1 Department of Biological Sciences, College of Natural Sciences, Wonkwang University, Iksan, Chunbuk, Korea.
These authors contributed equally to this work.
Cartilage. 2019 Apr;10(2):148-156. doi: 10.1177/1947603517725566. Epub 2017 Aug 13.
In this article, we explored the hypothesis that the long noncoding RNA, Nespas, promotes osteoarthritis (OA) by supporting abnormal lipid metabolism in human chondrocytes.
Human articular chondrocytes from osteoarthritis patients were used and expression level of Nespas were determined by real-time polymerase chain reaction. Introduction of Nespas and Nespas-associated genes/miRNAs were performed by using a lentiviral system. The effect of Nespas on mitochondrial function was determined by staining mitochondria and analyzing mitopotential and mitochondrial genes. Moreover, to identify the responsible molecules in Nespas-induced pathogenesis, profiling of peroxisomal genes and miRNAs were applied and interactome analysis was performed.
Highly elevated levels of Nespas and Acyl-CoA synthetase 6 (ACSL6) were observed in OA patients. Both Nespas overexpression and ACSL6 upregulation into human chondrocytes induced typical OA characteristics, such as downregulation of type II collagen; upregulation of type I collagen, metalloproteinase 13, and caspase-1 and -3; and dysfunction of mitochondria and peroxisome. Co-expression of Nespas and ACSL6 siRNA reduced caspase-1 and -3 levels. Moreover, Nespas overexpression significantly suppressed levels of miR-291a-3p, -196a-5p, -23a-3p, -24-3p, and let-7a-5p, and these miRs are known to potentially target ACSL6 according to ingenuity pathway analysis. We also confirmed that these miRs were significantly suppressed in human OA chondrocytes. Overexpression of miR-291a-3p, -196a-5p, -23a-3p, -24-3p, or let-7a-5p in the presence of Nespas suppressed levels of ACSL6, caspase-1 and -3.
Overall, we suggest that elevated Nespas level in OA are associated with OA pathogenesis by suppressing miRs targeting ACSL6 and subsequent ACSL6 upregulation.
在本文中，我们探讨了长非编码 RNA Nespas 通过支持人软骨细胞异常脂质代谢促进骨关节炎（OA）的假说。
使用来自 OA 患者的人关节软骨细胞，通过实时聚合酶链反应确定 Nespas 的表达水平。通过慢病毒系统进行 Nespas 和 Nespas 相关基因/miRNA 的导入。通过染色线粒体和分析线粒体潜能和线粒体基因来确定 Nespas 对线粒体功能的影响。此外，为了鉴定 Nespas 诱导发病机制中的负责分子，应用过氧化物酶体基因和 miRNA 谱分析并进行互作分析。
在 OA 患者中观察到 Nespas 和酰基辅酶 A 合成酶 6（ACSL6）的水平高度升高。Nespas 的过表达和 ACSL6 的上调都诱导了人软骨细胞中的典型 OA 特征，例如 II 型胶原的下调；I 型胶原、金属蛋白酶 13 和半胱氨酸蛋白酶 1 和 -3 的上调；以及线粒体和过氧化物酶体的功能障碍。Nespas 和 ACSL6 siRNA 的共表达降低了半胱氨酸蛋白酶 1 和 -3 的水平。此外，Nespas 的过表达显著抑制了 miR-291a-3p、-196a-5p、-23a-3p、-24-3p 和 let-7a-5p 的水平，根据 ingenuity 通路分析，这些 miRs 已知可能靶向 ACSL6。我们还证实，这些 miR 在人 OA 软骨细胞中显著受到抑制。在 Nespas 存在下过表达 miR-291a-3p、-196a-5p、-23a-3p、-24-3p 或 let-7a-5p 可抑制 ACSL6、半胱氨酸蛋白酶 1 和 -3 的水平。
总的来说，我们认为 OA 中升高的 Nespas 水平与 OA 发病机制有关，通过抑制靶向 ACSL6 的 miR 并随后上调 ACSL6。
