医学分子生物学杂志 ›› 2025, Vol. 22 ›› Issue (1): 1-7.doi: 10.3870/j.issn.1672-8009.2025.01.001

• 论著 •    下一篇

P300 通过 Nrf2 / HO-1 / NF-κB 信号通路抑制脊柱侧凸大鼠的椎间盘退变 #br#

  

  1. 新疆医科大学第七附属医院1运动损伤康复科,2预防保健科 乌鲁木齐市, 830000
  • 出版日期:2025-01-31 发布日期:2025-02-28
  • 基金资助:
    新疆维吾尔自治区自然科学基金 (No. 2022D01C416)

P300 Inhibits Intervertebral Disc Degeneration through Nrf2 / HO-1 / NF-κB Signaling in Rat Scoliosis Model #br#

  1. 1Department of Sports Injury Rehabilitation,2 Department of Prevention and Health Care, the Seventh Affiliated Hospital of Xinjiang Medical University, Urumqi, 830000, China
  • Online:2025-01-31 Published:2025-02-28

摘要: 目的 探讨组蛋白乙酰化转移酶 P300 对脊柱侧凸大鼠的椎间盘髓核细胞 (nucleus pulposus cells,NPCs) 退变的影响和潜在调控机制 方法培养椎间盘 NPCs, NPCs 分为 4 : 无处理组 (对照组)、10 μg / L 白细胞介素 1β (interleukin-1β, IL-1β) 诱导 NPCs 退变组 (IL-1β )、 10 μg / L IL-1β 联合 15 mg /L P300 处理组 (IL-1β + P300 ) 15 mg / L P300 单独处理组 ( P300 )。 CCK-8 法检测细胞增殖活力酶联免疫吸附法 (enzyme linked immunosorbent assay, ELISA) 检测肿瘤坏死因子 α ( TNF-α) 和白细胞介素 6 (IL-6) 的水平流式细胞术检测细胞凋亡蛋白质印迹法检测细胞中性别决定区 Y 框蛋白 9 (sex determining region Y-box 9, SOX9)、 胶原蛋白Ⅱ ( collagen type Ⅱ , COL-Ⅱ )、 基质金属蛋白酶 13 ( matrixmetalloproteinase-13, MMP-13)、 金属蛋白酶 ADAMTS (A disintegrin and metalloproteinase with thrombospondinmotifs) -5、 核因子 κB-α 抑制蛋白 ( IκBα)、 磷酸化的 IκBα ( p-IκBα)、 磷酸化的 NF-κB P65 ( p-P65) 以及核转录因子红系 2 相关因子 2 ( nuclear factor erythroid 2-related factor 2, Nrf2) 和血红素加氧酶-1 ( hemeoxygenase-1, HO-1) 的表达另外, 40 只成年 SPF 级雌性 SD 大鼠分为假手术组脊柱侧凸组脊柱侧凸 + P300 脊柱侧凸 + P300 + Nrf2-IN-3 其中脊柱侧凸组用手术去除大鼠的双上肢及尾部脊柱侧凸+ P300 组建模后静脉注射 P300 [15 mg / (kg· d), 30 d]。 脊柱侧凸 + P300 + Nrf2-IN-3 组建模后静脉注射P300 [15 mg / (kg· d), 30 d] Nrf2 的抑制剂 Nrf2-IN-3 [23. 5 mg / (kg· d), 30 d]。 30 d 后取 T12 ~ L1段椎间盘髓核组织, 用蛋白质印迹法检测组织中 SOX9、 COL-Ⅱ 、 MMP-13、 ADAMTS-5 的表达 结果  与对照组比较, IL-1β 组的细胞活力降低, 但细胞凋亡增加, TNF-α、 IL-6、 MMP-13、 ADAMTS-5、 p-P65、 pIκBα 的表达水平上调, SOX9、 COL-Ⅱ 、 IκBα、 Nrf2、 HO-1 的表达水平下调 ( P< 0. 05)。 而与 IL-1β 组比较, IL-1β + P300 组的细胞活力增加, 细胞凋亡减少, TNF-α、 IL-6、 MMP-13、 ADAMTS-5、 p-P65、 pIκBα 的表达水平下调, SOX9、 COL-Ⅱ 、 IκBα、 Nrf2、 HO-1 的表达水平上调 ( P< 0. 05)。 与假手术组比较, 脊柱侧凸组的 SOX9 COL-Ⅱ 表达水平减少, MMP-13 ADAMTS-5 的表达增加 ( P< 0. 05), 与脊柱侧凸组比较, 脊柱侧凸 + P300 组的 SOX9 COL-Ⅱ 表达水平增加, MMP-13 ADAMTS-5 的表达减少 (P< 0. 05)。 与脊柱侧凸 + P300 组比较, 脊柱侧凸 + P300 + Nrf2-IN-3 组中的 SOX9 COL-Ⅱ 表达水平减少, MMP-13 ADAMTS-5 的表达增加 (P< 0. 05)。 结论 P300 通过调控 Nrf2 / HO-1 / NF-κB 信号通路抑制脊柱侧凸大鼠的椎间盘退变

关键词: P300, Nrf2 / HO-1 / NF-κB 信号通路, 脊柱侧凸, 大鼠, 椎间盘退变

Abstract: Objective To investigate the effect and potential regulatory mechanism of histoneacetyltransferase P300 on the degeneration of nucleus pulposus cells (NPCs) in the intervertebral discs in the rat scoliosis model. Methods NPCs were cultured and divided into 4 groups: the untreated control group, the interleukin-1β ( IL-1β) -induced degeneration group ( IL-1β group), the IL-1β combined with P300 treatment group (IL-1β + P300 group), and the P300 treatment alone group ( P300 group). Cell proliferation activity was detected using the Cell Counting Kit-8 (CCK-8) assay. The levels of TNF-α and IL-6 were measured using enzyme-linked immunosorbent assay (ELISA). Cell apoptosis was determined using flow cytometry. The expression levels of sex determining region Y-box 9 ( SOX9 )、 collagen type Ⅱ ( COL-Ⅱ ), matrix metalloproteinase-13 (MMP-13), A disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS-5), inhibitor of nuclear factor-kappa B-α ( IκBα), phosphorylated IκBα ( p-IκBα), phosphorylated NF-κB P65 (p-P65), nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) in the cells were analyzed using Western blotting. Additionally, 40 adult specific pathogen-free (SPF) female SD rats were divided into 4 groups: sham surgery group, scoliosis model group, scoliosis model + P300 group, and scoliosis model + P300 + Nrf2-IN-3 group. The scoliosis model was established by removing the upper limbs and tail of the rats through surgery. In the scoliosis model + P300 group, P300 was intravenously injected [15 mg / (kg· d), 30 d] after modeling. In the scoliosis model + P300 + Nrf2-IN-3 group, P300 [15 mg / ( kg· d), 30 d] and the Nrf2 inhibitor Nrf2-IN-3 [23. 5 mg / ( kg · d), 30 d] were intravenously injected after modeling. After 30 days, the nucleus pulposus tissues from the T12-L1 intervertebral discs were collected, and the expression levels of SOX9, COL-Ⅱ , MMP-13, and ADAMTS-5 were determined usingWestern blotting. Results Compared with the control group, the IL-1β group showed decreasedcell viability, increased cell apoptosis, upregulated expression levels of TNF-α, IL-6, MMP-13, ADAMTS-5, p-P65, and p-IκBα, and downregulated expression levels of SOX9, COL-Ⅱ ,IκBα, Nrf2, and HO-1 (all P< 0. 05). Compared with the IL-1β group, the IL-1β + P300 groupshowed increased cell viability, decreased cell apoptosis, decreased expression levels of TNF-α, IL-6, MMP-13, ADAMTS-5, p-P65, and p-IκBα, and increased expression levels of SOX9,COL-Ⅱ , IκBα, Nrf2, and HO-1 ( all P < 0. 05). Compared with the sham surgery group, thescoliosis model group showed decreased expression levels of SOX9 and COL-Ⅱ , and increased expression levels of MMP-13 and ADAMTS-5 ( all P < 0. 05 ). Compared with the scoliosis modelgroup, the scoliosis model + P300 group showed increased expression levels of SOX9 and COL-Ⅱ ,and decreased expression levels of MMP-13 and ADAMTS-5 ( all P < 0. 05). Compared with thescoliosis model + P300 group, the scoliosis model + P300 + Nrf2-IN-3 group showed decreased expression levels of SOX9 and COL-Ⅱ , and increased expression levels of MMP-13 and ADAMTS-5(all P< 0. 05). Conclusion P300 inhibits intervertebral disc degeneration by regulating the Nrf2 /HO-1 / NF-κB signaling pathway in the rat scoliosis model.

Key words:

P300, Nrf2 / HO-1 / NF-κB signaling pathway, scoliosis, rats, intervertebral disc degeneration

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