Zhang Gensheng, Liu Ruiyu, Dang Xiaoqian, Liu Jichao, Jiao Haibin
Department of Orthopaedics, 3201 Hospital of Xi'an Jiaotong University Health Science Center, Hanzhong Shaanxi, 723000, P.R.China.
Department of Orthopaedics, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an Shaanxi, 710004, P.R.China.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2021 Mar 15;35(3):356-365. doi: 10.7507/1002-1892.202011026.
To investigate whether exosomes derived from miR-27a-overexpressing human umbilical vein endothelial cells (HUVECs)-exo (miR-27a) can promote bone regeneration and improve glucocorticoids (GC) induced osteonecrosis of femoral head (ONFH) (GC-ONFH).
The exo (miR-27a) were intended to be constructed and identified by transmission electron microscopy, nanoparticle tracking analysis, Western blot, and real-time fluorescent quantitative PCR (qRT-PCR). qRT-PCR was used to evaluate the effect of exo (miR-27a) in delivering miR-27a to osteoblasts (MC3T3-E1 cells). Alkaline phosphatase staining, alizarin red staining, and qRT-PCR were used to evaluate its effect on MC3T3-E1 cells osteogenesis. Dual-luciferase reporter (DLRTM) assay was used to verify whether miR-27a targeting Dickkopf WNT signaling pathway inhibitor 2 (DKK2) was a potential mechanism, and the mechanism was further verified by qRT-PCR, Western blot, and alizarin red staining in MC3T3-E1 cells. Finally, the protective effect of exo (miR-27a) on ONFH was verified by the GC-ONFH model in Sprague Dawley (SD) rats.
Transmission electron microscopy, nanoparticle tracking analysis, Western blot, and qRT-PCR detection showed that exo (miR-27a) was successfully constructed. exo (miR-27a) could effectively deliver miR-27a to MC3T3-E1 cells and enhance their osteogenic capacity. The detection of DLRTM showed that miR-27a promoted bone formation by directly targeting DDK2. Micro-CT and HE staining results of animal experiments showed that tail vein injection of exo (miR-27a) improved the osteonecrosis of SD rat GC-ONFH model.
exo (miR-27a) can promote bone regeneration and protect against GC-ONFH to some extent.
研究过表达miR-27a的人脐静脉内皮细胞来源的外泌体（HUVECs-exo(miR-27a)）是否能促进骨再生并改善糖皮质激素（GC）诱导的股骨头坏死（ONFH）（GC-ONFH）。
构建外泌体（miR-27a），并通过透射电子显微镜、纳米颗粒跟踪分析、蛋白质印迹法及实时荧光定量聚合酶链反应（qRT-PCR）进行鉴定。采用qRT-PCR评估外泌体（miR-27a）向成骨细胞（MC3T3-E1细胞）递送miR-27a的效果。采用碱性磷酸酶染色、茜素红染色及qRT-PCR评估其对MC3T3-E1细胞成骨的影响。采用双荧光素酶报告基因（DLRTM）检测验证miR-27a靶向Dickkopf WNT信号通路抑制因子2（DKK2）是否为潜在机制，并通过MC3T3-E1细胞中的qRT-PCR、蛋白质印迹法及茜素红染色进一步验证该机制。最后，通过Sprague Dawley（SD）大鼠的GC-ONFH模型验证外泌体（miR-27a）对ONFH的保护作用。
透射电子显微镜、纳米颗粒跟踪分析、蛋白质印迹法及qRT-PCR检测显示外泌体（miR-27a）构建成功。外泌体（miR-27a）能有效将miR-27a递送至MC3T3-E1细胞并增强其成骨能力。DLRTM检测显示miR-27a通过直接靶向DKK2促进骨形成。动物实验的Micro-CT及苏木精-伊红染色结果显示尾静脉注射外泌体（miR-27a）改善了SD大鼠GC-ONFH模型的骨坏死情况。
外泌体（miR-27a）在一定程度上可促进骨再生并预防GC-ONFH。
