Department of Orthopaedics, Peking University Third Hospital, Beijing, China.
J Cell Mol Med. 2020 Aug;24(15):8753-8762. doi: 10.1111/jcmm.15509. Epub 2020 Jun 24.
Thoracic ossification of the ligamentum flavum (TOLF) causes serious spinal canal stenosis. The underlying aetiology may relate to genetic and inflammatory factors. DNA methylation plays a critical role in osteogenesis and inflammation, whereas there is no genome-wide DNA methylation analysis about TOLF. The two subtypes of TOLF (single-level and multiple-level) have distinct clinical features. Using micro-computed tomography (micro-CT), we showed the ossification arose from the joint between two vertebrae at one/both sides of ligament flavum. With Illumina Infinium Human Methylation 850 BeadChip arrays, genome-wide DNA methylation profile was measured in ligament flavum of eight healthy and eight TOLF samples. Only 65 of the differentially methylated cytosine-phosphate-guanine dinucleotides were found in both subtype groups. Principal component analysis and heat map analysis showed a different methylation pattern in TOLF samples, and methylation patterns of two subtypes are also distinct. The Gene Ontology enrichment analysis was significantly enriched in differentiation and inflammation. Pyrosequencing analysis and quantitative real-time polymerase chain reaction were performed to validate the arrays results and expression levels, to test six differentially methylated genes (SLC7A11, HOXA10, HOXA11AS, TNIK, homeobox transcript antisense RNA, IFITM1), using another independent samples (P < 0.05). Our findings first demonstrated an altered Genome-wide DNA methylation profile in TOLF, and implied distinct methylated features in two subtypes.
黄韧带骨化（TOLF）可导致严重的椎管狭窄。其潜在病因可能与遗传和炎症因素有关。DNA 甲基化在成骨和炎症中起着关键作用，而关于 TOLF 的全基因组 DNA 甲基化分析尚未见报道。TOLF 有两种亚型（单节段和多节段），具有不同的临床特征。我们使用微计算机断层扫描（micro-CT）显示骨化起源于黄韧带两侧的两个椎体之间的关节。使用 Illumina Infinium Human Methylation 850 BeadChip 阵列，我们测量了 8 个健康和 8 个 TOLF 样本中黄韧带的全基因组 DNA 甲基化谱。仅在两个亚型组中发现了 65 个差异甲基化的胞嘧啶-磷酸-鸟嘌呤二核苷酸。主成分分析和热图分析显示 TOLF 样本的甲基化模式不同，两种亚型的甲基化模式也不同。基因本体论富集分析在分化和炎症中显著富集。焦磷酸测序分析和定量实时聚合酶链反应用于验证阵列结果和表达水平，以测试六个差异甲基化基因（SLC7A11、HOXA10、HOXA11AS、TNIK、同源框转录反义 RNA、IFITM1），使用另一个独立样本（P < 0.05）。我们的研究结果首次证明了 TOLF 中全基因组 DNA 甲基化谱的改变，并暗示了两种亚型中存在不同的甲基化特征。
