牙周炎(PD)是一种与牙周炎细菌相关的多因素炎性疾病。赖氨酸特异性脱甲基酶1(LSD1),一种组蛋白去甲基酶,通过与miRNA靶标结合,参与了口腔疾病中炎症反应过程的调节。这项研究调查了miRNA与LSD1结合的分子机制及其对成骨分化的后续作用。首先,分离人牙周膜干细胞(hPDLSCs),培养,和特点。然后对这些细胞进行脂多糖(LPS)处理以诱导炎症,之后开始成骨分化。采用qPCR和蛋白质印迹来监测LSD1表达的变化。随后,在hPDLSCs中沉默LSD1以评估其对成骨分化的影响。通过生物信息学和双荧光素酶报告基因测定,预测并确认miR-708-3p为LSD1的靶miRNA。随后,评估miR-708-3p表达,并通过过表达评估其在PDhPDLSCs中的作用。使用染色质免疫沉淀(ChIP)和蛋白质印迹测定,我们探索了miR-708-3p和LSD1通过二甲基化H3K4(H3K4me2)对osterix(OSX)转录的潜在调节。最后,我们研究了OSX在hPDLSCs中的作用。LPS处理hPDLSCs后,LSD1的表达增加,但是这种趋势在诱导成骨分化后被逆转。沉默LSD1加强了hPDLSCs的成骨分化。miR-708-3p被发现直接结合并负调节LSD1,导致通过H3K4me2的去甲基化抑制OSX转录。此外,miR-708-3p的过表达可促进hPDLSCs在炎症微环境中成骨分化。然而,OSX的表达降低部分减弱了保护作用。我们的发现表明miR-708-3p靶向调节LSD1以通过H3K4me2甲基化增强OSX转录,最终促进hPDLSCs成骨分化。
Periodontitis (PD) is a multifactorial inflammatory disease associated with periodontopathic bacteria. Lysine-specific demethylase 1 (
LSD1), a type of histone demethylase, has been implicated in the modulation of the inflammatory response process in oral diseases by binding to miRNA targets. This study investigates the molecular mechanisms by which miRNA binds to
LSD1 and its subsequent effect on osteogenic differentiation. First, human periodontal ligament stem cells (hPDLSCs) were isolated, cultured, and characterized. These cells were then subjected to lipopolysaccharide (LPS) treatment to induce inflammation, after which osteogenic differentiation was initiated. qPCR and western blot were employed to monitor changes in
LSD1 expression. Subsequently,
LSD1 was silenced in hPDLSCs to evaluate its impact on osteogenic differentiation. Through bioinformatics and dual luciferase reporter assay, miR-708-3p was predicted and confirmed as a target miRNA of LSD1. Subsequently, miR-708-3p expression was assessed, and its role in hPDLSCs in PD was evaluated through overexpression. Using chromatin immunoprecipitation (ChIP) and western blot assay, we explored the potential regulation of osterix (OSX) transcription by miR-708-3p and LSD1 via di-methylated H3K4 (H3K4me2). Finally, we investigated the role of OSX in hPDLSCs. Following LPS treatment of hPDLSCs, the expression of
LSD1 increased, but this trend was reversed upon the induction of osteogenic differentiation. Silencing LSD1 strengthened the osteogenic differentiation of hPDLSCs. miR-708-3p was found to directly bind to and negatively regulate LSD1, leading to the repression of OSX transcription through demethylation of H3K4me2. Moreover, overexpression of miR-708-3p was found to promote hPDLSCs osteogenic differentiation in inflammatory microenvironment. However, the protective effect was partially attenuated by reduced expression of OSX. Our findings indicate that miR-708-3p targetedly regulates LSD1 to enhance OSX transcription via H3K4me2 methylation, ultimately promoting hPDLSCs osteogenic differentiation.