多芯片整合分析腰椎间盘突出症发生相关的分子通路及关键基因研究
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摘要
目的利用多芯片整合的生物信息学方法系统性地分析严重退化和轻度退化的椎间盘(IVD)组织的基因表达差异,并探讨潜在的分子机制。方法从基因表达综合数据库(GEO)下载3组腰椎间盘突出症(LDH)的原始芯片数据。利用R语言SVA软件的Combat函数消除表达数据间的异质性。通过差异、基因功能富集及关联网络分析进一步探索差异基因的蛋白互作(PPI)网络、基因本体(GO)功能和分子通路。结果基于差异分析,本研究得到149个差异基因。相关分子通路包含HSA03010:核糖体活动(富集数目13,P=8.34×10-9)、HSA000 190:氧化磷酸化(富集数目12,P=7.30×10-8)、HSA045 12:细胞外基质反应(富集数目10,P=1.20×10-4。通过PPI网络分析,连接度较高的基因UBA52,RPLP0,RPL3,RPLP2和RPL27被进一步筛选出来。结论核糖体活动、氧化磷酸化及细胞外基质反应可能是LDH疾病潜在的分子机制,而UBA52,RPLP0,RPL3,RPLP2和RPL27被认为是LDH疾病发生、发展的关键基因。
Objective To identify the difference of gene expression between severely degraded and mildly degenerated intervertebral disc(IVD)tissues using multi-chip integrated bioinformatics methods,and explore the potential molecular mechanism.Methods Three groups(GSE23130,GSE17077 and GSE15227)of microarray data of lumbar disc herniation(LDH)were downloaded from the Gene Expression Omnibus(GEO)database.Subsequently,the Combat function and adjustment for batch of SVA package was used to remove the data heterogeneity.In addition,extensive differential expression analysis and network analysis methods were used to access the protein-protein interaction(PPI)network,gene ontology(GO)terms and pathway enrichment among the differential expression genes.Results The results of differential expression analysis showed that 149 differentially expressed genes were identified.The related molecular pathways including HSA03010:ribosome(count:13,P=8.34×10-9),HSA000 190:oxidative phosphorylation(count:12,P=7.30×10-8),and HAS045 12:ECM-receptor interaction(count:10,P=1.20×10-4)were significantly enriched among differential expression genes.Finally,the hub genes including UBA52,RPLP0,RPL3,RPLP2 and RPL27 of LDH were identified from PPI network.Conclusion The maps of ribosome,oxidative phosphorylation and ECM-receptor interaction are considered as potential mechanisms,and UBA52,RPLP0,RPL3,RPLP2 and RPL27 genes may be implicated in the occurrence and development of LDH.
Objective To identify the difference of gene expression between severely degraded and mildly degenerated intervertebral disc(IVD)tissues using multi-chip integrated bioinformatics methods,and explore the potential molecular mechanism.Methods Three groups(GSE23130,GSE17077 and GSE15227)of microarray data of lumbar disc herniation(LDH)were downloaded from the Gene Expression Omnibus(GEO)database.Subsequently,the Combat function and adjustment for batch of SVA package was used to remove the data heterogeneity.In addition,extensive differential expression analysis and network analysis methods were used to access the protein-protein interaction(PPI)network,gene ontology(GO)terms and pathway enrichment among the differential expression genes.Results The results of differential expression analysis showed that 149 differentially expressed genes were identified.The related molecular pathways including HSA03010:ribosome(count:13,P=8.34×10-9),HSA000 190:oxidative phosphorylation(count:12,P=7.30×10-8),and HAS045 12:ECM-receptor interaction(count:10,P=1.20×10-4)were significantly enriched among differential expression genes.Finally,the hub genes including UBA52,RPLP0,RPL3,RPLP2 and RPL27 of LDH were identified from PPI network.Conclusion The maps of ribosome,oxidative phosphorylation and ECM-receptor interaction are considered as potential mechanisms,and UBA52,RPLP0,RPL3,RPLP2 and RPL27 genes may be implicated in the occurrence and development of LDH.
引文
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[2]NOURI A,TETREAULT L,SINGH A,et al.Degenerative cervical myelopathy:epidemiology,genetics,and pathogenesis[J].Spine(Phila Pa 1976),2015,40(12):E675-693.
[3]LEE C H,CHUNG C K,KIM C H,et al.Health care burden of spinal diseases in the republic of korea:analysis of a nationwide database from 2012through 2016[J].Neurospine,2018,15(1):66-76.
[4]MARTIROSYAN N L,PATEL A A,CAROTENUTO A,et al.Genetic alterations in intervertebral disc disease[J].Front Surg,2016,3:59.
[5]ZHANG Y G,ZHANG F,SUN Z,et al.A controlled case study of the relationship between environmental risk factors and apoptotic gene polymorphism and lumbar disc herniation[J].Am J Pathol,2013,182(1):56-63.
[6]PERERA R S,DISSANAYAKE P H,SENARATH U,et al.Variants of ACAN are associated with severity of lumbar disc herniation in patients with chronic low back pain[J].PLoSOne,2017,12(7):e181580.
[7]KURZAWSKI M,RUT M,DZIEDZIEJKO V,et al.Common missense variant of SCN9Agene is associated with pain intensity in patients with chronic pain from disc herniation[J].Pain Med,2018,19(5):1010-1014.
[8]SADOWSKA A,TOULI E,HITZL W,et al.Inflammaging in cervical and lumbar degenerated intervertebral discs:analysis of proinflammatory cytokine and TRP channel expression[J].Eur Spine J,2018,27(3):564-577.
[9]CLOUGH E,BARRETT T.The gene expression omnibus database[J].Methods Mol Biol,2016,1418:93-110.
[10]GRUBER H E,HOELSCHER G L,INGRAM J A,et al.Genome-wide analysis of pain-,nerve-and neurotrophin-related gene expression in the degenerating human annulus[J].Mol Pain,2012,8:63.
[11]FU G,SAUNDERS G,STEVENS J.Holm multiple correction for large-scale gene-shape association mapping[J].BMCGenet,2014,15(Suppl 1):S5.
[12]SERBAN N,JIANG H.Multilevel functional clustering analysis[J].Biometrics,2012,68(3):805-814.
[13]LEEK J T,JOHNSON W E,PARKER H S,et al.The sva package for removing batch effects and other unwanted variation in high-throughput experiments[J].Bioinformatics,2012,28(6):882-883.
[14]HUANG D W,SHERMAN B T,TAN Q,et al.DAVIDBioinformatics Resources:expanded annotation database and novel algorithms to better extract biology from large gene lists[J].Nucleic Acids Res,2007,35(Web Server issue):W169-175.
[15]SZKLARCZYK D,MORRIS J H,COOK H,et al.The STRING database in 2017:quality-controlled protein-protein association networks,made broadly accessible[J].Nucleic Acids Res,2017,45(Database issue):D362-368.
[16]KOBAYASHI S,UCHIDA K,YAYAMA T,et al.Motor neuron involvement in experimental lumbar nerve root compression:a light and electron microscopic study[J].Spine(Phila Pa 1976),2007,32(6):627-634.
[17]ALAMIN T F,MUNOZ M,ZAGEL A,et al.Ribosomal PCR assay of excised intervertebral discs from patients undergoing single-level primary lumbar microdiscectomy[J].Eur Spine J,2017,26(8):2038-2044.
[18]SLOMNICKI L P,PIETRZAK M,VASHISHTA A,et al.Requirement of neuronal ribosome synthesis for growth and maintenance of the dendritic tree[J].J Biol Chem,2016,291(11):5721-5739.
[19]POIROT O,TIMSIT Y.Neuron-like networks between ribosomal proteins within the ribosome[J].Sci Rep,2016,6:26485.
[20]MESKAUSKAS A,PETROV A N,DINMAN J D.Identification of functionally important amino acids of ribosomal protein L3by saturation mutagenesis[J].Mol Cell Biol,2005,25(24):10863-10874.
[21]KOBAYASHI M,OSHIMA S,MAEYASHIKI C,et al.The ubiquitin hybrid gene UBA52regulates ubiquitination of ribosome and sustains embryonic development[J].Sci Rep,2016,6:36780.
[22]ARTERO-CASTRO A,PEREZ-ALEA M,FELICIANO A,et al.Disruption of the ribosomal P complex leads to stressinduced autophagy[J].Autophagy,2015,11(9):1499-1519.
[23]XIAO L,DING M,FERNANDEZ A,et al.Curcumin alleviates lumbar radiculopathy by reducing neuroinflammation,oxidative stress and nociceptive factors[J].Eur Cell Mater,2017,33:279-293.
[24]CHANG F,ZHANG T,GAO G,et al.Therapeutic effect of percutaneous endoscopic lumbar discectomy on lumbar disc herniation and its effect on oxidative stress in patients with lumbar disc herniation[J].Exp Ther Med,2018,15(1):295-299.
[25]GUTERL C C,SEE E Y,BLANQUER S B,et al.Challenges and strategies in the repair of ruptured annulus fibrosus[J].Eur Cell Mater,2013,25:1-21.
[26]HIROSE Y,CHIBA K,KARASUGI T,et al.A functional polymorphism in THBS2that affects alternative splicing and MMP binding is associated with lumbar-disc herniation[J].Am J Hum Genet,2008,82(5):1122-1129.