背景:很明显,长链非编码RNA(lncRNA)与牙周炎的发病机理有关。然而,详细的功能机制仍然未知。
目的:本研究旨在通过研究lncRNAs对蛋白质编码基因表达的调控来阐明其在牙周炎中的致病机制。
方法:用5μg/mL脂多糖(LPS)刺激人牙龈成纤维细胞-1(HGF-1)24小时,构建牙周炎细胞模型。进行qRTPCR和蛋白质印迹分析以确定由LPS诱导或参与炎症反应的基因的mRNA和蛋白质水平。使用ELISA测定细胞因子水平和炎性蛋白。进行转录组测序和分析以揭示lncRNA的表达特征。DESeq2(v1.4.5)用于分析差异表达基因。使用Phyper进行基因功能富集。AnimalTFDBv3.0用于分析参与牙周炎发病的转录因子。根据Pfam蛋白家族数据库鉴定了目标蛋白的Prot\ein结构域和家族。
结果:在LPS处理的HGF-1细胞中,我们检测到TNF-α和IL-1β的分泌,随着MDA和ROS的产生,表明LPS显著触发了HGF-1细胞的炎症反应和氧化应激。在对照组(ConT)和LPS处理组中检测到总共15,295个lncRNA。我们选择了10个显著差异共表达的lncRNA编码基因(MIR222HG,SNHG15,SNHG12,URS00005F6AA3,URS00009C153E,URS0000D57D7F,URS00019A4688,URS00019AF240,URS00019C6526和URS0001A00B79)作为诊断牙周炎进展的潜在生物标志物。由2个lncRNA编码基因(MIR222HG和SNHG15)和蛋白质编码基因(CBX5,NUPR1,CHAC1和MAB21L3)组成的相互作用网络可能与牙周炎的发病机理有关。ceRNA网络分析显示差异表达的lncRNAs参与炎症反应,免疫浸润,胶原纤维合成,脂多糖诱导牙周炎的骨改建。
结论:本研究确定了与牙周炎发病机制有关的关键分子,包括那些参与炎症调节的,胶原纤维合成,和骨骼重塑。我们的发现可能有助于解释lncRNAs如何参与牙周炎的病理过程。
BACKGROUND: It is evident that long non-coding RNAs (
lncRNAs) are implicated in the pathogenesis of periodontitis. However, the detailed functional mechanisms remain unknown.
OBJECTIVE: This study aimed to elucidate the pathogenic mechanisms of
lncRNAs in periodontitis by investigating their regulation of protein-coding gene expression.
METHODS: Human Gingival Fibroblasts-1 (HGF-1) were stimulated with 5 μg/mL of Lipopolysaccharide (LPS) for 24 hours to construct the periodontitis cell model. qRTPCR and western blot analyses were carried out to determine mRNA and protein levels of genes induced by LPS or involved in the inflammatory response. Cytokine levels and inflammatory proteins were assayed using ELISA. Transcriptome sequencing and analysis were conducted to reveal the expression signatures of
lncRNAs. DESeq2 (v1.4.5) was used to analyze differentially expressed genes. Gene function enrichment was carried out using Phyper. AnimalTFDB v3.0 was used to analyze transcription factors involved in the pathogenesis of periodontitis. Prot\\ein domains and families of the target proteins were identified based on the Pfam protein family database.
RESULTS: In LPS-treated HGF-1 cells, we detected the secretion of TNF-α and IL-1β, along with the production of MDA and ROS, indicating that LPS significantly triggered inflammatory responses and oxidative stress in HGF-1 cells. A total of 15,295
lncRNAs were detected in both the control (ConT) and LPS-treated groups. We selected 10 significantly differentially co-expressed lncRNA-coding genes (MIR222HG, SNHG15, SNHG12, URS00005F6AA3, URS00009C153E, URS0000D57D7F, URS00019A4688, URS00019AF240, URS00019C6526, and URS0001A00B79) as potential biomarkers for diagnosing the progression of periodontitis. An interaction network consisting of 2 lncRNA- encoding genes (MIR222HG and SNHG15) and protein-encoding genes (CBX5, NUPR1, CHAC1, and MAB21L3) may be involved in the pathogenesis of periodontitis. The ceRNA network analysis revealed the differentially expressed
lncRNAs to be involved in inflammatory response, immune infiltration, collagen fiber synthesis, and bone remodeling in LPS-induced periodontitis.
CONCLUSIONS: This study has identified pivotal molecules implicated in the pathogenesis of periodontitis, including those involved in inflammation regulation, collagen fiber synthesis, and bone remodeling. Our findings may contribute to explaining how lncRNAs participate in the pathological process of periodontitis.