Intervertebral disc degeneration (IVDD) is the major cause of low-back pain. Histone deacetylase 9 (HDAC9) was dramatically decreased in the degenerative nucleus pulposus (NP) samples of patients with intervertebral disc degeneration (IVDD) according to bioinformatics analysis of Gene Expression Omnibus (GEO) GSE56081 dataset. This study aims to investigate the role of HDAC9 in IVDD progression. The contribution of HDAC9 to the progression of IVDD was assessed using HDAC9 knockout (HDAC9KO) mice and NP-targeted HDAC9-overexpressing mice by IVD injection of adenovirus-mediated HDAC9 under a Col2a1 promoter. Magnetic resonance imaging (MRI) and histological analysis were used to examine the degeneration of IVD. NP cells were isolated from mice to investigate the effects of HDAC9 on apoptosis and viability. mRNA-seq and coimmunoprecipitation/mass spectrometry (co-IP/MS) analysis were used to analyze the HDAC9-regulated factors in the primary cultured NP cells. HDAC9 was statistically decreased in the NP tissues in aged mice. HDAC9KO mice spontaneously developed age-related IVDD compared with wild-type (HDAC9WT) mice. In addition, overexpression of HDAC9 in NP cells alleviated IVDD symptoms in a surgically-induced IVDD mouse model. In an in vitro assay, knockdown of HDAC9 inhibited cell viability and promoted cell apoptosis of NP cells, and HDAC9 overexpression had the opposite effects in NP cells isolated from HDAC9KO mice. Results of mRNA-seq and co-IP/MS analysis revealed the possible proteins and signaling pathways regulated by HDAC9 in NP cells. RUNX family transcription factor 3 (RUNX3) was screened out for further study, and RUNX3 was found to be deacetylated and stabilized by HDAC9. Knockdown of RUNX3 restored the effects of HDAC9 silencing on NP cells by inhibiting apoptosis and increasing viability. Our results suggest that HDAC9 plays an important role in the development and progression of IVDD. It might be required to protect NP cells against the loss of cell viability and apoptosis by inhibiting RUNX3 acetylation and expression during IVDD. Together, our findings suggest that HDAC9 may be a potential therapeutic target in IVDD.

中文翻译：

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HDAC9在椎间盘退变中的分子机制和治疗潜力

椎间盘退变（IVDD）是腰痛的主要原因。根据基因表达综合 (GEO) GSE56081 数据集的生物信息学分析，椎间盘退变 (IVDD) 患者的退变髓核 (NP) 样本中组蛋白脱乙酰酶 9 (HDAC9) 显着降低。本研究旨在探讨 HDAC9 在 IVDD 进展中的作用。使用 HDAC9 敲除 (HDAC9KO) 小鼠和 NP 靶向 HDAC9 过表达小鼠，通过 IVD 注射 Col2a1 启动子下的腺病毒介导的 HDAC9 来评估 HDAC9 对 IVDD 进展的贡献。磁共振成像（MRI）和组织学分析用于检查IVD的变性。从小鼠体内分离 NP 细胞，以研究 HDAC9 对细胞凋亡和活力的影响。使用mRNA-seq和免疫共沉淀/质谱(co-IP/MS)分析来分析原代培养的NP细胞中的HDAC9调节因子。老年小鼠 NP 组织中的 HDAC9 显着减少。与野生型 (HDAC9WT) 小鼠相比，HDAC9KO 小鼠自发产生与年龄相关的 IVDD。此外，在手术诱导的 IVDD 小鼠模型中，NP 细胞中 HDAC9 的过度表达减轻了 IVDD 症状。在体外测定中，HDAC9的敲低抑制了NP细胞的细胞活力并促进了细胞凋亡，而HDAC9的过表达在从HDAC9KO小鼠分离的NP细胞中具有相反的作用。mRNA-seq 和 co-IP/MS 分析的结果揭示了 NP 细胞中 HDAC9 可能调控的蛋白质和信号通路。筛选出RUNX家族转录因子3（RUNX3）进行进一步研究，发现RUNX3被HDAC9去乙酰化并稳定。RUNX3 的敲低通过抑制细胞凋亡和增加活力来恢复 HDAC9 沉默对 NP 细胞的影响。我们的结果表明 HDAC9 在 IVDD 的发生和进展中发挥着重要作用。可能需要通过在 IVDD 期间抑制 RUNX3 乙酰化和表达来保护 NP 细胞免遭细胞活力丧失和凋亡。总之，我们的研究结果表明 HDAC9 可能是 IVDD 的潜在治疗靶点。