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模拟骨关节炎：miR-16-5p 通过 MAPK 通路靶向 MAP2K1 减轻 IL-1β 诱导的软骨细胞功能障碍。

Modeling Osteoarthritis: MiR-16-5p Attenuates IL-1β Induced Chondrocyte Dysfunction by Targeting MAP2K1 through the MAPK Pathway.

机构信息

Department of Orthopedics, Wuhan Hospital of Traditional Chinese Medicine, Wuhan, Hubei, China.

Department of Massage, Wuhan Hospital of Traditional Chinese Medicine, Wuhan, Hubei, China.

出版信息

Ann Clin Lab Sci. 2023 Mar;53(2):248-258.

PMID:37094858

Abstract

OBJECTIVE

Osteoarthritis (OA) is a chronic joint disease characterized by cartilage degeneration, significantly reducing the quality of life. Previous report has confirmed that MAP2K1 acts as a potential therapeutic target in OA. Nevertheless, its specific function and related molecular mechanism in OA remain uncharacterized. Our report revealed the biological significance of MAP2K1 and elucidated its regulatory mechanism in OA.

METHODS

Interleukin (IL)-1β was utilized to stimulate human chondrocyte cell line CHON-001 for establishing the models of OA. Cell apoptosis and viability were determined by flow cytometry analysis and CCK-8 assay. Protein levels and gene expression were quantified by western blotting and RT-qPCR. Binding relation between miR-16-5p and MAP2K1 (mitogen-activated protein kinase kinase 1) was confirmed by luciferase reporter assay.

RESULTS

IL-1β treatment triggered CHON-001 cell injury by repressing cell viability and facilitating cell apoptosis. Moreover, IL-1β stimulation upregulated MAP2K1 level in CHON-001 cells. MAP2K1 depletion attenuated IL-1β-elicited CHON-001 cell injury. Mechanistically, miR-16-5p targeted MAP2K1 in CHON-001 cells. In rescue assays, MAP2K1 upregulation counteracted the suppressive impact of miR-16-5p enhancement on IL-1β-triggered CHON-001 cell dysfunction. In addition, upregulated miR-16-5p suppressed IL-1β-elicited activation of MAPK pathway in CHON-001 cells.

CONCLUSIONS

MiR-16-5p mitigates IL-1β-induced damage to chondrocyte CHON-001 by targeting MAP2K1 and inactivating the MAPK signaling.

摘要

目的

骨关节炎（OA）是一种慢性关节疾病，其特征为软骨退化，显著降低生活质量。先前的报告已经证实 MAP2K1 可作为 OA 的潜在治疗靶点。然而，其在 OA 中的具体功能和相关分子机制仍未被阐明。我们的报告揭示了 MAP2K1 的生物学意义，并阐明了其在 OA 中的调控机制。

方法

利用白细胞介素（IL）-1β刺激人软骨细胞系 CHON-001 建立 OA 模型。通过流式细胞术分析和 CCK-8 测定来检测细胞凋亡和活力。通过 Western blot 和 RT-qPCR 来定量蛋白质水平和基因表达。通过荧光素酶报告实验来验证 miR-16-5p 与 MAP2K1（丝裂原活化蛋白激酶激酶 1）之间的结合关系。

结果

IL-1β 处理通过抑制细胞活力和促进细胞凋亡来引发 CHON-001 细胞损伤。此外，IL-1β 刺激可上调 CHON-001 细胞中的 MAP2K1 水平。MAP2K1 耗竭可减轻 IL-1β 诱导的 CHON-001 细胞损伤。机制上，miR-16-5p 在 CHON-001 细胞中靶向 MAP2K1。在挽救实验中，MAP2K1 的过表达抵消了 miR-16-5p 增强对 IL-1β 触发的 CHON-001 细胞功能障碍的抑制作用。此外，上调的 miR-16-5p 抑制了 IL-1β 诱导的 CHON-001 细胞中 MAPK 通路的激活。