Cluster analysis of long noncoding RNA after sciatic nerve injury and effect of long noncoding RNA MX1 on the migration and proliferation of rat Schwann cells
Luo Xuanxiang1, Pan Bin2, Feng Hu2
1Department of Orthopedic,Nanjing Gaochun People's Hospital, Nanjing 211300, China; 2Department of Orthopedic, Affiliated Hospital of Xuzhou Medical University, Xuzhou 221000, China
Abstract：Objective This study was performed to investigate the differentially expressed genes of long noncoding RNA after sciatic nerve injury and effects of long noncoding RNA MX1 on the migration and proliferation of rat Schwann cells. Methods Twenty-four 10-week-old male SD rats without specific pathogens were randomly divided into 4 groups (6 rats per group), namely, 0 d (T0), 3 d (T1), 7 d (T2), and 14 d (T3). The rats were selected to establish animal models of sciatic nerve injury. Animal models of sciatic nerve injury were established. RNA was extracted from the residual tissue of the injured sciatic nerve on days 0, 3, 7, and 14 after surgery according to the time point of the different groups. RNA gene chips were prepared, and heatmap clustering analysis was performed to screen out the differentially expressed genes at each time point. The gene LncRNA MX1 with significant differentially expressed genes was selected. Icell-r030 rat Schwann cells were cultured and divided into the control group, control siRNA group(si RNA group), and LncRNA MX1 siRNA group(si RNA-MX1 group). Transfection was performed with corresponding reagents. After transfection, the expression of LncRNA MX1mRNA in the three groups of Schwann cells was detected by real-time fluorescence quantitative polymerase chain reaction (qRT-PCR). Migration ability of the Schwann cells was detected by Transwell chamber, and the proliferation ability of the Schwann cells was detected by 5-ethynyl-2'-deoxyuridine (EDU) assay. Results The rats in each group were modeled successfully, and no animals died during the experiment. Compared with the T0 group, a total of 3 066 differentially expressed LncRNA genes were found in the T1 group, including 1 634 upregulated and 1 432 downregulated LncRNA genes. A total of 2 498 LncRNA genes were differentially expressed in the T2 group, including 1 634 upregulated and 864 downregulated LncRNA genes. In the T3 group, 3 567 LncRNA genes were differentially expressed, among which 1 643 LncRNA genes were upregulated, and 1 924 were downregulated. Fold change value of LncRNA MX1 in the differentially expressed LncRNA genes in each group was larger, and the differentially expressed LncRNA MX1 was significant. qRT-PCR results of the rat Schwann cells showed that the relative expression level of LncRNA MX1 in the siRNA-MX1 group was 1.0±0.2 after transfection with LncRNA MX1 siRNA, which was lower than those of the control group (2.3±0.2) and siRNA group (2.2±0.2). The difference was statistically significant (F=78.47, P<0.001). No significant difference was found between the control group and the siRNA group (P>0.05). The results of migration and proliferation experiments showed that the number of cell migration in the siRNA-MX1 group was 24.1±4.2. The ratio of EDU positive cells to DAPI positive cells was 27.5%±2.8%, which was lower than those of the control group (50.3±7.8; 44.1%±7.2%) and siRNA group (49.2±6.2; 41.8%±7.0%). The differences were statistically significant (F=93.15, 121.26, all P values<0.001). No significant difference was found between the control group and the siRNA group (all P values>0.05). Conclusion The differential expression of LncRNA MX1 after sciatic nerve injury in rats is significant. Downregulation of LncRNA MX1 expression in the rat Schwann cells can significantly reduce cell migration and proliferation.
罗选翔, 潘彬, 冯虎. 坐骨神经损伤后长链非编码RNA的聚类分析及长链非编码RNA MX1对大鼠雪旺细胞迁移、增殖能力的影响[J]. 中华解剖与临床杂志, 2022, 27(8): 574-580.
Luo Xuanxiang, Pan Bin, Feng Hu. Cluster analysis of long noncoding RNA after sciatic nerve injury and effect of long noncoding RNA MX1 on the migration and proliferation of rat Schwann cells. Chinese Journal of Anatomy and Clinics, 2022, 27(8): 574-580.
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2022, Vol. 27 
