椎间盘灌注培养条件下压应力频率对纤维环细胞基质合成的影响

Effects of loading frequency on matrix synthesis of annulus fibrosus cells in an intervertebral disc perfusion culture system

中文关键词:  椎间盘  退变  器官培养  压应力  频率

英文关键词:Intervertebral disc  Degeneration  Organ culture  Loading  Frequency

中文摘要:

  【摘要】 目的：观察压应力频率对椎间盘纤维环（AF）细胞基质合成的影响。方法：获取4～6月龄巴马香猪胸腰段椎间盘72个后，随机分为无应力对照组和应力组（分别为0.1Hz组、0.5Hz组、1Hz组、3Hz组、5Hz组），每组12个样本，将各组椎间盘置于生物反应器中进行灌注培养。无应力对照组不施加应力刺激，各应力组每天施加对应频率的应力刺激2h。灌注培养5d后分离全层纤维环组织行HE染色、阿利新蓝染色、胞外基质基因Ag?鄄grecan、Collagen Ⅰ、Collagen Ⅱ定量PCR、糖胺多糖（GAG）含量和羟脯氨酸（HYP）含量的检测。结果：培养5d后，各组纤维环组织仍保持天然的层状叠加结构。与对照组比较时，除5Hz组外其他应力组（0.1Hz、0.5Hz、1Hz、3Hz）纤维环组织阿利新蓝染色强度增强、各胞外基质基因的表达得到维持（0.1Hz）或上调（0.5Hz、1Hz、3Hz）、GAG和HYP的含量也维持在同等水平（0.1Hz）或显著升高（0.5Hz、1Hz、3Hz）。然而各应力组组间比较时，5Hz组纤维环组织阿利新蓝染色强度、胞外基质基因的表达水平及GAG和HYP的含量比其他应力组（0.1Hz、0.5Hz、1Hz、3Hz）都下调（P＜0.05）。结论：椎间盘体外灌注培养条件下，压应力刺激频率影响纤维环细胞的基质合成，低频率（0.1～3Hz）的应力刺激增加基质合成，高频率（5Hz）的应力刺激减少基质合成。

英文摘要:

  【Abstract】 Objectives: To investigate the effects of loading frequency on matrix synthesis of annulus fibrosus (AF) cells. Methods: Discs from porcine(4-6 months old) were isolated and randomly divided into control group (no loading) and loading groups(0.1Hz, 0.5Hz, 1Hz, 3Hz and 5Hz group, respectively). All discs were put in tissue culture chamber for perfusion culturing. No loading was applied to the discs in the control group while different loading frequencies(0.1Hz, 0.5Hz, 1 Hz, 3Hz and 5Hz, respectively) were applied to the discs in the loading groups. After culturing for 5 days, the full-thickness AF tissues were used for the analysis of HE staining, alcian blue staining, matrix genes(Aggrecan, Collagen Ⅰ, collagen Ⅱ) expression, GAG content and HYP content. Results: After 5 days, AF in each group remained the natural lamellar structure. Compared with the control group, stronger intensity of alcian blue staining, up-regulated expression of matrix genes and increased GAG content and HYP content were observed in loading groups except for the 5Hz group. When compared among loading groups, the alcian blue staining intensity, expression of matrix genes, GAG content and HYP content in 5Hz group all decreased compared with those of other loading groups(0.1Hz, 0.5Hz, 1Hz and 3Hz group). Conclusions: Under condition of intervertebral disc perfusion culturing, matrix synthesis of AF cells is influenced by loading frequency, which increases at low frequency(0.1-3Hz) and decreases at high frequency(5Hz).