兔腰椎间盘退变动物模型中锌指蛋白A20和NF-κB的表达及其作用研究

发布时间：2018-05-20 15:38

  本文选题：椎间盘退变 + 锌指蛋白A20　； 参考：《重庆医科大学》2017年硕士论文

【摘要】：目的:检测锌指蛋白A20、NF-κB在兔正常及退变腰椎间盘髓核组织中的表达,并在体外培养原代正常及退变的兔腰椎间盘髓核细胞,并进一步用内毒素(LPS)刺激髓核细胞,观察锌指蛋白A20、NF-κB及相关炎性因子的表达及其相互关系。方法:构建兔腰椎间盘退变动物模型,获取兔腰椎间盘退变模型中正常及退变的椎间盘组织,HE染色观察椎间盘髓核及纤维环的形态变化,免疫组化检测椎间盘髓核细胞锌指蛋白A20、NF-κB p65蛋白及Ⅱ型胶原(COL-Ⅱ)的表达。体外分离、培养获取的正常和退变兔腰椎间盘髓核细胞,分为正常组、退变组、内毒素(LPS)刺激组和NF-κB抑制剂组,Real-time PCR分别检测各组A20、IL-1β、TNF-α、NF-κB、Ⅱ型胶原mRNA的表达变化。Western blot观察A20、p65、Ⅱ型胶原蛋白的表达变化。ELISA检测细胞培养液中IL-1β、TNF-α的表达。结果:组织学观察发现退变组髓核细胞数量明显减少,髓核细胞聚集成团,Ⅱ型胶原表达下降,A20、p65蛋白表达升高;细胞实验发现,与正常组相比,退变组A20、TNF-α、IL-1β、p65表达在mRNA及蛋白水平均明显升高,Ⅱ型胶原表达降低(p0.05);在LPS刺激组中A20、TNF-α、IL-1β、p65表达较退变组更为显著,Ⅱ型胶原降低明显(p0.05),而NF-κB抑制剂组A20、TNF-α、IL-1β、p65表达下降,Ⅱ型胶原表达回升,且与LPS刺激组有显著统计学意义(p0.05)。结论:兔腰椎间盘退变时,锌指蛋白A20表达增加,NF-κB活性增强;当加入LPS刺激后,锌指蛋白A20、及其相关炎性因子表达进一步升高,且锌指蛋白A20与NF-κB呈正相关,提示髓核细胞炎症反应与兔椎间盘退变的发生、发展密切相关,其中A20可能发挥了重要作用。
[Abstract]:Objective: to detect the expression of zinc finger protein A20 (NF- 魏 B) in normal and degenerative nucleus pulposus of lumbar intervertebral disc in rabbits, and to culture primary normal and degenerative nucleus pulposus cells of rabbit lumbar intervertebral disc in vitro, and to further stimulate nucleus pulposus cells with endotoxin (LPS). To observe the expression and correlation of zinc finger protein A 20, NF- 魏 B and related inflammatory factors. Methods: a rabbit model of lumbar disc degeneration was established. The normal and degenerated intervertebral disc tissues were stained with HE to observe the morphological changes of the nucleus pulposus and the fibrous ring of the intervertebral disc. The expression of zinc finger protein A20, NF- 魏 B p65 and collagen 鈪，

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