甲基化测序和转录组整合分析黄韧带肥厚的分子机制  

Molecular mechanisms of ligament flavum hypertrophy:analysis based on methylation sequencing and transcriptome integration

作　　者：何杨 唐步源 卢昌怀 He Yang;Tang Buyuan;Lu Changhuai(Hunan University of Chinese Medicine,Changsha 410208,Hunan Province,China;No.2 Spinal Ward,Department of Orthopedics and Traumatology,the First Hospital of Traditional Chinese Medicine in Changde,Changde 415000,Hunan Province,China)

机构地区：[1]湖南中医药大学,湖南省长沙市410208 [2]常德市第一中医医院骨伤科脊柱二病区,湖南省常德市415000

出　　处：《中国组织工程研究》2025年第5期1013-1020,共8页Chinese Journal of Tissue Engineering Research

基　　金：湖南省自然科学基金面上项目(2021JJ30049),项目负责人:卢昌怀;湖南省研究生科研创新项目(2022CX201),项目负责人:何杨。

摘　　要：背景:黄韧带肥厚是腰椎管狭窄症的最主要病因,是多个病理因素共同作用的结果,目前黄韧带肥厚的分子机制、作用通路尚不明确,缺乏有效的非手术治疗方案。目的:使用甲基化测序和转录组整合分析方法,研究黄韧带肥厚发生发展的分子机制。方法:收集5个正常黄韧带组织和5个肥厚黄韧带组织,采用甲基化测序记录异常甲基化位点和甲基化状态,采用转录组整合分析记录异常表达的基因,取二者交集中甲基化水平与表达水平负相关的基因。采用GO和KEGG富集分析研究差异基因表达的主要功能通路和分子功能。使用PPI互作分析筛选调控黄韧带肥厚的关键基因。结果与结论:对两组黄韧带行甲基化测序发现高甲基化位点37173个,低甲基化位点10583个,转录组整合分析发现异常表达基因720个,其中463个上调基因、257个下调基因。两者重叠基因383个,其中192个基因甲基化水平与表达水平负相关。GO功能通路分析结果显示分子功能富集到10个条目,细胞组分富集到15个条目,生物学途径富集到30个条目。KEGG通路富集分析结果显示,192个基因主要富集到PI3K-Akt信号通路、Rap1信号通路、Focal adhesion信号通路、信号通路等9条通路。PPI互作分析筛选出PPARG、EGFR、CNR1、TNF和COL11A25个基因可能是调控黄韧带肥厚的关键基因,主要通过调控组织纤维化、细胞增殖分化、炎症因子水平、胶原纤维表达水平等途径影响黄韧带肥厚的发生发展。BACKGROUND:Ligament flavum hypertrophy is the main cause of lumbar spinal stenosis,which is the result of multiple pathological factors working together.Currently,the molecular mechanism and pathway of action of ligament flavum hypertrophy are unclear,and there is a lack of effective non-surgical treatment options.OBJECTIVE:To investigate the molecular mechanisms of ligament flavum hypertrophy using methylation sequencing and transcriptome integration analysis methods.METHODS:Five normal ligament flavum tissue samples and five hypertrophic ligament flavum tissue samples were collected.Abnormal methylation sites and methylation status were recorded by methylation sequencing and abnormally expressed genes were recorded by transcriptome integration analysis.The genes that showed a negative correlation between methylation level and expression level at the intersection of the two were selected.GO and KEGG enrichment analyses were used to study the major functional pathways and molecular functions of differentially expressed genes.Key genes regulating ligamentum flavum hypertrophy were screened using protein-protein interaction analysis.RESULTS AND CONCLUSION:Methylation sequencing of the two groups of the ligament flavum showed 37173 hypermethylation sites and 10583 low methylation sites.Transcriptome integration analysis found 720 abnormally expressed genes,of which 463 were upregulated and 257 were down-regulated.There were 383 overlapping genes,of which 192 genes showed a negative correlation between the methylation level and the expression level.GO functional pathway analysis results showed that molecular function was enriched to 10 terms,cellular component was enriched to 15 terms,and biological process(BP)was enriched to 30 terms.The results of KEGG pathway enrichment analysis showed that 192 genes were mainly enriched to 9 pathways,such as PI3K-Akt signaling pathway,Rap1 signaling pathway,and focal adhesion signaling pathway.The protein-protein interaction analysis identified five genes,PPARG,EGFR,CNR1,TNF and

关 键 词：黄韧带肥厚 甲基化测序 转录组整合分析 腰椎管狭窄症 DNA甲基化 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学] R274