基因治疗家兔退变颈椎间盘的动物实验研究

Experimental study on the effect of gene therapy on IL-1β,TNF-α of degenerated cervical intervertebral discs,cartilage and plate calification in rabbits with cervical intervertebral disc degeneration

目的探讨基因治疗退变颈椎间盘对细胞因子含量及软骨终板钙化的影响。方法选用25只4月龄新西兰兔,建立家兔颈椎动力平衡失调模型,诱导颈椎间盘退变(正常对照组不作处理)。术后7个月,将带有转化生长因子β1(TGF-β1)基因的重组质粒DNA注入基因治疗组(浅层、全层)家兔C2～3、C3～4、C4～5椎间盘中(每个椎间盘用量为20μl)。1个月后用耳缘静脉气栓法处死动物,用双抗夹心ELISA法检测颈椎间盘中白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)抗体含量,并在形态学上评定颈椎间盘退变程度,测定各组动物颈椎软骨终板钙化层厚度。结果模型浅层组、模型全层组与正常对照组比较,颈椎间盘中IL-1β、TNF-α含量明显升高(P<0.01),颈椎软骨终板钙化层也明显增厚(P<0.01),基因治疗退变颈椎间盘对IL-1β、TNF-α含量及颈椎软骨终板钙化有明显抑制作用(P<0.05)。结论退变颈椎间盘组织释放多种细胞因子和炎症介质,它们可加速颈椎间盘退变;而软骨终板钙化是颈椎间盘退变的始动因素,二者呈高度正相关。基因治疗退变颈椎间盘对IL-1β、TNF-α含量及软骨终板钙化有明显的抑制作用。

Objective To observe the changes of cytokines in degenerated cervical intervertebral discs and the relationship between cartilage end-plate calcification and degeneration of cervical intervertebral discs in rabbits,and to investigate the effect of gene therapy on the contents of IL-1β,TNF-α in degenerated cervical intervertebral discs and on the thickness of the calcified layer of the cartilage end-plate of cervical spine.Methods 25 New Zealand rabbits（4-month-old） were enrolled in the study,whose weight was 2～3kg.The rabbits were randomly divided into five groups：normal group,model group 1（shallow layer）,model group 2（whole layer）,gene-treatment group 1（shallow layer）,gene-treatment group 2（whole layer）.The model of the cervical dynamic unbalance were established by surgical intervention except for the rabbits in normal group.After eight months,the contents of IL-1β,TNFαin cervical intervertebral discs of every group were detected by ELISA,and the thickness of the calcified layer of the cartilage end-plates in every group were measured.Results The contents of IL-1β,TNF-α in degenerated cervical intervertebral discs and the thickness of the calcified layer of the cartilage end-plate in gene-treatment groups were significantly decreased,as compared with those in model groups（P0.05）.Conclusion The gene therapy can decrease the contents of IL-1β,TNF-α in degenerated cervical intervertebral discs and the thickness of the calcified layer of the cartilage end-plate in cervical spine of rabbits.