Radiation exposure of peripheral mononuclear blood cells alters the composition and function of their extracellular vesicles

Normal tissue toxicity is a dose-limiting factor in radiation therapy. One component of normal tissue that is continuously exposed during therapeutic irradiation is the circulating population of peripheral blood mononuclear cells (PBMC). PBMCs are highly sensitive to ionizing radiation (IR), however little is known about how ionizing radiation (IR) affects the PBMC response on a systemic level. Therefore it was the aim of this study to investigate whether IR was capable to induce changes in the composition and function of extracellular vesicles (EVs) secreted from PBMCs after radiation exposure to different doses.Whole blood samples from healthy donors were irradiated with 0 Gy, 0.1 Gy, 2 Gy or 6 Gy X-rays and PBMC-secreted EVs were isolated 72 hours later. Proteome and miRNome analysis of EVs showed a dose-dependent increase in the number of significantly deregulated proteins and microRNAs. For both, proteome and microRNA data, principal component analysis showed a dose-dependent separation of control and exposed groups. Integrated pathway analysis of the radiation-regulated EV proteins and microRNAs consistently predicted an association of deregulated molecules with apoptosis, cell death and survival. Functional studies identified endothelial cells as an efficient EV recipient system, in which irradiation of recipient cells further increased the uptake. Furthermore we detected an apoptosis suppressive effect of EVs from irradiated PBMCs in endothelial recipient cells. In summary, our study demonstrates that IR modifies the communication between PBMCs and endothelial cells. We identified EVs from irradiated PBMC donors as transmitters of protective signals to irradiated endothelial cells. Thus our data may lead to the discovery of biomarker candidates for radiation dosimetry and even more importantly, they suggest EVs as a novel systemic communication pathway between irradiated and non-irradiated normal, non-cancer tissues.

正常组织毒性是放射治疗中的剂量限制性因素。治疗性照射过程中持续暴露的正常组织组分之一，是循环外周血单个核细胞（peripheral blood mononuclear cells, PBMC）群体。外周血单个核细胞对电离辐射（ionizing radiation, IR）具有高度敏感性，但目前对于电离辐射如何在系统层面影响PBMC的应答机制尚不清楚。因此本研究旨在探讨不同剂量电离辐射照射后，PBMC分泌的细胞外囊泡（extracellular vesicles, EVs）的组成与功能是否会发生改变。本研究采集健康供者的全血样本，分别给予0 Gy、0.1 Gy、2 Gy及6 Gy的X射线照射，并于照射后72小时分离PBMC分泌的细胞外囊泡。对细胞外囊泡的蛋白质组与miRNA组分析结果显示，显著失调的蛋白质及microRNA数量呈剂量依赖性升高。针对蛋白质组及microRNA数据的主成分分析（principal component analysis）结果显示，对照组与照射组呈剂量依赖性分离。对辐射调控的细胞外囊泡蛋白质及microRNA进行的整合通路分析，一致预测失调分子与细胞凋亡、细胞死亡及细胞存活存在显著关联。功能实验证实，内皮细胞（endothelial cells）是高效的细胞外囊泡摄取系统，而受体细胞的照射可进一步提升其摄取效率。此外，本研究还发现，照射后的PBMC分泌的细胞外囊泡，可对内皮受体细胞产生抑制细胞凋亡的作用。综上，本研究证实电离辐射可改变PBMC与内皮细胞之间的信号交流。本研究发现，来自照射后PBMC供者的细胞外囊泡，可向受照射的内皮细胞传递保护性信号。因此，本研究数据有望筛选出辐射剂量学的潜在生物标志物；更为重要的是，本研究揭示细胞外囊泡是受照射与未受照射的正常非癌组织之间一种全新的系统性通信通路。