Department of Orthopaedics and Traumatology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, 21 Sassoon Road, Hong Kong, China; HKU Shenzhen Institute of Research and Innovation, China; Center for Reproduction, Development and Growth, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China; Stem Cell & Regenerative Medicine Consortium, The University of Hong Kong, Hong Kong, China.
Department of Orthopaedics and Traumatology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, 21 Sassoon Road, Hong Kong, China.
Osteoarthritis Cartilage. 2016 Oct;24(10):1826-1836. doi: 10.1016/j.joca.2016.05.012. Epub 2016 May 19.
Recent evidence suggests a role of fibrogenesis in intervertebral disc (IVD) degeneration. We aim to explore if fibrotic genes may serve as IVD degeneration indicators, and if their expression is associated with myofibroblast activity.
Transcriptional expression of fibrosis markers (COL1A1, COL3A1, FN1, HSP47, MMP12, RASAL1) were analyzed in degenerated (D) and non-degenerated (ND) human nucleus pulposus (NP) and annulus fibrosus (AF) cells, along with traditional (SOX9, ACAN) and newly established degeneration markers (CDH2, KRT19, KRT18, FBLN1, MGP, and COMP). Protein expression was investigated by immunohistochemistry in human IVDs, and in rodent IVDs undergoing natural ageing or puncture-induced degeneration. Co-expression with myofibroblast markers was examined by double staining on human and rat specimens. Disc degeneration severity and extent of fibrosis were determined by histological scoring and picrosirius red staining respectively.
Human D-NP showed more intensive staining for picrosirius red than ND-NP. Among the genes examined, D-NP showed significantly higher MMP12 expression along with lower KRT19 expression. Protein expression analysis revealed increased MMP12(+) cells in human D-IVD. Histological scoring indicated mild degeneration in the punctured rat discs and discs of ageing mouse. Higher MMP12 positivity was found in peripheral NP and AF of the degenerative rat discs and in NP of the aged mice. In addition, human D-NP and D-AF showed increased α-SMA(+) cells, indicating enhanced myofibroblast activity. MMP12 was found co-expressed with α-SMA, FSP1 and FAP-α in human and rat degenerative IVDs.
Our study suggests that in addition to a reduced KRT19 expression, an increased expression of MMP12, a profibrotic mediator, is characteristic of disc degenerative changes. Co-expression study indicates an association of the increased MMP12 positivity with myofibroblast activity in degenerated IVDs. Overall, our findings implicate an impact of MMP12 in disc cell homeostasis. The precise role of MMP12 in IVD degeneration warrants further investigation.
最近的证据表明，纤维化在椎间盘（IVD）退变中起作用。我们旨在探讨纤维化基因是否可作为 IVD 退变的指标，以及它们的表达是否与肌成纤维细胞活性相关。
分析退变（D）和非退变（ND）人髓核（NP）和纤维环（AF）细胞中纤维化标记物（COL1A1、COL3A1、FN1、HSP47、MMP12、RASAL1）的转录表达，以及传统（SOX9、ACAN）和新建立的退变标记物（CDH2、KRT19、KRT18、FBLN1、MGP 和 COMP）。通过免疫组织化学染色法在人 IVD 以及经历自然老化或穿刺诱导退变的鼠 IVD 中检测蛋白表达。通过双染色法在人及大鼠标本上检测与肌成纤维细胞标记物的共表达。通过组织学评分和苦味酸天狼猩红染色分别确定椎间盘退变严重程度和纤维化程度。
人 D-NP 比 ND-NP 显示出更强的苦味酸天狼猩红染色。在所检查的基因中，D-NP 显示 MMP12 表达显著升高，同时 KRT19 表达降低。蛋白表达分析显示人 D-IVD 中 MMP12(+)细胞增多。组织学评分显示穿刺鼠椎间盘和老龄鼠椎间盘有轻度退变。退变鼠椎间盘的 NP 和 AF 及老龄鼠 NP 中 MMP12 阳性率较高。此外，人 D-NP 和 D-AF 显示α-SMA(+)细胞增多，表明肌成纤维细胞活性增强。人及鼠退变 IVD 中 MMP12 与α-SMA、FSP1 和 FAP-α共表达。
我们的研究表明，除了 KRT19 表达降低外，促纤维化介质 MMP12 的表达增加也是椎间盘退行性变化的特征。共表达研究表明，退变 IVD 中 MMP12 阳性率增加与肌成纤维细胞活性有关。总的来说，我们的研究结果表明 MMP12 对椎间盘细胞稳态有影响。MMP12 在 IVD 退变中的确切作用还需要进一步研究。
