanded while retaining the reticular morphology. Polygon rendering imaging revealed that the volume of RCAS1 at 48 h post-IR was greater than that for unirradiated cells (93.7 ± 19.0 μm~3 vs. 33.0 ±4.2 μm~3, respectively; P ＜ 0.001): a 2.8-fold increase. Spot signal imaging showed that the number of RCAS1 spot signals post-IR was greater than that for unirradiated cells [3.4 ± 0.8 (×10~3) versus 1.3 ±0.2 (× 10~3), respectively;P＜0.001]: a 2.7-fold increase. This is the first study to report quantitative volumetric data of the Golgi apparatus in response to IR using super-resolution 3D-SIM microscopy.