Norwegian College of Fishery Science, UiT The Arctic University of Norway, Tromsø, Norway.
Institute of Medical Biology, Faculty of Health Sciences, UiT The Arctic University of Norway, Tromsø, Norway.
FEBS J. 2018 Aug;285(16):3114-3133. doi: 10.1111/febs.14601. Epub 2018 Jul 12.
Exosomes are distinguished from other types of extracellular vesicles by their small and relatively uniform size (30-100 nm) and their composition which reflects their endo-lysosomal origin. Involvement of these extracellular organelles in intercellular communication and their implication in pathological conditions has fuelled intensive research on mammalian exosomes; however, currently, very little is known about exosomes in lower vertebrates. Here we show that, in primary cultures of head kidney leukocytes from Atlantic salmon (Salmo salar), phosphorothioate CpG oligodeoxynucleotides induce secretion of vesicles with characteristics very similar to these of mammalian exosomes. Further experiments revealed that the oligonucleotide-induced exosome secretion did not depend on the CpG motifs but it relied on the phosphorothioate modification of the internucleotide linkage. Exosome secretion was also induced by genomic bacterial and eukaryotic DNA in toll-like receptor 9-negative piscine and human cell lines demonstrating that this is a phylogenetically conserved phenomenon which does not depend on activation of immune signaling pathways. In addition to exosomes, stimulation with phosphorothioate oligonucleotides and genomic DNA induced secretion of LC3B-II, an autophagosome marker, which was associated with vesicles of diverse size and morphology, possibly derived from autophagosome-related intracellular compartments. Overall, this work reveals a previously unrecognized biological activity of phosphorothioate ODNs and genomic DNA - their capacity to induce secretion of exosomes and other types of extracellular vesicles. This finding might help shed light on the side effects of therapeutic phosphorothioate oligodeoxynucleotides and the biological activity of extracellular genomic DNA which is often upregulated in pathological conditions.
外泌体通过其小而相对均匀的大小（30-100nm）和反映其内体溶酶体起源的组成与其他类型的细胞外囊泡区分开来。这些细胞外细胞器在细胞间通讯中的作用及其在病理条件下的意义，促使人们对哺乳动物外泌体进行了深入研究；然而，目前对于低等脊椎动物的外泌体知之甚少。在这里，我们展示了在大西洋鲑（Salmo salar）头肾白细胞的原代培养物中，硫代磷酸酯 CpG 寡脱氧核苷酸诱导具有与哺乳动物外泌体非常相似特征的囊泡的分泌。进一步的实验表明，寡核苷酸诱导的外泌体分泌不依赖于 CpG 基序，而是依赖于核苷酸间连接的硫代磷酸酯修饰。在 Toll 样受体 9 阴性鱼类和人类细胞系中，基因组细菌和真核 DNA 也诱导外泌体的分泌，这表明这是一种进化上保守的现象，不依赖于免疫信号通路的激活。除了外泌体之外，用硫代磷酸酯寡核苷酸和基因组 DNA 刺激还诱导了 LC3B-II 的分泌，LC3B-II 是自噬体的标志物，与不同大小和形态的囊泡相关联，可能来源于自噬体相关的细胞内隔室。总的来说，这项工作揭示了硫代磷酸酯寡核苷酸和基因组 DNA 的一种以前未被认识的生物学活性-它们诱导外泌体和其他类型的细胞外囊泡分泌的能力。这一发现可能有助于阐明治疗用硫代磷酸酯寡核苷酸的副作用以及在病理条件下经常上调的细胞外基因组 DNA 的生物学活性。
