Possible relationship between mitochondrial changes and oxidative stress under low dose-rate irradiation

Objectives: High dose-rate ionizing radiation (IR) causes severe DSB damage, as well as reactive oxygen species (ROS) accumulation and oxidative stress. However, it is unknown what biological processes are affected by low dose-rate IR; therefore, the molecular relationships between mitochondria changes and oxidative stress in human normal cells was investigated after low dose-rate IR. Methods: We compared several cellular response between high and low dose-rate irradiation using cell survival assay, ROS/RNS assay, immunofluorescence and western blot analysis. Results: Reduced DSB damage and increased levels of ROS, with subsequent oxidative stress responses, were observed in normal cells after low dose-rate IR. Low dose-rate IR caused several mitochondrial changes, including morphology mass, and mitochondrial membrane potential, suggesting that mitochondrial damage was caused. Although damaged mitochondria were removed by mitophagy to stop ROS leakage, the mitophagy-regulatory factor, PINK1, was reduced following low dose-rate IR. Although mitochondrial dynamics (fission/fusion events) are important for the proper mitophagy process, some mitochondrial fusion factors decreased following low dose-rate IR. Discussion: The dysfunction of mitophagy pathway under low dose-rate IR increased ROS and the subsequent activation of the oxidative stress response.</p

Bisulfite sequencing of the <i>Apc</i> promoter region in small intestinal tumors of B6/B6-Chr18^MSM-F1 <i>Apc</i>^<i>Min/+</i> mice.

Methylation status of the 93 CpG dinucleotides in the promoter region of Apc. Each circle represents a CpG site in the genomic DNA sequence and each row of circles represents the analysis of a single cloned allele. Closed circles: methylated CpG dinucleotides; open circles: unmethylated CpG dinucleotides. Bisulfite efficiency averaged 98.5 ± 1.33% for CpG island 1 and 98.8 ± 1.23% for CpG island 2 in all samples examined.</p

Sequence analysis of <i>Apc</i> in intestinal tumors from B6/B6-Chr18^MSM-F1 <i>Apc</i>^<i>Min/+</i> mice.

Sequence analysis of Apc in intestinal tumors from B6/B6-Chr18MSM-F1 ApcMin/+ mice.</p

Analysis of LOH of Chr18 in mouse intestinal tumors.

Each row shows the LOH of one microsatellite marker on Chr18, and each column shows the data for one intestinal tumor. Open circles indicate the loss of either the wild-type or Min alleles, and filled circles indicate the retention of both alleles. Based on the LOH results, each tumor was categorized as 0 Gy Unidentified, 2 Gy Unidentified, or 2 Gy Deletion. (TIF)</p

Mutations identified in the Unidentified type of intestinal tumors.

Mutations identified in the Unidentified type of intestinal tumors.</p

Bisulfite sequencing of the sites corresponding to human <i>APC</i> promoter 1A in small intestinal tumors of B6/B6-Chr18^MSM-F1 <i>Apc</i>^<i>Min/+</i> mice.

Methylation status of the 21 CpG dinucleotides in the promoter region of Apc. Each circle represents a CpG site in the genomic DNA sequence and each row of circles represents the analysis of a single cloned allele. Closed circles: methylated CpG dinucleotides; open circles: unmethylated CpG dinucleotides. (TIF)</p

<i>Apc</i> genomic region analyzed in this study.

Nucleotide positions are numbered relative to the transcription start site of Apc (NM_007462). Capital letters represent CpG islands obtained from the UCSC Genome Browser (http://genome.ucsc.edu/). The CpG sites are highlighted in red. Primer pairs (boxed sequences) were designed using MethPrimer (http://www.urogene.org/cgi-bin/methprimer/methprimer.cgi). Arrows indicate the direction from 5’ to 3’. Due to the relative length of this CpG island, it was split into two segments, and bisulfite sequencing was performed for each segment. (TIF)</p

Figure S1 from Radiation-Induced Myofibroblasts Promote Tumor Growth via Mitochondrial ROS–Activated TGFβ Signaling

Induction of alpha-SMA after acute single radiation</p

Supplemental Figure Legends from Radiation-Induced Myofibroblasts Promote Tumor Growth via Mitochondrial ROS–Activated TGFβ Signaling

S1. The percentage of TIG-3 and MRC-5 cells with alpha-SMA staining exposed to indicated doses is shown in the graph. S2. Tumor volumes of HeLa, HeLa+MRC-5 0FR tumors, HeLa+MRC-5 0.05FR tumors, HeLa+MRC-5 NAC 0FR tumors, and HeLa+MRC-5 0.05FR tumors.</p

DNA copy number aberrations in intestinal tumors from the array CGH analysis.

DNA copy number aberrations in intestinal tumors from the array CGH analysis.</p