Supplementary Material for: Altered Long Noncoding RNA and Messenger RNA Expression in Experimental Intracerebral Hemorrhage - a Preliminary Study

Background/Aims: Functional recovery in the chronic phase is a difficult problem in intracerebral hemorrhage (ICH) treatment. Long noncoding RNAs (lncRNAs) are demonstrated to be involved in central nervous system (CNS) disorders. However, the roles of lncRNAs in post-ICH injury and repair are poorly understood, especially those that may be attributed to long-term neurological deficit. The present study depicted the lncRNA and messenger RNA (mRNA) profile by microarray at late stage after an experimental ICH. Methods: LncRNA and mRNA microarray was used to first identify differentially expressed genes. Gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to determine bio-functions and signaling pathways, with which differentially expressed genes are most closely related. Quantitative real-time polymerase chain reaction (PCR) was used to validate the results of microarray. Finally, the lncRNA-mRNA co-expression network was constructed to find the interaction of genes. Results: A total of 625 differentially expressed lncRNAs and 826 expressed mRNAs were identified. Altered genes were enriched in mitochon-drial matrix, G-protein coupled receptor signaling pathway, and olfactory transduction, which may be associated with ICH-induced pathophysiologic changes in the long term. A co-expression network profile based on 5 validated differentially expressed lncRNAs and 205 interacted mRNAs was composed of 210 nodes and 298 connections. Conclusion: Mitochondrial matrix, reduced G-protein coupled receptor activity, and impaired olfactory transduction may be involved in the sequelae following ICH. Further, these dysregulated lncRNAs and mRNAs may be the promising therapeutic targets to overcome obstacles in functional recovery following ICH.研究背景与目的：脑出血（intracerebral hemorrhage, ICH）慢性期的功能恢复是临床治疗的棘手难题。现有研究证实，长链非编码RNA（long noncoding RNAs, lncRNAs）参与中枢神经系统（central nervous system, CNS）疾病的病理生理过程，但目前对于lncRNAs在脑出血后损伤与修复环节中的作用机制尚不明确，尤其是与长期神经功能缺损相关的lncRNAs尚未得到充分阐释。本研究通过基因芯片技术，对实验性脑出血模型造模后期的lncRNAs与信使RNA（messenger RNA, mRNA）表达谱进行了系统性分析。 研究方法：首先采用lncRNA和mRNA基因芯片筛选差异表达基因；随后通过基因本体论（Gene Ontology, GO）富集分析与京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）通路分析，明确与差异表达基因高度相关的生物学功能及信号通路；采用实时定量聚合酶链式反应（Quantitative real-time polymerase chain reaction, PCR）对芯片检测结果进行验证；最后构建lncRNA-mRNA共表达网络，以探究基因间的相互作用关系。 研究结果：本研究共筛选得到625个差异表达的lncRNAs与826个差异表达的mRNA。富集分析显示，差异表达基因主要富集于线粒体基质、G蛋白偶联受体信号通路以及嗅觉转导通路，上述通路可能与脑出血诱导的长期病理生理改变密切相关。基于5个经验证的差异表达lncRNAs与205个互作mRNA构建的共表达网络，共包含210个节点与298条连接。 研究结论：线粒体基质功能异常、G蛋白偶联受体活性降低以及嗅觉转导功能受损，可能参与脑出血后的后遗症发生发展过程。此外，这些异常表达的lncRNAs与mRNA有望成为改善脑出血后功能恢复障碍的潜在治疗靶点。