Department of Hematology, the Second Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, PR China.
Department of Hematology, the Second Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, PR China.
Int Immunopharmacol. 2024 Dec 5;142(Pt B):113080. doi: 10.1016/j.intimp.2024.113080. Epub 2024 Sep 16.
Multiple myeloma (MM), characterized with bone marrow microenvironment disorder, accounts for about 20% of hematological cancer deaths globally. Tissue extracellular communication, especially extracellular vesicles, has been defined as important mediator among cell-to-cell cross-talk. Our previous study revealed an elevated level of H19 in MM, whereas, its role in MM exosomes in the development of osteolysis remains largely unknown.
MM exosomes referring to 5TGM1 cells were isolated and characterized using transmission electron microscopy (TEM), nanoparticle tracking and western blot analysis. The biological effects of blocking H19 were examined on osteolysis in vivo of C57Bl6/KalwRij mice, as well as on the osteoclast differentiation in vitro of RAW264.7 cells, by the application of TRAP, either with osteogenic differentiation in vitro of bone marrow mesenchymal stem cells (BMSCs), by the detection of alkaline phosphatase (ALP), alizarin red dye staining (ARS). The targeted relationships among H19/hnRNPA2B1/BET proteins were validated through RNA immunoprecipitation (RIP) and RNA pull-down assays.
5TGM1 cells derived-exosomes lacking H19 dramatically blocked osteolysis and boosted osteogeneis in C57Bl6/KalwRij mice, either with osteoclastic differentiation of RAW264.7 cells and osteogenic differentiation of BMSCs, thereby enhancing their resorptive activity. Physically, H19 interacted with hnRNPA2B1 by preferentially adhering to it and enhancing its nuclear-cytoplasmic translocation. Further mechanistic research validated that H19 promoted the stabilization of BET proteins through hnRNA2B1 to be involved in osteoclast differentiation for contributing to MM progression.
Altogether, our findings suggest that H19, serving as an essential role for exosomes in the bone marrow environment, might be a viable diagnostic and therapeutic target for MM therapy.
多发性骨髓瘤（MM），其特征为骨髓微环境紊乱，约占全球 20%的血液系统癌症死亡病例。细胞外组织通讯，特别是细胞外囊泡，已被定义为细胞间通讯的重要介质。我们之前的研究表明，H19 在 MM 中升高，然而，它在 MM 外泌体中对溶骨性骨破坏的作用在很大程度上仍然未知。
使用透射电子显微镜（TEM）、纳米颗粒跟踪和 Western blot 分析分离和表征 5TGM1 细胞来源的 MM 外泌体。通过应用 TRAP，在 C57Bl6/KalwRij 小鼠体内观察到阻断 H19 对破骨细胞分化的影响，以及在体外 RAW264.7 细胞中的破骨细胞分化，通过骨形态发生蛋白（BMPs）诱导的骨髓间充质干细胞（BMSCs）体外成骨分化，通过碱性磷酸酶（ALP）检测、茜素红染料染色（ARS）。通过 RNA 免疫沉淀（RIP）和 RNA 下拉实验验证 H19/hnRNPA2B1/BET 蛋白之间的靶向关系。
缺乏 H19 的 5TGM1 细胞衍生的外泌体可显著阻断 C57Bl6/KalwRij 小鼠的溶骨性骨破坏，并增强破骨细胞分化、RAW264.7 细胞和 BMSCs 的成骨分化，从而增强其吸收活性。实际上，H19 通过优先与其结合并增强其核质易位与 hnRNPA2B1 相互作用。进一步的机制研究验证了 H19 通过 hnRNA2B1 促进 BET 蛋白的稳定性，从而参与破骨细胞分化，促进 MM 的进展。
总的来说，我们的研究结果表明，H19 在外泌体在骨髓环境中起着重要作用，可能是 MM 治疗的一种可行的诊断和治疗靶点。
