Anderson D Greg, Li Xudong, Balian Gary
Department of Orthopaedic Surgery, University of Virginia, School of Medicine, Charlottesville, VA 22908, USA.
Spine (Phila Pa 1976). 2005 Jun 1;30(11):1242-6. doi: 10.1097/01.brs.0000164097.47091.4c.
A biochemical and gene expression study was conducted to determine the effects of the 30-kDa N-terminal fibronectin fragment (Fn-f) on the glycosaminoglycan content of nucleus pulposus (NP) explant cultures, and on the gene expression profile of NP cells in alginate culture.
To determine the effects of Fn-f on NP cells in alginate culture and disc explant cultures.
The macroscopic and histologic features of disc degeneration have been well described, but the molecular biology of disc degeneration remains poorly understood. Although fibronectin and fibronectin fragments are known to accumulate in degenerative discs, the role of fibronectin fragments on the degenerative process has not been elucidated. This study sought to define the effects of Fn-f on the expression of key matrix and degradative genes and on disc matrix proteins.
New Zealand white rabbits discs were harvested. NP cells were either isolated and grown in alginate culture or cultured as explanted tissue. The cultured cells were exposed to 10 nmol/L, 100 nmol/L, and 1 micromol/L concentrations of 30-kDa N-terminal Fn-f or a control substance and then analyzed histologically, biochemically, and with gene expression studies.
Alginate-cultured NP cells maintained a histologic appearance and phenotypic expression pattern similar to disc cells in vivo. Exposure of these cells to Fn-f led to the up-regulation of the MMP-9, MMP-13, and Fas genes and the down-regulation of the Type II collagen and aggrecan genes. In explant culture, Fn-f exposure led to a 60% reduction in glycosaminoglycan content compared with controls.
Treatment of NP cells in vitro with Fn-f led to changes in matrix proteins and gene expression similar to those seen during disc degeneration in vivo. This supports a possible detrimental role of the N-terminal fibronectin fragment in degenerative disc disease.
进行了一项生化和基因表达研究，以确定30 kDa N端纤连蛋白片段（Fn-f）对髓核（NP）外植体培养物中糖胺聚糖含量以及对藻酸盐培养中NP细胞基因表达谱的影响。
确定Fn-f对藻酸盐培养和椎间盘外植体培养中NP细胞的影响。
椎间盘退变的宏观和组织学特征已得到充分描述，但椎间盘退变的分子生物学仍知之甚少。尽管已知纤连蛋白和纤连蛋白片段在退变椎间盘中积累，但纤连蛋白片段在退变过程中的作用尚未阐明。本研究旨在确定Fn-f对关键基质和降解基因表达以及椎间盘基质蛋白的影响。
采集新西兰白兔的椎间盘。NP细胞要么分离出来并在藻酸盐培养中生长，要么作为外植组织进行培养。将培养的细胞暴露于10 nmol/L、100 nmol/L和1 μmol/L浓度的30 kDa N端Fn-f或一种对照物质，然后进行组织学、生化分析以及基因表达研究。
藻酸盐培养的NP细胞保持了与体内椎间盘细胞相似的组织学外观和表型表达模式。将这些细胞暴露于Fn-f导致MMP-9、MMP-13和Fas基因上调，以及II型胶原蛋白和聚集蛋白聚糖基因下调。在外植体培养中，与对照组相比，暴露于Fn-f导致糖胺聚糖含量降低60%。
体外使用Fn-f处理NP细胞导致基质蛋白和基因表达发生变化，类似于体内椎间盘退变时所见。这支持了N端纤连蛋白片段在椎间盘退变疾病中可能具有有害作用的观点。
