■骨关节炎(OA)是一种以关节软骨退化为特征的慢性炎症性退行性疾病。环状RNA已被证明在OA过程中起重要作用。在这里,这项工作旨在探讨circSCAPER在OA进展中的潜在作用和机制。
■circscaper的水平,通过qRT-PCR或Western印迹检测miR-127-5p和toll样受体4(TLR4)。通过流式细胞术确定细胞凋亡。使用蛋白质印迹检查Aggrecan和基质金属肽酶的表达以评估细胞外基质(ECM)降解。通过测量炎症因子的释放来确定炎症反应和氧化应激,随着细胞内活性氧和丙二醛的产生。miR-127-5p与circscaper或TLR4之间的相互作用通过双荧光素酶报告基因测定,RNA免疫沉淀和下拉测定。
■用白细胞介素-1β(IL-1β)处理软骨细胞以在体外模拟OA条件。OA软骨和IL-1β诱导的软骨细胞中CircSCAPER增加。功能上,circscaper的敲减减弱IL-1β诱发的凋亡,ECM降解,体外炎症和氧化应激。发现OA软骨中的circSCAPER上调伴随着miR-127-5p降低和TLR4增加。机械上,circscaper充当miR-127-5p的海绵,正向调节软骨细胞中TLR4的表达。IL-1β治疗降低miR-127-5p表达,但上调TLR4表达,miR-127-5p的再表达抑制IL-1β引起的软骨细胞损伤,TLR4过表达被废除。此外,miR-127-5p抑制逆转了circscaper敲低对IL-1β处理下软骨细胞的保护作用。
■CircSCAPER沉默保护IL-1β诱导的细胞凋亡,ECM降解,通过miR-127-5p/TLR4轴在软骨细胞中的炎症和氧化应激。
UNASSIGNED: Osteoarthritis (OA) is a chronic inflammatory degenerative disease characterized by articular cartilage degradation. Circular RNAs have been shown to play significant roles in OA process. Herein, this work aimed to investigate the potential role and mechanism of circSCAPER in OA progression.
UNASSIGNED: Levels of circSCAPER, miR-127-5p and toll-like receptor 4 (TLR4) were detected by qRT-PCR or western blotting. Cell apoptosis was determined by flow cytometry. The expression of Aggrecan and Matrix metallopeptidase was examined using western blot to assess extracellular matrix (ECM) degradation. Inflammatory response and oxidative stress were determined by measuring the release of inflammatory factors, along with the generation of intracellular reactive oxygen species and malondialdehyde. The interaction between miR-127-5p and circSCAPER or TLR4 was determined by dual-luciferase reporter, RNA immunoprecipitation and pull-down assays.
UNASSIGNED: Chondrocytes were treated with interleukin-1β (IL-1β) to mimic OA condition in vitro. CircSCAPER was increased in OA cartilages and IL-1β-induced chondrocytes. Functionally, knockdown of circSCAPER attenuated IL-1β-evoked apoptosis, ECM degradation, inflammation and oxidative stress in vitro. CircSCAPER up-regulation in OA cartilages was discovered to be accompanied by decreased miR-127-5p and increased TLR4. Mechanistically, circSCAPER acted as a sponge for miR-127-5p to positively regulate TLR4 expression in chondrocytes. IL-1β treatment reduced miR-127-5p expression but up-regulated TLR4 expression, re-expression of miR-127-5p suppressed IL-1β-caused chondrocyte injury, which was abolished by TLR4 overexpression. Moreover, miR-127-5p inhibition reversed the protective action of circSCAPER knockdown on chondrocytes under IL-1β treatment.
UNASSIGNED: CircSCAPER silencing protected against IL-1β-induced apoptosis, ECM degradation, inflammation and oxidative stress in chondrocytes via miR-127-5p/TLR4 axis.