Wang X F, Zhang A P, Sun Z Y, Liu C, Kuang L H, Tian J W
Department of Orthopedic Surgey, Hospital of Zaozhuang Mining Corporation, Zaozhuang 277101, China.
Zhonghua Yi Xue Za Zhi. 2017 May 9;97(17):1324-1329. doi: 10.3760/cma.j.issn.0376-2491.2017.17.011.
To investigate the changes in the expression of NF-κB signaling pathway in human degenerative intervertebral discs. From October 2014 to March 2016, 55 nucleus pulposus of surgical patients with degenerative human intervertebral disc were collected for study in Department of Orthopedic Surgey, Hospital of Zaozhuang Mining Corporation, and Department of Orthopedics, Shanghai General Hospital Affiliated Shanghai Jiaotong University, School of Medicine.The collected nucleus pulposus tissues were divided into two groups: experimental group(30) and control group(25). Cell culture observed normal and degenerative nucleus pulposus cells morphological changes; immunofluorescence observed NF-κB p65 changes in the nucleus of nucleus pulposus cells.Real-time PCR was observed changes in aggregated proteoglycans and matrix metalloproteinase gene mRNA.Finally, the use of blockers of nucleus pulposus cells were treated 24 hours, Western blot analysis the changing of p65, ADAMTS-4, MMP-13, aggregate proteoglycans and collagen Ⅱ protein expression. Compared with the experimental group, the nucleus pulposus cells in the control group had larger cell volume, abundant cytoplasm and faster growth rate.Cell immunofluorescence show Nondegenerative nucleus pulposus cells p65 protein was mainly localized in the cytoplasm, degeneration of nucleus pulposus cells p65 protein was mainly concentrated in the nucleus.RT-PCR showed degenerative group of matrix metalloproteinases (MMP-1, MMP-3, MMP-13), aggrecanase(ADAMTS-4, ADAMTS-5) and IL-6 mRNA expression was significantly higher than Nondegenerative group; aggrecan and type Ⅱ collagen expression than those without degeneration group was significantly lower.Expression of nucleus pulposus degeneration in nuclear protein p65 with the degenerative level increased gradually increased.BAY11-7082 blocked the activity of NF-κB signaling pathway, which could significantly down-regulate the expression of ADAMTS-4 and MMP-13 protein and significantly up-regulate the expression of Agg and COLⅡ protein.With the increase of BAY11-7082 concentration, gradually strengthened. The activation of the NF-κB signaling pathway in a degenerative intervertebral disc is gradually increased, regulating the over-expression of matrix-degrading enzymes.It plays an important role in the degradation of extra-cellular matrix.
探讨人退变椎间盘核因子-κB（NF-κB）信号通路表达的变化。2014年10月至2016年3月，收集枣庄矿业集团医院骨科及上海交通大学医学院附属上海第一人民医院骨科手术患者的55份退变人椎间盘髓核用于研究。将收集的髓核组织分为两组：实验组（30份）和对照组（25份）。细胞培养观察正常和退变髓核细胞的形态变化；免疫荧光观察髓核细胞核内NF-κB p65的变化。实时荧光定量聚合酶链反应（Real-time PCR）观察聚集蛋白聚糖和基质金属蛋白酶基因mRNA的变化。最后，用NF-κB信号通路阻滞剂处理髓核细胞24小时，蛋白质免疫印迹法分析p65、含血小板反应蛋白基序的解聚蛋白样金属蛋白酶-4（ADAMTS-4）、基质金属蛋白酶-13（MMP-13）、聚集蛋白聚糖和Ⅱ型胶原蛋白表达的变化。与实验组比较，对照组髓核细胞体积较大，胞质丰富，生长速度较快。细胞免疫荧光显示非退变髓核细胞p65蛋白主要定位于细胞质，退变髓核细胞p65蛋白主要集中于细胞核。RT-PCR显示退变组基质金属蛋白酶（MMP-1、MMP-3、MMP-13）、聚集蛋白聚糖酶（ADAMTS-4、ADAMTS-5）及白细胞介素-6（IL-6）mRNA表达明显高于非退变组；聚集蛋白聚糖和Ⅱ型胶原表达明显低于非退变组。髓核退变中核蛋白p65表达随退变程度增加而逐渐升高。BAY11-7082阻断NF-κB信号通路活性，可显著下调ADAMTS-4和MMP-13蛋白表达，显著上调聚集蛋白聚糖（Agg）和Ⅱ型胶原（COLⅡ）蛋白表达。且随BAY11-7082浓度增加，作用逐渐增强。退变椎间盘中NF-κB信号通路激活逐渐增加，调控基质降解酶的过度表达，在细胞外基质降解中起重要作用。
