Li Feng, Sun Jianguang, Huang Shilei, Su Gao, Pi Guofu
Cell Physiol Biochem. 2018;45(3):1241-1251. doi: 10.1159/000487455. Epub 2018 Feb 9.
BACKGROUND/AIMS: Osteoarthritis (OA) is the most frequently occurring joint disease and characterized by degeneration of cartilage. As the unique cell type in cartilage, chondrocytes play a crucial role during OA. Our study explored the influence of long non-coding RNA (lncRNA) growth arrest-specific transcript 5 (GAS5) on lipopolysaccharides (LPS)-induced injury in ATDC5 cells.
Cell viability, apoptosis and expression of inflammatory cytokines were all assessed to evaluate LPS-induce inflammatory injury. Expression of GAS5 in LPS-induced cells was evaluated by qRT-PCR. After cell transfection, effect of abnormally expressed GAS5 on LPS-induced inflammatory injury was determined. Then, the possible target of GAS5 was screened by bioinformatics and verified by qRT-PCR and luciferase activity assay. Together, whether aberrant expression of target gene affected the modulation of GAS5 in LPS-induced inflammatory injury was also assessed. Finally, the influences of aberrant expressed Kruppel-like factor 2 (KLF2) on nuclear factor κB (NF-κB) and Notch pathways were detected by Western blot analysis.
LPS reduced cell viability and promoted cell apoptosis and secretion of inflammatory cytokines, along with down-regulation of GAS5. LPS-induced injury was alleviated by GAS5 overexpression while was exacerbated by GAS5 silence. KLF2 was predicted and verified as a target of GAS5, and GAS5 functioned through regulating expression of KLF2. Besides, aberrant expression of KLF2 regulated expressions of key kinases involved in the NF-κB and Notch pathways.
GAS5 might ameliorate LPS-induced inflammatory injury in ATDC5 chondrocytes by inhibiting the NF-κB and Notch signaling pathways.
背景/目的：骨关节炎（OA）是最常见的关节疾病，其特征为软骨退变。软骨细胞作为软骨中独特的细胞类型，在骨关节炎发生过程中起关键作用。本研究探讨长链非编码RNA（lncRNA）生长停滞特异性转录本5（GAS5）对脂多糖（LPS）诱导的ATDC5细胞损伤的影响。
通过评估细胞活力、凋亡及炎性细胞因子表达来评价LPS诱导的炎性损伤。采用qRT-PCR检测LPS诱导细胞中GAS5的表达。细胞转染后，确定异常表达的GAS5对LPS诱导炎性损伤的影响。然后，通过生物信息学筛选GAS5可能的靶标，并经qRT-PCR和荧光素酶活性测定进行验证。同时，评估靶基因的异常表达是否影响GAS5对LPS诱导炎性损伤的调控作用。最后，通过蛋白质印迹分析检测异常表达的 Kruppel样因子2（KLF2）对核因子κB（NF-κB）和Notch信号通路的影响。
LPS降低细胞活力，促进细胞凋亡及炎性细胞因子分泌，同时下调GAS5表达。GAS5过表达减轻LPS诱导的损伤，而GAS5沉默则加剧该损伤。KLF2被预测并验证为GAS5的靶标，GAS5通过调节KLF2表达发挥作用。此外KLF2的异常表达调节NF-κB和Notch信号通路中关键激酶的表达。
GAS5可能通过抑制NF-κB和Notch信号通路减轻LPS诱导的ATDC5软骨细胞炎性损伤。
