肿瘤坏死因子-α作用下椎间盘退变动物模型的建立

Establishment of animal model of intervertebral disc degeneration induced by TNF-α

目的:制作家兔椎间盘退变(IVDD)的动物模型,研究肿瘤坏死因子-α(TNF-α)对髓核组织的影响。方法:取健康成年家兔30只,手术暴露L2~L6,L3/4、L4/5作为实验组,每个椎间盘髓核注射20 ng/μL TNF-α15μL;L5/6作为对照组,每个椎间盘髓核注射15μL磷酸缓冲盐溶液;L2/3作为空白组,不做处理。于术后4周、8周、12周行腰椎X线及磁共振(MRI)检查,每次随机检查10只家兔,检查完后处死并取其髓核组织经行免疫组织化学及原位末端标记检查。结果:术后4~12周,X线见实验组椎间隙高度进行性下降,MRI可见实验组椎间隙T2加权像信号逐渐降低;免疫组织化学及TUNEL显示实验组髓核组织内Ⅰ型胶原及细胞凋亡率随之时间推移逐渐增加,Ⅱ型胶原逐渐减少。对照组和空白组差异无统计学意义(P>0.05)。结论:实验成功建立了TNF-α作用下的IVDD动物模型。TNF-α介导的椎间盘术后椎间隙高度丢失,同时TNF-α可诱导椎间盘髓核细胞退变及凋亡。微量注射器的穿刺作用对髓核细胞的退变及凋亡无明显影响。

Objective ：To establish the animal model of intervertebral disc degeneration （IVDD） induced by tumor necrosis iactor-α （TNF-α） , and investigate the influence of TNF-α on nucleus pulposus tissue. Methods：The nucleus pulposus of intervertebral disc in L2 to L6, ,L3/4 and L4/5 in 30 New Zealand white rabbits were injected with 15 μL of 20 ng/μL TNF-α, and set as the experimental group. The nucleus pulposus of intervertebral disc in L5/6, was injected with 15 μL of PBS,and set as the control group. The non-treat-ment L2/3 of rabbit was set as the blank group. Ten rabbits were randomly selected,and detected using X-ray and MRI aher 4,8 and 12 weeks of operation,respectively. The nucleus pulposus tissue of rabbits were examined using immunohistochemistry and TUNEL. Re-suits ：After 4 to 12 weeks of operation, the height and T2 weighted image signal of intervertebral disc in experimental group gradually decreased under X-ray and MRI, respectively. The results of immunohistochemistry and TUNEL showed that the expression level of type Ⅰ collagen and cell apoptosis in experimental group gradually increased with the time extending,the expression level of type Ⅱ collagen gradually decreased, and the differences of which between the control group and blank group was not statistically significant（ P 〉 0.05 ）. Conclusions ：The IVDD animal model induced by TNF-α is successthlly established. The height of intervertebral disc loses under the mediating of TNF-v, and TNF-α can induce the degeneration and apoptosis of nucleus pulposus cells. The microsyringe puncture can not affect the degeneration and apoptosis of nucleus pulposus cells.