Department of Anesthesiology, The Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, 710004, China.
Xi'an Jiaotong University Health Science Center, Xi'an, Shaanxi, 710061, China.
Chem Biol Interact. 2018 Dec 25;296:89-97. doi: 10.1016/j.cbi.2018.09.010. Epub 2018 Sep 19.
Accumulating evidence has suggested that microRNAs are critical regulators of intervertebral disc degeneration (IDD). The excessive apoptosis and extracellular matrix degradation of nucleus pulposus (NP) cells contribute to the initiation of IDD. However, the precise regulatory role of miRNAs in NP cell apoptosis and extracellular matrix degradation remains largely unknown. MicroRNA-30d (miR-30d) has been reported to be involved in regulating apoptosis and bone homeostasis. In this study, we aimed to investigate the role of miR-30d in regulating apoptosis and the extracellular matrix degradation of NP cells, along with the potential underlying molecular mechanism. Herein, our results showed that miR-30d was significantly increased in degenerative NP tissues compared with normal controls. Functional experiments showed that the inhibition of miR-30d promoted the viability and reduced the apoptosis of NP cells in vitro. Moreover, miR-30d inhibition increased the expression of type II collagen and aggrecan and inhibited the expression of matrix metalloproteinase. In contrast, the overexpression of miR-30d showed the opposite effects. Bioinformatics analysis, the dual-luciferase reporter assay, real-time quantitative PCR and western blot analysis showed that miR-30d directly targeted the 3'-untranslated region of SRY-related high mobility group box 9 (SOX9) and negatively regulated SOX9 expression. Correlation analysis showed that miR-30d expression was inversely correlated with SOX9 expression in degenerative NP tissues. Moreover, siRNA-mediated silencing of SOX9 expression significantly blocked the protective effects of miR-30d inhibition against NP cell apoptosis and extracellular matrix degradation. Overall, these results demonstrate that the inhibition of miR-30d attenuates the apoptosis and extracellular matrix degradation of degenerative human NP cells by up-regulating SOX9, suggesting a potential therapeutic target for IDD.
越来越多的证据表明 microRNAs 是椎间盘退变 (IDD) 的关键调节因子。核髓核 (NP) 细胞的过度凋亡和细胞外基质降解导致 IDD 的发生。然而，miRNAs 在 NP 细胞凋亡和细胞外基质降解中的精确调节作用在很大程度上仍不清楚。microRNA-30d (miR-30d) 已被报道参与调节细胞凋亡和骨稳态。在本研究中，我们旨在研究 miR-30d 在调节 NP 细胞凋亡和细胞外基质降解中的作用及其潜在的分子机制。研究结果表明，与正常对照组相比，退变的 NP 组织中 miR-30d 显著增加。功能实验表明，miR-30d 的抑制促进了 NP 细胞的活力并减少了其凋亡。此外，miR-30d 抑制增加了 II 型胶原和聚集蛋白聚糖的表达并抑制了基质金属蛋白酶的表达。相反，miR-30d 的过表达则表现出相反的效果。生物信息学分析、双荧光素酶报告基因检测、实时定量 PCR 和 Western blot 分析表明，miR-30d 可直接靶向性别决定区 Y 相关高迁移率族框 9 (SOX9) 的 3'-非翻译区并负调控 SOX9 表达。相关性分析表明，退变的 NP 组织中 miR-30d 表达与 SOX9 表达呈负相关。此外，siRNA 介导的 SOX9 表达沉默显著阻断了 miR-30d 抑制对 NP 细胞凋亡和细胞外基质降解的保护作用。综上所述，这些结果表明，抑制 miR-30d 通过上调 SOX9 减轻退变的人 NP 细胞的凋亡和细胞外基质降解，提示其可能成为 IDD 的潜在治疗靶点。
