Gene expression profiles of irradiated lung tissue in three mouse strains

The aim of our research is to clarify the mechanisms generating heterogeneity in response to C-ion irradiation that arise from individual genetic variations in humans. In this study, we performed whole lung C-ion irradiation using three different strains of mice to examine whether strain-dependent differences in radiation effects occur in high-LET C-ion thoracic irradiation. Murine strain-variance was evaluated by histopathological examination of intra-alveolar hemorrhage that is likely to occur during the early phase after irradiation, and of lung fibrosis during the late phase, occurring more than three months after irradiation. We also performed microarray analysis to identify the key genes that are differentially regulated in different mouse strains after C-ion irradiation and to determine the mechanism of strain-dependent pulmonary damage after high-LET C-ion irradiation. Eight-week old female inbred C3H/He Slc, C57BL/6J Jms Slc and A/J Jms Slc mice (3 kinds of mice) were used. The whole thorax of three mice was locally irradiated at 10 Gy with C-ion beams, with a reference beam, 137Cs gamma-rays (2 kinds of beam). Three mice of each strain (3 mice) were sacrificed, and immediately dissected for lung extraction at 6 h (6h sample). Lungs for three mice of each strain without irradiation were extracted at the same time as control samples (control sample). Hybridization to microarrays (Whole Mouse Genome 4x44K OligoMicroarray Kit, single color, Cyamine 3-CTP) consisting of 44,000 total spots was conducted using an Agilent Gene Expression hybridization kit. The arrays from three independent replicates for each sample (3 arrays) were scanned on an Agilent dual-laser Microarray Scanner (all from Agilent Technologies) following the manufacturer's instructions. A total of 108 samples were analyzed. 3 replicate arrays x 3 mice x 3 strains x 2 timing for sampling x 2 kinds of beam.

本研究旨在阐明人类个体遗传差异所导致的碳离子束照射应答异质性的产生机制。本研究选用三种不同品系的小鼠开展全肺碳离子照射实验，以探究高线性能量转移（high-LET）碳离子胸部照射是否存在品系依赖性的辐射效应差异。通过组织病理学检查，分别评估照射后早期阶段易出现的肺泡内出血情况，以及照射后三个月以上晚期阶段的肺纤维化程度，以此评价小鼠品系间的差异。此外，本研究还开展了微阵列（microarray）分析，以筛选碳离子照射后不同小鼠品系中差异调控的关键基因，并阐明高线性能量转移（high-LET）碳离子照射后品系依赖性肺损伤的发生机制。本研究选用8周龄雌性近交系C3H/He Slc、C57BL/6J Jms Slc及A/J Jms Slc三种小鼠品系。随后，对每品系3只小鼠的全胸廓进行局部照射，照射剂量为10 Gy，辐射源包括碳离子束及参照束137Cs γ射线（共2种辐射束）。分别在照射后6小时（6h样本）处死3只小鼠，即刻解剖摘取肺组织；同时，采集每品系未照射小鼠各3只的肺组织作为对照样本。采用包含总计44000个探针点的Agilent全小鼠基因组4x44K寡核苷酸微阵列试剂盒（单荧光标记，Cyamine 3-CTP），使用Agilent基因表达杂交试剂盒完成杂交实验。按照制造商说明书，使用Agilent双激光微阵列扫描仪对每个样本的3次独立重复杂交阵列（共3张阵列）进行扫描。本研究共计分析108个样本，样本量计算公式为：3张重复阵列 × 3只小鼠 × 3个品系 × 2个采样时间点 × 2种辐射束。