NEAT1/miR-193a-3p/SOX5 axis regulates cartilage matrix degradation in human osteoarthritis
NEAT1/miR-193a-3p/SOX5 axis regulates cartilage matrix degradation in human osteoarthritis
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DOI：
10.1002/cbin.11291
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发表时间：
2020-04-01
影响因子：
3.9
通讯作者：
Wu, Fei
中科院分区：
文献类型：
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作者：
Liu, Feng;Liu, Xiangyang;Wu, Fei
据报道，长链非编码RNA（lncRNA）参与骨关节炎（OA）的进展。本研究的目的是探讨lncRNA核富集丰富转录本1（NEAT 1）在OA中的功能作用。采用逆转录定量聚合酶链反应（RT-qPCR）分析OA软骨组织和软骨细胞中microRNA（miR-193a）-3p、NEAT 1和性别决定区Y盒蛋白5（SOX 5）的表达，以及促炎细胞因子白细胞介素-6（IL-6）、IL-1 β、肿瘤坏死因子-α（TNF-α）和IL-8的水平。流式细胞仪检测软骨细胞凋亡情况。使用蛋白质印迹分析测定细胞外基质ACAN、II型胶原α 1链（Col2a1）、基质金属蛋白酶-3（MMP-3）、MMP-13、去整合素和具有血小板反应蛋白基序的金属蛋白酶（ADAMTS）-5和SOX 5的蛋白水平。进行双荧光素酶报告基因测定以确定NEAT 1、miR-193a-3p和SOX 5之间的靶标关系。我们发现miR-193a-3p在OA软骨组织和软骨细胞中表达下调，而NEAT 1和SOX 5表达上调。miR-193a-3p的上调和NEAT 1的敲低都抑制了炎症、凋亡，并降低了MMP-3、MMP-13和ADAMTS-5的蛋白水平，同时提高了软骨细胞中ACAN和Col2a1的表达。NEAT 1靶向miR-193a-3p，而SOX 5被miR-193a-3p靶向。miR-193 a-3 p的沉默逆转了NEAT 1敲低介导的对炎症、细胞凋亡和细胞外基质产生的影响。SOX 5的引入消除了miR-193a-3p上调对软骨细胞中炎症、凋亡和细胞外基质产生的影响。总之，NEAT1/miR-193a-3p/SOX 5轴调节人OA中的软骨基质降解。
Long non-coding RNAs (lncRNAs) were reported to be involved in the progression of osteoarthritis (OA). The aim of this work was to explore the functional role of lncRNA nuclear-enriched abundant transcript 1 (NEAT1) in OA. Reverse-transcription quantitative polymerase chain reaction (RT-qPCR) was employed to analyze the expression of microRNA (miR-193a)-3p, NEAT1, and sex-determining region Y-box protein 5 (SOX5), as well as the levels of pro-inflammatory cytokines interleukin-6 (IL-6), IL-1 beta, tumor necrosis factor-alpha (TNF-alpha), and IL-8 in OA cartilage tissue and chondrocytes. In addition, flow cytometry was used to measure the apoptosis of chondrocytes. The protein levels of extracellular matrix ACAN, collagen type II alpha 1 chain (Col2a1), matrix metalloproteinase-3 (MMP-3), MMP-13, a disintegrin, and metalloproteinase with thrombospondin motifs (ADAMTS)-5 and SOX5 were determined using western blot analysis. Dual-luciferase reporter assay was performed to determine the target relationship among NEAT1, miR-193a-3p, and SOX5. We found that miR-193a-3p expression was downregulated, while NEAT1 and SOX5 were upregulated in OA cartilage tissue and chondrocytes. Both upregulation of miR-193a-3p and knockdown of NEAT1 suppressed inflammation, apoptosis, and reduced the protein levels of MMP-3, MMP-13, and ADAMTS-5, while elevating ACAN and Col2a1 expression in chondrocytes. NEAT1 targeted miR-193a-3p, and SOX5 was targeted by miR-193a-3p. Silencing of miR-193a-3p reversed the NEAT1 knockdown-mediated effect on the inflammation, apoptosis, and production of the extracellular matrix. The introduction of SOX5 abolished the impact of the upregulation of miR-193a-3p on inflammation, apoptosis, and production of extracellular matrix in chondrocytes. In conclusion, NEAT1/miR-193a-3p/SOX5 axis regulates cartilage matrix degradation in human OA.