Department of Orthopedics, The Fifth People's Hospital of Shanghai, Fudan University, China.
Department of Orthopedics, The Fifth People's Hospital of Shanghai, Fudan University, China.
Exp Cell Res. 2019 Oct 15;383(2):111541. doi: 10.1016/j.yexcr.2019.111541. Epub 2019 Jul 29.
The skeletal system is a dynamic organ that continuously undergoes coupled trabeculae and blood vessels remodeling, indicating the possible existence of molecular crosstalk between endothelial and osteoblastic cells. Since the cross-talk between bone-forming osteoblasts (OBs) and vessel-forming endothelial cells (ECs) have progressively gained investigators' attention, few studies focused on the regulatory function of extracellular vesicles derived from OBs on ECs. In this study, the effect of the exosomes derived from mature osteoblasts (MOBs) on the ECs was investigated. Firstly, exosomes derived from mature osteoblasts (MOB-Exos) were isolated and identified by NanoSight light scatter technology, electron microscopy and Western bolting. Fluorescent labeling of MOB-Exos revealed its internalization by ECs. RNA interference technique was used to knock down matrix metalloproteinase-2 (MMP2) in MOB-Exos. Then ECs were co-cultured with MOB-Exos and MMP2 knockdown MOB-Exos. Wound healing migration assay, transwell migration assay, CCK-8 assay and tube formation assay of ECs were conducted to determine the angiogenic capability of ECs. Then the VEGF/Erk1/2 pathway markers were detected by Western blot. Our results showed that MOB-Exos could promote the proliferation, migration and tube formation of ECs. Meanwhile, the promoted angiogenetic capacities of ECs were impaired when MMP2 in MOB-Exos was knocked down. In addition, immunoblotting indicated that MOB-Exos could promote the activation of the VEGF/Erk1/2 pathway of ECs; whereas the activation of the VEGF/Erk1/2 pathway was attenuated when the ECs were co-cultured with the MMP2 knockdown MOB-Exos. In conclusion, the MMP-2 existing in exosomes derived from MOBs could promote the angiogenesis of ECs in vitro, which might be realized through VEGF/Erk1/2 signaling pathway.
骨骼系统是一个动态器官，不断进行小梁和血管的重塑，表明内皮细胞和成骨细胞之间可能存在分子串扰。由于成骨细胞（OBs）和血管形成内皮细胞（ECs）之间的串扰逐渐引起研究人员的关注，因此很少有研究关注源自 OBs 的细胞外囊泡对 ECs 的调节功能。在这项研究中，研究了成熟成骨细胞（MOB）衍生的外泌体对 ECs 的影响。首先，通过纳米粒子光散射技术、电子显微镜和 Western blot 鉴定了成熟成骨细胞（MOB）衍生的外泌体（MOB-Exos）。MOB-Exos 的荧光标记显示其被 ECs 内化。使用 RNA 干扰技术敲低 MOB-Exos 中的基质金属蛋白酶-2（MMP2）。然后将 ECs 与 MOB-Exos 和 MMP2 敲低 MOB-Exos 共培养。进行 ECs 的划痕愈合迁移实验、Transwell 迁移实验、CCK-8 实验和管形成实验，以确定 ECs 的血管生成能力。然后通过 Western blot 检测 VEGF/Erk1/2 通路标志物。我们的结果表明，MOB-Exos 可以促进 ECs 的增殖、迁移和管形成。同时，当 MOB-Exos 中的 MMP2 被敲低时，ECs 的促血管生成能力受损。此外，免疫印迹表明 MOB-Exos 可以促进 ECs 中 VEGF/Erk1/2 通路的激活；而当 ECs 与 MMP2 敲低的 MOB-Exos 共培养时，VEGF/Erk1/2 通路的激活会减弱。总之，MOB 衍生的外泌体中的 MMP-2 可以促进体外 ECs 的血管生成，这可能是通过 VEGF/Erk1/2 信号通路实现的。
