中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2020, Vol. 15 ›› Issue (1): 86-95.doi: 10.4103/1673-5374.264469

• 原著:脑损伤修复保护与再生 • 上一篇    下一篇

新生儿缺氧缺血性脑损伤发病相关关键mRNA单核苷酸多态性靶点筛选与RNA测序

  

  • 出版日期:2020-01-15 发布日期:2020-05-20
  • 基金资助:

    云南省科技创新研究团队项目；中国国家自然科学基金项目（81601074）；四川省科学基金项目（2017SZ0145）

Single-nucleotide polymorphism screening and RNA sequencing of key messenger RNAs associated with neonatal hypoxic-ischemia brain damage

Liu-Lin Xiong1, 2, Lu-Lu Xue3, Mohammed Al-Hawwas2, Jin Huang3, Rui-Ze Niu3, Ya-Xin Tan3, Yang Xu4, Ying-Ying Su4, Jia Liu3,#br# Ting-Hua Wang3, 4#br#   

  1. 1 Department of Anesthesiology, National Traditional Chinese Medicine Clinical Research Base and Western Medicine Translational Medicine Research Center, Affiliated Traditional Chinese Medicine Hospital, Southwest Medical University, Luzhou, Sichuan Province, China

    2 School of Pharmacy and Medical Sciences, Division of Health Sciences, University of South Australia, Adelaide, South Australia, Australia

    3 Department of Animal Zoology, Kunming Medical University, Kunming, Yunnan Province, China

    4 Institute of Neurological Disease, Department of Anesthesiology, Translational Neuroscience Center, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China

  • Online:2020-01-15 Published:2020-05-20
  • Contact: Jia Liu, PhD, liujiaaixuexi@126.com; Ting-Hua Wang, PhD, wangtinghua@vip.163.com.
  • Supported by:
    This study was supported by the program Innovative Research Team in Science and Technology in Yunnan Province of China (to THW); the National Natural Science Foundation of China, No. 81601074, Sichuan Provincial Scientific Foundation Grant of China, No. 2017SZ0145.

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摘要：

单核苷酸多态性是由单个核苷酸的变异所引起的DNA序列多态性，其可影响疾病易感性，但是目前单核苷酸多态性与新生儿缺氧缺血性脑损伤发病的关系尚不确定。(1)实验使用7d的新生大鼠建立新生儿缺氧缺血性脑损伤模型，并使用RNA测序在缺氧缺血性脑病中分析mRNA和单核苷酸多态性的表达谱。通过基因本体论和途径分析鉴定出与新生儿缺氧缺血性脑损伤相关的表现出单核苷酸多态性的基因，并鉴定这些基因引发疾病的可能机制；(2)结果发现有89个表现出单核苷酸多态性的上调基因，且其主要位于染色体1和2上，且主要参与血管生成、伤口愈合、谷氨酸能突触以及钙激活氯离子通道的过程。信号通路分析表明，差异表达的基因在谷氨酸能突触、长期抑制和催产素信号通路中起作用。通过Pubmed进行高通量筛选和单核苷酸多态性基因测序的交叉分析发现3种新的参与新生儿缺氧缺血性脑损伤的基因，CSRNP1，DUSP5和LRRC25，且经实时定量PCR能这3种基因也在氧糖剥夺人胚胎大脑皮质神经元表达上调；(3)实验结果说明表现出单核苷酸多态性的基因CSRNP1，DUSP5和LRRC25，可能参与新生儿缺氧缺血性脑损伤的发生和发展，且其可能成为未来治疗新生儿缺氧缺血性脑损伤的新靶点。动物实验于2017年2月5日由昆明医科大学动物护理和使用委员会批准（批准号kmmu2019038），人胚胎原代皮质神经元相关研究于2015年9月30日经昆明医科大学伦理委员会批准（批准号2015-9）。

orcid: 0000-0003-2012-8936 (Ting-Hua Wang)
         0000-0001-5678-7437 (Jia Liu)

关键词:

mRNA, 单核苷酸多态性, 新生儿缺氧缺血性脑损伤, RNA测序, CSRNP1, DUSP5, LRRC25, 神经再生

Abstract: A single-nucleotide polymorphism (SNP) is an alteration in one nucleotide in a certain position within a genome. SNPs are associated with disease susceptibility. However, the influences of SNPs on the pathogenesis of neonatal hypoxic-ischemic brain damage remain elusive. Seven- day-old rats were used to establish a hypoxic ischemic encephalopathy model. SNPs and expression profiles of mRNAs were analyzed in hypoxic ischemic encephalopathy model rats using RNA sequencing. Genes exhibiting SNPs associated with hypoxic ischemic encephalopathy were identified and studied by gene ontology and pathway analysis to identify their possible involvement in the disease mechanism. We identified 89 up-regulated genes containing SNPs that were mainly located on chromosome 1 and 2. Gene ontology analysis indicated that the up-regulated genes containing SNPs are mainly involved in angiogenesis, wound healing and glutamatergic synapse and biological processing of calcium-activated chloride channels. Signaling pathway analysis indicated that the differentially expressed genes play a role in glutamatergic synapses, long-term depression and oxytocin signaling. Moreover, intersection analysis of high throughput screening following PubMed retrieval and RNA sequencing for SNPs showed that CSRNP1, DUSP5 and LRRC25 were most relevant to hypoxic ischemic encephalopathy. Significant up-regulation of genes was confirmed by quantitative real-time polymerase chain reaction analysis of oxygen-glucose-deprived human fetal cortical neurons. Our results indicate that CSRNP1, DUSP5 and LRRC25, containing SNPs, may be involved in the pathogenesis of hypoxic ischemic encephalopathy. These findings indicate a novel direction for further hypoxic ischemic encephalopathy research. This animal study was approved on February 5, 2017 by the Animal Care and Use Committee of Kunming Medical University, Yunnan Province, China (approval No. kmmu2019038). Cerebral tissue collection from a human fetus was approved on September 30, 2015 by the Ethics Committee of Kunming Medical University, China (approval No. 2015-9).

Key words: CSRNP1, DUSP5, gene ontology analysis, human fetal cortical neurons, LRRC25, mRNA, neonatal hypoxic ischemic encephalopathy, pathogenesis, signaling pathway analysis