Prostate cancer (PCa) is the most frequent cancer in men aged 65 and over. PCa mainly metastasizes in the bone, forming osteosclerotic lesions, inducing pain, fractures, and nerve compression. Cancer cell-derived exosomes participate in the metastatic spread, ranging from oncogenic reprogramming to the formation of pre-metastatic niches. Moreover, exosomes were recently involved in the dialog between PCa cells and the bone metastasis microenvironment. Phospholipase D (PLD) isoforms PLD1/2 catalyze the hydrolysis of phosphatidylcholine to yield phosphatidic acid (PA), regulating tumor progression and metastasis. PLD is suspected to play a role in exosomes biogenesis. We aimed to determine whether PCa-derived exosomes, through PLD, interact with the bone microenvironment, especially osteoblasts, during the metastatic process. Here we demonstrate for the first time that PLD2 is present in exosomes of C4-2B and PC-3 cells. C4-2B-derived exosomes activate proliferation and differentiation of osteoblasts models, by stimulating ERK 1/2 phosphorylation, by increasing the tissue-nonspecific alkaline phosphatase activity and the expression of osteogenic differentiation markers. Contrariwise, when C4-2B exosomes are generated in the presence of halopemide, a PLD pan-inhibitor, they lose their ability to stimulate osteoblasts. Furthermore, the number of released exosomes diminishes significantly (−40%). When the PLD product PA is combined with halopemide, exosome secretion is fully restored. Taken together, our results indicate that PLD2 stimulates exosome secretion in PCa cell models as well as their ability to increase osteoblast activity. Thus, PLD2 could be considered as a potent player in the establishment of PCa bone metastasis acting through tumor cell derived-exosomes.

前列腺癌（PCa）是65岁以上男性中最常见的癌症。PCa主要在骨中转移，形成骨硬化性病变，引起疼痛，骨折和神经受压。癌细胞衍生的外来体参与转移扩散，范围从致癌重编程到转移前小生境的形成。而且，外泌体最近参与了PCa细胞与骨转移微环境之间的对话。磷脂酶D（PLD）异构体PLD1 / 2催化磷脂酰胆碱的水解，产生磷脂酸（PA），调节肿瘤的进展和转移。怀疑PLD在外泌体生物发生中起作用。我们旨在确定在转移过程中PCa衍生的外泌体是否通过PLD与骨微环境（尤其是成骨细胞）相互作用。在这里，我们首次证明C4-2B和PC-3细胞的外泌体中存在PLD2。C4-2B衍生的外来体通过刺激ERK 1/2磷酸化，增加组织非特异性碱性磷酸酶活性和成骨分化标志物的表达来激活成骨细胞模型的增殖和分化。相反，当在PLD泛抑制剂halopemide存在下产生C4-2B外泌体时，它们会失去刺激成骨细胞的能力。此外，释放的外泌体的数量显着减少（-40％）。当PLD产物PA与卤胺合用时，外泌体分泌被完全恢复。两者合计，我们的结果表明PLD2刺激PCa细胞模型中外泌体的分泌，以及它们增加成骨细胞活性的能力。从而，