MiR-103-3p regulates chondrocyte autophagy, apoptosis, and ECM degradation through the PI3K/Akt/mTOR pathway by targeting CPEB3.

IF 2.8 3区 医学 Q1 ORTHOPEDICS
Jun Li, Farui Sun, Yuanjin Zhang, Xian Pan, Bo Li, Guofu Zhang, Qian Zhou
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引用次数: 0

Abstract

Background: Chondrocyte apoptosis is associated with the severity of cartilage destruction and matrix degeneration in the progression of osteoarthritis. Increasing evidence indicates that autophagy has a significant cytoprotective effect against chondrocyte apoptosis. Here, we investigated the role of microRNA-103-3p (miR-103-3p) in regulating chondrocyte function and elucidated the underlying mechanism.

Methods: MiR-103-3p expression in interleukin-1β (IL-1β)-stimulated chondrocytes was evaluated using RT-qPCR. The targets of miR-103-3p predicted by online databases were verified using biotin-based pulldown assay and luciferase reporter assay. IL-1β stimulated-chondrocytes were transfected with miR-103-3p inhibitor along with siRNA targeting cytoplasmic polyadenylation element-binding protein3 (siCPEB3), the autophagy inhibitor 3-MA, or the PI3K agonist 740 Y-P. Chondrocyte proliferation was evaluated using cell counting kit-8. Apoptosis was detected by flow cytometry. The levels of apoptosis-, extracellular matrix (ECM)-, autophagy-, and the PI3K/Akt/mTOR pathway-related proteins in chondrocytes were detected using immunoblotting or immunofluorescence.

Results: We found that IL-1β stimulation upregulated miR-103-3p and downregulated CPEB3 in mouse chondrocytes. Inhibiting miR-103-3p reduced IL-1β-induced apoptosis and ECM macromolecule degradation while enhancing autophagy in chondrocytes. MiR-103-3p targeted CPEB3, and its downregulation rescued the expression of level in IL-1β stimulated-chondrocytes. MiR-103-3p downregulation inhibited the PI3K/Akt/mTOR pathway in IL-1β stimulated-chondrocytes by upregulating CPEB3. 3-MA, 740 Y-P, or CPEB3 knockdown counteracted the effect of miR-103-3p downregulation on chondrocyte apoptosis, ECM macromolecule degradation, and autophagy.

Conclusion: Overall, inhibition of miR-103-3p reduces IL-1β-induced apoptosis and ECM macromolecule degradation in chondrocytes by enhancing autophagy through the CPEB3/PI3K/Akt/mTOR pathway.

MiR-103-3p通过PI3K/Akt/mTOR通路靶向CPEB3调控软骨细胞自噬、凋亡和ECM降解。
背景:在骨关节炎的进展过程中,软骨细胞凋亡与软骨破坏和基质退行性变的严重程度有关。越来越多的证据表明,自噬对软骨细胞凋亡具有显著的细胞保护作用。在这里,我们研究了microRNA-103-3p (miR-103-3p)在调节软骨细胞功能中的作用,并阐明了其潜在的机制。方法:采用RT-qPCR方法检测IL-1β刺激软骨细胞中MiR-103-3p的表达。通过基于生物素的拉下实验和荧光素酶报告基因实验验证在线数据库预测的miR-103-3p靶点。IL-1β刺激的软骨细胞转染miR-103-3p抑制剂以及靶向细胞质多腺苷化元件结合蛋白3 (siCPEB3)的siRNA、自噬抑制剂3-MA或PI3K激动剂740 Y-P。使用细胞计数试剂盒-8评估软骨细胞增殖。流式细胞术检测细胞凋亡。采用免疫印迹法和免疫荧光法检测软骨细胞中凋亡-、细胞外基质(ECM)-、自噬-和PI3K/Akt/mTOR通路相关蛋白的水平。结果:我们发现IL-1β刺激上调小鼠软骨细胞中miR-103-3p并下调CPEB3。抑制miR-103-3p可减少il -1β诱导的细胞凋亡和ECM大分子降解,同时增强软骨细胞的自噬。MiR-103-3p靶向CPEB3,其下调挽救了IL-1β刺激的软骨细胞中的表达水平。MiR-103-3p下调通过上调CPEB3抑制IL-1β刺激的软骨细胞中的PI3K/Akt/mTOR通路。3-MA、740 Y-P或CPEB3敲低可抵消miR-103-3p下调对软骨细胞凋亡、ECM大分子降解和自噬的影响。结论:总体而言,抑制miR-103-3p可通过CPEB3/PI3K/Akt/mTOR途径增强自噬,从而减少il -1β诱导的软骨细胞凋亡和ECM大分子降解。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
CiteScore
4.10
自引率
7.70%
发文量
494
审稿时长
>12 weeks
期刊介绍: Journal of Orthopaedic Surgery and Research is an open access journal that encompasses all aspects of clinical and basic research studies related to musculoskeletal issues. Orthopaedic research is conducted at clinical and basic science levels. With the advancement of new technologies and the increasing expectation and demand from doctors and patients, we are witnessing an enormous growth in clinical orthopaedic research, particularly in the fields of traumatology, spinal surgery, joint replacement, sports medicine, musculoskeletal tumour management, hand microsurgery, foot and ankle surgery, paediatric orthopaedic, and orthopaedic rehabilitation. The involvement of basic science ranges from molecular, cellular, structural and functional perspectives to tissue engineering, gait analysis, automation and robotic surgery. Implant and biomaterial designs are new disciplines that complement clinical applications. JOSR encourages the publication of multidisciplinary research with collaboration amongst clinicians and scientists from different disciplines, which will be the trend in the coming decades.
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