Abstract Context: Osteoarthritis (OA) is a degenerative arthrosis sickness. Astragaloside IV (AS-IV) functions by relieving inflammatory damage. Objective: We aimed to investigate the mechanism by which AS-IV protects ATD cells from lipopolysaccharide (LPS)-induced damage. Materials and methods: ATDC5 cells were transfected with miR-203 inhibitor and NC inhibitor (150 nM) or pEX-MyD88 and sh-MyD88 (50 nM) for 48 h, pre-treated by 15 μg/mL AS-IV for 24 h, then treated by 5 μg/mL LPS for 12 h. Dual-luciferase activity testing was used to determine whether miR-203 could bind to MyD88. CCK-8 and flow cytometry were used to detect cell activity and apoptosis, respectively, and qRT-PCR, western blots, and ELISA were performed to detect expression levels of miR-203 and inflammatory cytokines. Results: Based on the 50% inhibiting concentration (IC50), there was no significant difference of AS-IV (0 to 15 μg/mL) on cell viability. Fifteen μg/mL was the optimal concentration of AS-IV in treating LPS-induced inflammatory damage in subsequent experiments since this was a semi-lethal concentration. AS-IV significantly reduces LPS-induced viability, apoptosis and the release of TNF-α, IL-6 and iNOS mainly through up-regulating miR-203. Further, MyD88 was a target gene of miR-203 and negatively regulated by miR-203. Knockdown of MyD88 inhibited LPS-induced inflammatory damage by inhibiting the NF-κB signal pathway. Discussion and conclusions: AS-IV protects ATDC5 cells against LPS-induced damage mainly via regulating miR-203/MyD88. Our results support a theoretical basis for in-depth study of the function of AS-IV and the clinical cure of OA.

中文翻译：

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黄芪甲苷通过调节 miR-203/MyD88 保护 ATDC5 细胞免受脂多糖引起的损伤

摘要背景：骨关节炎（OA）是一种退行性关节病。黄芪甲苷 (AS-IV) 通过缓解炎症损伤发挥作用。目的：我们旨在研究 AS-IV 保护 ATD 细胞免受脂多糖 (LPS) 诱导损伤的机制。材料和方法：ATDC5细胞用miR-203抑制剂和NC抑制剂（150 nM）或pEX-MyD88和sh-MyD88（50 nM）转染48小时，用15 μg/mL AS-IV预处理24小时，然后用 5 μg/mL LPS 处理 12 小时。双荧光素酶活性测试用于确定 miR-203 是否可以与 MyD88 结合。CCK-8和流式细胞术分别用于检测细胞活性和细胞凋亡，并进行qRT-PCR、蛋白质印迹和ELISA检测miR-203和炎性细胞因子的表达水平。结果：基于 50% 抑制浓度 (IC50)，AS-IV（0 至 15 μg/mL）对细胞活力没有显着差异。在随后的实验中，15 μg/mL 是 AS-IV 治疗 LPS 诱导的炎症损伤的最佳浓度，因为这是一个半致死浓度。AS-IV 主要通过上调 miR-203 显着降低 LPS 诱导的活力、细胞凋亡和 TNF-α、IL-6 和 iNOS 的释放。此外，MyD88 是 miR-203 的靶基因，受 miR-203 负调控。MyD88 的敲低通过抑制 NF-κB 信号通路来抑制 LPS 诱导的炎症损伤。讨论与结论：AS-IV 主要通过调节 miR-203/MyD88 保护 ATDC5 细胞免受 LPS 诱导的损伤。我们的研究结果为深入研究AS-IV的功能和OA的临床治愈提供了理论基础。