Abstract:

Objective: To investigate the formation of stress granule (SG) in human periodontal ligament cells (hPDLCs) under stress, and its effect on the apoptosis, the expression of inflammatory factors, and the generation of reactive oxygen species (ROS) under lipopolysaccharide (LPS) stimulation.Methods: Sodium arsenite (SA) was used to induce SG formation in hPDLCs, and immunofluorescence staining was used to quantify SG formation. Live and dead cell staining, real-time quantitative polymerase chain reaction (RT-qPCR) and immunofluorescence staining were used to detect the localization and expression of cysteinyl aspartate-specific proteinase 3 (Caspase-3) in hPDLCs to evaluate the effect of SG formation on apoptosis. RT-qPCR was used to detect the mRNA expression changes of inflammatory-related cytokines interleukin 1β (IL-1β) and interleukin 6 (IL-6) during LPS stimulation. ROS detection was used to assess the effect of SG on LPS-induced ROS levels of hPDLCs.Results: SA successfully induced SG formation in hPDLCs. The results of immunofluorescence staining showed that Caspase-3 was localized in SG, which significantly inhibited the apoptosis of hPDLCs. RT-qPCR results indicated that SG could downregulate the mRNA relative expression of LPS-stimulated inflammatory cytokines in hPDLCs. ROS detection showed that SG inhibited LPS-induced ROS generation in hPDLCs.Conclusion: SG can protect hPDLCs from stress-induced apoptosis and inhibit the expression of LPS-triggered inflammatory-related factors and ROS generation.

Key words: stress granule, apoptosis, periodontitis, reactive oxygen species

摘要：

目的： 探讨应激状态下人牙周膜细胞（human periodontal ligament cells，hPDLCs）中应激颗粒（stress granule，SG）的形成及其对细胞凋亡、脂多糖（lipopolysaccharide，LPS）诱导下炎症相关因子表达和活性氧（reactive oxygen species，ROS）生成的影响。方法： 采用亚砷酸钠（sodium arsenite，SA）诱导hPDLCs形成SG，通过免疫荧光染色对SG进行量化分析；通过活死细胞染色、实时定量聚合酶链反应（real-time quantitative polymerase chain reaction，RT-qPCR）和免疫荧光检测hPDLCs中半胱氨酰天冬氨酸特异性蛋白酶3（cysteinyl aspartate-specific proteinase 3，Caspase-3）的定位及表达，评估SG对细胞凋亡的影响；采用RT-qPCR检测LPS刺激下炎症因子白细胞介素1β（interleukin 1β，IL-1β）和白细胞介素6（interleukin 6，IL-6）的mRNA表达变化；应用ROS检测试剂盒检测SG对LPS诱导下hPDLCs中的ROS水平影响。结果：SA成功诱导hPDLCs形成SG；免疫荧光结果显示Caspase-3定位于SG，且SG的形成显著抑制了hPDLCs凋亡；RT-qPCR结果显示，SG可下调LPS刺激下hPDLCs相关炎症因子的mRNA相对表达；ROS检测结果显示SG能抑制LPS刺激下hPDLCs的ROS生成。结论：SG可保护hPDLCs免受应激诱导的细胞凋亡，并显著抑制LPS触发的炎症因子的表达及ROS生成。

关键词: 应激颗粒, 凋亡, 牙周炎, 活性氧

CLC Number: 

• R782