Department of Orthopaedic Trauma and Microsurgery, Zhongnan Hospital of Wuhan University, Wuhan, Hubei Province, China.
Department of Orthopaedic, Huangshi No. 4 Hospital, Huangshi, Hubei Province, China.
J Gene Med. 2020 Nov;22(11):e3246. doi: 10.1002/jgm.3246. Epub 2020 Jul 14.
The present study aimed to investigate whether forkhead box M1 (FOXM1), as a putative target of miR-877-5p, participated in interleukin (IL)-1β-induced cartilage degeneration in experimental osteoarthritis (OA) models in vitro and in vivo.
In vitro and in vivo models of OA were established using IL-1β treated primary mouse chondrocytes and anterior cruciate ligament transection (ACLT) operation in mice. miR-877-5p mimics or agomir-miR-877-5p were used as therapeutic agents in both in vitro and in vivo models of OA. Cell viability and apoptosis were evaluated using cell counting kit-8 and terminal deoxynucleotidyl transferase dUTP nick end labeling staining, respectively. A quantitative reverse transcriptase-polymerase chain reaction and western blotting were used to measure gene and protein expression, respectively.
FOXM1 was up-regulated in IL-1β-stimulated chondrocytes and the proximal tibia of ACLT-operated mice. Bioinformatics algorithms deduced a highly conserved sequence in the 3'-UTR of FOXM1 that could be bound with miR-877-5p. A luciferase assay indicated that miR-877-5p directly targeted the 3'-UTR of FOXM1. Overexpression of miR-877-5p could reduce protein expression of FOXM1 in chondrocytes. Concurrently, IL-1β-evoked up-regulation of FOXM1 protein expression was neutralized in chondrocytes following transfection with miR-877-5p mimics. miR-877-5p mimics or agomir-miR-887-5p could inhibit IL-1β-induced inflammation in both in vitro and in vivo models of OA. miR-877-5p might have beneficial effects on the synthesis of cartilage matrix via the promotion of SRY-box transcription factor 9 and type II collagen expression and inhibition of matrix metalloproteinase 9 expression.
miR-877-5p can improve chondrocyte function in both in vivo and in vitro models of OA, based on post-transcriptional repression of FOXM1 as a postulated molecular mechanism.
本研究旨在探讨叉头框转录因子 M1（FOXM1）是否作为 miR-877-5p 的潜在靶点，参与体外和体内实验性骨关节炎（OA）模型中白细胞介素（IL）-1β诱导的软骨退变。
使用 IL-1β处理的原代小鼠软骨细胞和前交叉韧带横断（ACLT）手术建立 OA 的体内外模型。miR-877-5p 模拟物或 agomir-miR-877-5p 用于 OA 的体内外模型的治疗剂。使用细胞计数试剂盒-8 和末端脱氧核苷酸转移酶 dUTP 缺口末端标记染色分别评估细胞活力和细胞凋亡。定量逆转录-聚合酶链反应和 Western blot 分别用于测量基因和蛋白表达。
FOXM1 在 IL-1β刺激的软骨细胞和 ACLT 手术小鼠的胫骨近端均上调。生物信息学算法推断出 FOXM1 的 3'-UTR 中存在一个高度保守的序列，可以与 miR-877-5p 结合。荧光素酶报告基因实验表明，miR-877-5p 可直接靶向 FOXM1 的 3'-UTR。在软骨细胞中转染 miR-877-5p 模拟物可降低 FOXM1 蛋白表达。同时，在转染 miR-877-5p 模拟物后，IL-1β 诱导的 FOXM1 蛋白表达上调在软骨细胞中被中和。miR-877-5p 模拟物或 agomir-miR-877-5p 可抑制体内外 OA 模型中 IL-1β 诱导的炎症。miR-877-5p 可能通过促进性别决定区 Y 框转录因子 9 和 II 型胶原表达以及抑制基质金属蛋白酶 9 表达，对软骨基质的合成产生有益影响。
miR-877-5p 可能通过抑制 FOXM1 的转录后表达，作为一种潜在的分子机制，改善体内外 OA 模型中的软骨细胞功能。
