Jin Fa, Ou Weiyang, Wei Boyang, Fan Haiyan, Wei Chengcong, Fang Dazhao, Li Guangxu, Liu Wenchao, Liu Jiahui, Jin Lei, He Xuying, Duan Chuanzhi
Neurosurgery Center, Department of Cerebrovascular Surgery, The National Key Clinical Specialty, The Engineering Technology Research Center of Education Ministry of China on Diagnosis and Treatment of Cerebrovascular Disease, Guangdong Provincial Key Laboratory on Brain Function Repair and Regeneration, The Neurosurgery Institute of Guangdong Province, Zhujiang Hospital, Southern Medical University, Guangzhou, 510282, People's Republic of China.
J Inflamm Res. 2021 Jun 22;14:2667-2680. doi: 10.2147/JIR.S315281. eCollection 2021.
Ischemic stroke is one of the leading causes of mortality and disability worldwide. Following stroke, there is secondary neuroinflammation that promotes further injury. Identifying the long non-coding RNA (lncRNA) involved in neuroinflammation after cerebral ischemic stroke will promote the discovery of potential therapeutic targets.
We identified differentially expressed genes from genome-wide RNA-seq profiles of mice with focal ischemia using Gene Ontology Term Enrichment, Kyoto Encyclopedia of Genes and Genomes, and Gene Set Enrichment analyses. Immune cell infiltration deconvolution, protein-protein interaction network construction, and co-expression network analyses were also used to screen lncRNAs. In further experiments, lncRNA knockdown animal models were developed by intraventricular injection of the antisense oligonucleotide before performing middle cerebral artery occlusion (MCAO). An enzyme-linked immunosorbent assay was performed to measure the level of cytokines. Hematoxylin-eosin staining and immunohistochemical staining were used to observe the changes in morphology.
Enrichment analysis revealed that differential mRNAs induced neuroinflammation after MCAO. Immune deconvolution showed that the proportion of microglia gradually increased while monocytes decreased within 24 h. We identified six hub lncRNAs (, , , , , and ) that were highly correlated with activated-microglia mRNAs (cor > 0.8). We found that Neat1 had the highest correlation coefficient with pro-inflammatory factor mRNA levels. In vivo experiments demonstrated that had abnormally high expression after MCAO. Knockdown of could significantly alleviate brain damage by reducing the number of activated microglia and reducing their release of proinflammatory cytokines.
We identified inflammation-associated lncRNA as crucial, which means it is a potential target for ischemic stroke treatment.
缺血性中风是全球范围内导致死亡和残疾的主要原因之一。中风后会出现继发性神经炎症，进一步加重损伤。确定脑缺血性中风后参与神经炎症的长链非编码RNA（lncRNA）将有助于发现潜在的治疗靶点。
我们使用基因本体术语富集、京都基因与基因组百科全书和基因集富集分析，从局灶性缺血小鼠的全基因组RNA测序图谱中鉴定差异表达基因。还使用免疫细胞浸润反卷积、蛋白质-蛋白质相互作用网络构建和共表达网络分析来筛选lncRNA。在进一步的实验中，在进行大脑中动脉闭塞（MCAO）之前，通过脑室内注射反义寡核苷酸建立lncRNA敲低动物模型。进行酶联免疫吸附测定以测量细胞因子水平。苏木精-伊红染色和免疫组织化学染色用于观察形态学变化。
富集分析显示，差异mRNA在MCAO后诱导神经炎症。免疫反卷积显示，在24小时内，小胶质细胞的比例逐渐增加，而单核细胞的比例下降。我们鉴定出六个与活化小胶质细胞mRNA高度相关的枢纽lncRNA（， ， ， ， ，和 ）（相关性>0.8）。我们发现Neat1与促炎因子mRNA水平的相关系数最高。体内实验表明，在MCAO后表达异常高。敲低 可通过减少活化小胶质细胞的数量并减少其促炎细胞因子的释放，显著减轻脑损伤。
我们确定炎症相关lncRNA 至关重要，这意味着它是缺血性中风治疗的潜在靶点。
