Distinct vascular genomic effects of proton and gamma radiation. Distinct vascular genomic effects of proton and gamma radiation

We performed comparative RNA sequencing of the early (4 hrs) dose response (0.5 – 200 cGy whole body dose, 10 dose levels) of the mouse aorta to proton and gamma radiation. Total-body proton radiation of conscious animals was performed using a proton beam produced by a cyclotron system, while total-body gamma radiation of animals was performed using a Caesium-137 gamma source. A trend analysis identified genes that showed a dose response, using data permutation to estimate a false discovery rate (q-value) for each gene. We identified 29 and 194 genes (q-value ≤ 0.1) that were upregulated with increasing doses of proton and gamma radiation, respectively. No genes were down-regulated. While fewer genes were dose-responsive to proton radiation, the magnitude of the effect was greater than with gamma radiation. These highly responsive genes were enriched for pathways involved in the response to DNA damage, apoptosis, cellular stress and inflammation (p < 0.01). Gamma radiation responsive genes included the same pathways, but extended to genes in vasculature specific pathways. Genes responsive to both radiation types (19 genes at q-value ≤ 0.1) showed almost perfectly superimposable dose-response relationships. We observed the same superimposable dose response relationship of gamma and proton radiations in a subset of genes validated by quantitative PCR not only in the aorta but also in liver, lung, heart and kidney. Despite a relative similar relative biological effectiveness of protons and gamma photons and the activation of canonical radiation response pathways by both radiation types, we detected marked differences in the genomic response. It seems plausible that these genomic differences translate into differences in the biological processes leading to cardiovascular pathologies. Overall design: RNA-Seq 20 samples, mouse aorta. 10 mice were treated with ten different centigrays of gamma radiation and 10 mice were treated with ten different centigrays of high energy protons (0 cGy, 5 cGy, 10 cGy, 25 cGy, 50 cGy, 75 cGy, 100 cGy, 125 cGy, 150 cGy, 200 cGy). One sample per dose.

本研究针对小鼠主动脉在质子与γ射线照射后的早期（4小时）剂量响应（全身照射剂量范围0.5~200 cGy，共10个剂量梯度）开展了比较RNA测序。清醒状态小鼠的全身质子照射采用回旋加速器产生的质子束完成，而全身γ射线照射则使用铯-137（¹³⁷Cs）γ放射源进行。本研究通过趋势分析筛选出呈现剂量响应的基因，并采用数据置换法为每个基因估算错误发现率（q值，q-value）。本研究分别筛选出29个和194个随质子、γ射线照射剂量升高而上调的基因（q值≤0.1），未检测到下调表达的基因。尽管响应质子照射的剂量响应基因数量更少，但其表达变化幅度显著高于γ射线照射组。上述高响应基因显著富集于DNA损伤响应、细胞凋亡、细胞应激及炎症相关通路（p<0.01）。γ射线响应基因同样覆盖上述通路，且额外包含血管特异性通路相关基因。对两种射线均产生响应的基因（共19个，q值≤0.1），其剂量响应曲线几乎完全重合。通过定量聚合酶链反应（quantitative PCR, qPCR）验证的部分基因中，我们观测到γ射线与质子照射的剂量响应曲线同样完全重合，且该现象不仅存在于主动脉组织，还在肝脏、肺脏、心脏及肾脏组织中被发现。尽管质子与γ光子的相对生物有效性（relative biological effectiveness, RBE）相近，且两种射线均能激活经典辐射响应通路，但本研究仍检测到二者在基因组响应层面存在显著差异。上述基因组层面的差异或可转化为导致心血管病变的生物学过程差异，该推论具备合理性。实验整体设计：针对小鼠主动脉组织开展RNA测序，共纳入20个样本。其中10只小鼠接受10个不同梯度的γ射线全身照射，另外10只小鼠接受对应梯度的高能质子全身照射，剂量梯度涵盖0 cGy、5 cGy、10 cGy、25 cGy、50 cGy、75 cGy、100 cGy、125 cGy、150 cGy及200 cGy，每个剂量梯度对应1个样本。