Hypertrophy of ligamentum flavum (HLF) is a common lumbar degeneration disease (LDD) with typical symptoms of low back pain and limb numbness owing to an abnormal pressure on spinal nerves. Previous studies revealed HLF might be caused by fibrosis, inflammatory, and other bio-pathways. However, a global analysis of HLF is needed severely. A genome-wide DNA methylation and single-nucleotide polymorphism analysis were performed from five LDD patients with HLF and five LDD patients without HLF. Comprehensive integrated analysis was performed using bioinformatics analysis and the validated experiments including Sanger sequencing, methylation-specific PCR, qPCR and ROC analysis. Furthermore, the function of novel genes in ligamentum flavum cells (LFCs) was detected to explore the molecular mechanism in HLF through knock down experiment, overexpression experiment, CCK8 assay, apoptosis assay, and so on. We identified 69 SNP genes and 735 661 differentially methylated sites that were enriched in extracellular matrix, inflammatory, and cell proliferation. A comprehensive analysis demonstrated key genes in regulating the development of HLF including ACSM5. Furthermore, the hypermethylation of ACSM5 that was mediated by DNMT1 led to downregulation of ACSM5 expression, promoted the proliferation and fibrosis, and inhibited the apoptosis of LFCs. This study revealed that DNMT1/ACSM5 signaling could enhance HLF properties in vitro as a potential therapeutic strategy for HLF.

中文翻译：

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全基因组 DNA 甲基化和单核苷酸多态性的综合分析确定 ACSM5 是腰椎黄韧带肥大的抑制因子


黄韧带肥大症（HLF）是一种常见的腰椎退行性疾病（LDD），其典型症状是由于脊神经受到异常压力而导致腰痛和四肢麻木。先前的研究表明 HLF 可能是由纤维化、炎症和其他生物途径引起的。然而，迫切需要对 HLF 进行全球分析。对五名患有 HLF 的 LDD 患者和五名不患有 HLF 的 LDD 患者进行了全基因组 DNA 甲基化和单核苷酸多态性分析。使用生物信息学分析和经过验证的实验（包括桑格测序、甲基化特异性 PCR、qPCR 和 ROC 分析）进行全面的集成分析。进一步通过敲低实验、过表达实验、CCK8实验、凋亡实验等检测黄韧带细胞（LFCs）中新基因的功能，探讨HLF发生的分子机制。我们鉴定了 69 个 SNP 基因和 735 661 个差异甲基化位点，这些位点富含细胞外基质、炎症和细胞增殖。综合分析显示了调节 HLF 发生的关键基因，包括 ACSM5。此外，DNMT1介导的ACSM5高甲基化导致ACSM5表达下调，促进LFC的增殖和纤维化，并抑制细胞凋亡。这项研究表明，DNMT1/ACSM5 信号传导可以在体外增强 HLF 特性，作为 HLF 的潜在治疗策略。