转染hIGF-1基因增强兔退变椎间盘蛋白多糖的表达  

Enhancement effect of human insulin-like growth factor-1 gene on aggrecan expression of degenerative intervertebral disk in rabbits

作　　者：黄宗强[1] 刘尚礼[2] 郑召民[3] 

机构地区：[1]郑州大学第一附属医院骨科,河南郑州450052 [2]中山大学第二附属医院骨科,广东广州510120 [3]中山大学第一附属医院骨科,广东广州510089

出　　处：《基础医学与临床》2008年第8期805-809,共5页Basic and Clinical Medicine

基　　金：国家自然科学基金(30471746);广东省科委攻关项目(C30702)

摘　　要：目的探讨人胰岛素样生长因子（hIGF—1）基因在退变椎间盘中的表达及对椎间盘中蛋白多糖（aggrecan）的影响。方法制备新西兰大白兔腰椎间盘退变（IDD）模型24只，随机分为Ad／CMV—hIGF—1、hIGF—1生长因子及PBS组，每组8只。L4—5、L5-6椎间盘中分别注射第2代Ad／CMV—hIGF—1（8×10^8PFU）、hIGF—1生长因子（100μg／，L）、PBS均25μL。注射后1、2、4和8周，Western blot检测hIGF—1蛋白表达；RT—PCR检测aggrecan mRNA的表达。结果hIGF-1蛋白带出现在7．6×10^3ku。Ad／CMV-hIGF-1组hIGF-1蛋白表达持续达4周以上，hIGF-1组表达持续约2周；PBS注射组无hIGF—1蛋白表达。aggrecan电泳条带出现在200—300bp；在注射后1—4周，Ad／CMV—hIGF—1组内aggrecan mRNA相对表达量进行性增加，8周轻度下降，4个时期总的比较（F=8．51，P〈0．05），注射后1—8周，hIGF—1组、PBS组aggrecan mRNA相对表达量进行性下降。结论hIGF—1能够增强椎间盘aggrecan的表达。Objective To study hIGF—1 gene expression and the influence on aggrecan expression of degenerative intervertebral disk. Methods Twenty four male New-Zealand rabbits IDD models were established and randomly divided into Ad/CMV-hIGF—1 ,hIGF-1 growth factor and PBS group. Twenty five mL of Ad/CMV-hIGF—1 （T = 80 × 10^9 PFU/L）, hIGF—1 growth factor（ 100 μg/L） and PBS were respectively injected into L4-5, L5-6 intervertebral disk through fluoroscopic guidance. One, two, four and eight weeks after-operation, rabbits were sacrificed, intervertebral disk samples were collected. Total protein of equal mass intervertebral disks were extracted, isolated by SDS-PAGE and transferred to polyvinylidene difluoride （PVDF） Millipore. The hIGF-1 growth factor expression were indentified with Western blot. Aggrecan gene fragments were amplified with RT-PCR, and relative expressionwas checked with GAPDH as intern control. Results The hIGF-1 interest protein existed at 7. 6 kilo-Dalton. One week after injection, its expression quantifies were almost equal between Ad/CMV-hIGF—1 and hIGF-1 growth factor group. At two week after injection, it obviously declined in hIGF-1 growth factor group. Four week after injection, it was still expressed in Ad/CMV-hIGF—1 group. Eight week after injection, it did not express in three groups. Aggrecan gene relative expression increased significantly from one to four weeks after injection, declined slightly by the end of eight weeks in Ad/CMV-hIGF—1 group. However, it appeared to decrease continuously in the other two groups with time. Conclusion Ad/CMV-hIGF—1 successfully infected degenerative intervertebral disk. The hIGF-1 gene expression lasted four weeks and could stimulate aggrecan synthesis in Ad/CMV-hIGF—1 group.

关 键 词：椎间盘 退变 人胰岛素样生长因子 

分 类 号：Q786[生物学—分子生物学] R681.53[医药卫生—骨科学]