Zhang Yu-Xin, Wang Sai-Nan, Chen Jing, Hu Jian-Guo, Lü He-Zuo
Clinical Laboratory, The First Affiliated Hospital of Bengbu Medical College, Bengbu, China.
Anhui Key Laboratory of Tissue Transplantation, The First Affiliated Hospital of Bengbu Medical College, Bengbu, China.
PeerJ. 2020 Jan 3;8:e8367. doi: 10.7717/peerj.8367. eCollection 2020.
Recent studies have found that probenecid has neuroprotective and reparative effects on central nervous system injuries. However, its effect on genome-wide transcription in acute spinal cord injury (SCI) remains unknown. In the present study, RNA sequencing (RNA-Seq) is used to analyze the effect of probenecid on the local expression of gene transcription 8 h after spinal injury.
An Infinite Horizon impactor was used to perform contusive SCI in mice. The SCI model was made by using a rod (1.3 mm diameter) with a force of 50 Kdynes. Sham-operated mice only received a laminectomy without contusive injury. The injured mice were randomly assigned into either the control (SCI_C) or probenecid injection (SCI_P) group. In the latter group, the probenecid drug was intraperitoneally injected (0.5 mg/kg) immediately following injury. Eight hours after the injury or laminectomy, the spinal cords were removed from the mice in both groups. The total RNAs were extracted and purified for library preparation and transcriptome sequencing. Differential gene expressions (DEGs) of the three groups-sham, SCI_C and SCI_P-were analyzed using a DESeq software. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of DEGs were performed using a GOseq R package and KOBAS software. Real-time quantitative reverse-transcriptase polymerase chain reaction was used to validate RNA-Seq results.
RNA-Seq showed that, compared to the SCI_C group, the number of DEGs was 641 in the SCI_P group (286 upregulated and 355 downregulated). According to GO analysis, DEGs were most enriched in extracellular matrix (ECM), collagen trimer, protein bounding and sequence specific DNA binding. KEGG analysis showed that the most enriched pathways included: cell adhesion molecules, Leukocyte transendothelial migration, ECM-receptor interactions, PI3K-Akt signaling pathways, hematopoietic cell lineages, focal adhesions, the Rap1 signaling pathway, etc. The sequence data have been deposited into the Sequence Read Archive (https://www.ncbi.nlm.nih.gov/sra/PRJNA554464).
近期研究发现，丙磺舒对中枢神经系统损伤具有神经保护和修复作用。然而，其对急性脊髓损伤（SCI）全基因组转录的影响尚不清楚。在本研究中，采用RNA测序（RNA-Seq）分析丙磺舒对脊髓损伤后8小时基因转录局部表达的影响。
使用无限视野撞击器对小鼠进行挫伤性SCI。通过使用直径1.3毫米的棒以50达因的力制作SCI模型。假手术小鼠仅接受椎板切除术，无挫伤性损伤。将受伤小鼠随机分为对照组（SCI_C）或丙磺舒注射组（SCI_P）。在后者组中，受伤后立即腹腔注射丙磺舒药物（0.5毫克/千克）。损伤或椎板切除术后8小时，从两组小鼠中取出脊髓。提取并纯化总RNA用于文库制备和转录组测序。使用DESeq软件分析假手术、SCI_C和SCI_P三组的差异基因表达（DEG）。使用GOseq R包和KOBAS软件对DEG进行基因本体论（GO）和京都基因与基因组百科全书（KEGG）富集分析。使用实时定量逆转录聚合酶链反应验证RNA-Seq结果。
RNA-Seq显示，与SCI_C组相比，SCI_P组的DEG数量为641个（上调286个，下调355个）。根据GO分析，DEG在细胞外基质（ECM）、三聚体胶原蛋白、蛋白质结合和序列特异性DNA结合中富集最多。KEGG分析表明，最富集的途径包括：细胞粘附分子、白细胞跨内皮迁移、ECM-受体相互作用、PI3K-Akt信号通路、造血细胞系、粘着斑、Rap1信号通路等。序列数据已存入序列读取存档库（https://www.ncbi.nlm.nih.gov/sra/PRJNA554464）。
