Pan Bin, Shi Zhong-Ju, Yan Jia-Yin, Li Jia-He, Feng Shi-Qing
Department of Orthopedics, Tianjin Medical University General Hospital, Tianjin, China.
Neural Regen Res. 2017 Dec;12(12):2084-2091. doi: 10.4103/1673-5374.221168.
Schwann cells play a critical role in peripheral nerve regeneration through dedifferentiation and proliferation. In a previous study, we performed microarray analysis of the sciatic nerve after injury. Accordingly, we predicted that long non-coding RNA NONMMUG014387 may promote Schwann cell proliferation after peripheral nerve injury, as bioinformatic analysis revealed that the target gene of NONMMUG014387 was collagen triple helix repeat containing 1 (Cthrc1). Cthrc1 may promote cell proliferation in a variety of cells by activating Wnt/PCP signaling. Nonetheless, bioinformatic analysis still needs to be verified by biological experiment. In this study, the candidate long non-coding RNA, NONMMUG014387, was overexpressed in mouse Schwann cells by recombinant adenovirus transfection. Plasmid pHBAd-MCMV-GFP-NONMMUG014387 and pHBAd-MCMV-GFP were transfected into Schwann cells. Schwann cells were divided into three groups: control (Schwann cells without intervention), Ad-GFP (Schwann cells with GFP overexpression), and Ad-NONMMUGO148387 (Schwann cells with GFP and NONMMUGO148387 overexpression). Cell Counting Kit-8 assay was used to evaluate proliferative capability of mouse Schwann cells after NONMMUG014387 overexpression. Polymerase chain reaction and western blot assay were performed to investigate target genes and downstream pathways of NONMMUG014387. Cell proliferation was significantly increased in Schwann cells overexpressing lncRNA NONMMUG014387 compared with the other two groups. Further, compared with the control group, mRNA and protein levels of Cthrc1, Wnt5a, ROR2, RhoA, Rac1, JNK, and ROCK were visibly up-regulated in the Ad-NONMMUGO148387 group. Our findings confirm that long non-coding RNA NONMMUG014387 can promote proliferation of Schwann cells surrounding the injury site through targeting Cthrc1 and activating the Wnt/PCP pathway.
施万细胞通过去分化和增殖在周围神经再生中发挥关键作用。在先前的一项研究中，我们对损伤后的坐骨神经进行了微阵列分析。据此，我们预测长链非编码RNA NONMMUG014387可能促进周围神经损伤后施万细胞的增殖，因为生物信息学分析显示NONMMUG014387的靶基因是含胶原三螺旋重复序列1（Cthrc1）。Cthrc1可能通过激活Wnt/PCP信号通路促进多种细胞的增殖。尽管如此，生物信息学分析仍需通过生物学实验进行验证。在本研究中，通过重组腺病毒转染在小鼠施万细胞中过表达候选长链非编码RNA NONMMUG014387。将质粒pHBAd-MCMV-GFP-NONMMUG014387和pHBAd-MCMV-GFP转染到施万细胞中。施万细胞分为三组：对照组（未干预的施万细胞）；Ad-GFP组（过表达GFP的施万细胞）；Ad-NONMMUGO148387组（过表达GFP和NONMMUGO148387的施万细胞）。使用细胞计数试剂盒-8检测法评估NONMMUG014387过表达后小鼠施万细胞的增殖能力。进行聚合酶链反应和蛋白质印迹分析以研究NONMMUG014387的靶基因和下游通路。与其他两组相比，过表达lncRNA NONMMUG014387的施万细胞中细胞增殖显著增加。此外，与对照组相比，Ad-NONMMUGO148387组中Cthrc1、Wnt5a、ROR2、RhoA、Rac1、JNK和ROCK的mRNA和蛋白质水平明显上调。我们的研究结果证实，长链非编码RNA NONMMUG014387可通过靶向Cthrc1并激活Wnt/PCP通路促进损伤部位周围施万细胞的增殖。
