源自MiR-133b修饰的脂肪间充质干细胞的外泌体对脊髓损伤后神经功能恢复的影响。
Effect of exosomes derived from MiR-133b-modified ADSCs on the recovery of neurological function after SCI.
作者信息
Ren Z-W, Zhou J-G, Xiong Z-K, Zhu F-Z, Guo X-D
机构信息
Department of Orthopaedics, Affiliated Concord Hospital of Tongji Medical College, Huazhong University of Science And Technology, Wuhan, China.
出版信息
Eur Rev Med Pharmacol Sci. 2019 Jan;23(1):52-60. doi: 10.26355/eurrev_201901_16747.
OBJECTIVE
To investigate the recovery effect of exosomes derived from micro ribonucleic acid (miR)-133b-modified adipose-derived stem cells (ADSCs) on neurological function after spinal cord injury (SCI) and its mechanism.
MATERIALS AND METHODS
The SCI model of rats was used and divided into the following 5 groups: sham-operation group, 4 d SCI group, phosphate-buffered saline (PBS) group, miR-control group and miR-133b group. At 96 h after operation, rats were euthanatized, and spinal tissues were removed. Next, the level of miR-133b was detected via reverse transcription-polymerase chain reaction (RT-PCR), and the expression of RhoA protein was measured via Western blotting. Moreover, expressions of proteins associated with the axon regeneration pathway, including phosphorylated-cAMP-response element binding protein (p-CREB), CREB, phosphorylated-signal transducer and activator of transcription 3 (p-STAT3) and STAT3, along with expressions of neurofilament (NF), growth associated protein 43 (GAP43), glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP), were tested by Western blotting and immunofluorescence staining.
RESULTS
The miR-133b mimics significantly upregulated the expression of miR-133b in adipose-derived stem cells (ADSCs), compared to blank group (p<0.05). The expression of miR-133b was significantly decreased in 4 d SCI group compared with that in sham-operation group (p<0.001). The RhoA expression was statistically increased in 4 d SCI group compared with that in sham-operation group (p<0.001), and it was partially impaired by using miR-133b compared with that in 4 d SCI group (p<0.001). Expressions of NF, GAP43, GFAP and MBP were remarkably higher in 4 d SCI group than those in sham-operation group (p<0.01), and they were also significantly increased in miR-133b group than those in 4 d SCI group (p<0.01). Besides, our data showed a significant increase of p-CREB/CREB, p-STAT3/STAT3, NF, GAP-43, GFAP and MBP in miR-133b group compared to those in 4 d SCI group, with statistical reduction of RhoA (p<0.05).
CONCLUSIONS
We showed that exosomes derived from miR-133b-modified ADSCs can significantly promote the recovery of neurological function of SCI animals through affecting the signaling pathway related to axon regeneration and expressions of NF, GAP-43, GFAP and MBP.
目的
探讨微小核糖核酸（miR）-133b修饰的脂肪来源干细胞（ADSCs）分泌的外泌体对脊髓损伤（SCI）后神经功能的恢复作用及其机制。
材料与方法
采用大鼠SCI模型，分为以下5组：假手术组、SCI 4 d组、磷酸盐缓冲液（PBS）组、miR对照组和miR-133b组。术后96 h处死大鼠，取出脊髓组织。接下来，通过逆转录-聚合酶链反应（RT-PCR）检测miR-133b水平，通过蛋白质免疫印迹法检测RhoA蛋白的表达。此外，通过蛋白质免疫印迹法和免疫荧光染色检测与轴突再生途径相关的蛋白质表达，包括磷酸化环磷酸腺苷反应元件结合蛋白（p-CREB）、CREB、磷酸化信号转导和转录激活因子3（p-STAT3）和STAT3，以及神经丝（NF）、生长相关蛋白43（GAP43）、胶质纤维酸性蛋白（GFAP）和髓鞘碱性蛋白（MBP）的表达。
结果
与空白组相比，miR-133b模拟物显著上调了脂肪来源干细胞（ADSCs）中miR-133b的表达（p<0.05）。与假手术组相比，SCI 4 d组中miR-133b的表达显著降低（p<0.001）。与假手术组相比，SCI 4 d组中RhoA表达有统计学意义的增加（p<0.001），与SCI 4 d组相比，使用miR-133b后RhoA表达部分受损（p<0.001）。与假手术组相比，SCI 4 d组中NF、GAP43、GFAP和MBP的表达显著更高（p<0.01），与SCI 4 d组相比，miR-133b组中它们的表达也显著增加（p<0.01）。此外，我们的数据显示，与SCI 4 d组相比，miR-133b组中p-CREB/CREB、p-STAT3/STAT3、NF、GAP-43、GFAP和MBP显著增加，RhoA有统计学意义的降低（p<0.05）。
结论
我们发现，miR-133b修饰的ADSCs分泌的外泌体可通过影响与轴突再生相关的信号通路以及NF、GAP-43、GFAP和MBP的表达，显著促进SCI动物神经功能的恢复。
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