Department of Nanomedicine (DNP), Graduate School of Medical and Dental Science, Tokyo Medical and Dental University, 1-5-45, Yushima, Bunkyo-ku, 113-8510, Tokyo, Japan.
Current Address: Kanagawa Dental University, Yokohama Clinic, Tsuruya-cho 3-31-6, Kanagawa-ku, Yokohama, Kanagawa, 221-0835, Japan.
Stem Cell Res Ther. 2017 Oct 3;8(1):219. doi: 10.1186/s13287-017-0660-9.
The therapeutic potential of mesenchymal stem cells (MSCs) may be attributed partly to humoral factors such as growth factors, cytokines, and chemokines. Human term placental tissue-derived MSCs (PlaMSCs), or conditioned medium left over from cultures of these cells, have been reported to enhance angiogenesis. Recently, the exosome, which can transport a diverse suite of macromolecules, has gained attention as a novel intercellular communication tool. However, the potential role of the exosome in PlaMSC therapeutic action is not well understood. The purpose of this study was to evaluate PlaMSC-derived exosome angiogenesis promotion in vitro and in vivo.
MSCs were isolated from human term placental tissue by enzymatic digestion. Conditioned medium was collected after 48-h incubation in serum-free medium (PlaMSC-CM). Angiogenic factors present in PlaMSC-CM were screened by a growth factor array. Exosomes were prepared by ultracentrifugation of PlaMSC-CM, and confirmed by transmission electron microscopy, dynamic light scattering, and western blot analyses. The proangiogenic activity of PlaMSC-derived exosomes (PlaMSC-exo) was assessed using an endothelial tube formation assay, a cell migration assay, and reverse transcription-PCR analysis. The in-vivo angiogenic activity of PlaMSC-exo was evaluated using a murine auricle ischemic injury model.
PlaMSC-CM contained both angiogenic and angiostatic factors, which enhanced endothelial tube formation. PlaMSC-exo were incorporated into endothelial cells; these exosomes stimulated both endothelial tube formation and migration, and enhanced angiogenesis-related gene expression. Laser Doppler blood flow analysis showed that PlaMSC-exo infusion also enhanced angiogenesis in an in-vivo murine auricle ischemic injury model.
PlaMSC-exo enhanced angiogenesis in vitro and in vivo, suggesting that exosomes play a role in the proangiogenic activity of PlaMSCs. PlaMSC-exo may be a novel therapeutic approach for treating ischemic diseases.
间充质干细胞（MSCs）的治疗潜力部分归因于其分泌的体液因子，如生长因子、细胞因子和趋化因子。已报道来源于人足月胎盘组织的间充质干细胞（PlaMSCs）或这些细胞培养后的条件培养基可促进血管生成。最近，作为一种新型细胞间通讯工具，外泌体（可以转运多种生物大分子）引起了人们的关注。然而，外泌体在 PlaMSC 治疗作用中的潜在作用尚不清楚。本研究旨在评估 PlaMSC 来源的外泌体在体外和体内促进血管生成的作用。
通过酶消化法从人足月胎盘组织中分离 MSCs。在无血清培养基中孵育 48 小时后收集条件培养基（PlaMSC-CM）。通过生长因子阵列筛选 PlaMSC-CM 中存在的血管生成因子。通过超速离心法制备 PlaMSC-CM 中的外泌体，并通过透射电子显微镜、动态光散射和 Western blot 分析进行确认。使用内皮管形成试验、细胞迁移试验和逆转录-PCR 分析评估 PlaMSC 来源的外泌体（PlaMSC-exo）的促血管生成活性。使用小鼠耳廓缺血损伤模型评估 PlaMSC-exo 的体内促血管生成活性。
PlaMSC-CM 含有促血管生成和抗血管生成因子，可增强内皮管形成。PlaMSC-exo 被内吞到内皮细胞中；这些外泌体刺激内皮细胞的管形成和迁移，并增强与血管生成相关的基因表达。激光多普勒血流分析显示，PlaMSC-exo 输注也可增强体内小鼠耳廓缺血损伤模型中的血管生成。
PlaMSC-exo 增强了体外和体内的血管生成，提示外泌体在 PlaMSC 的促血管生成活性中发挥作用。PlaMSC-exo 可能是治疗缺血性疾病的一种新的治疗方法。
