hIGF-1基因转染促进退变椎间盘Ⅱ型胶原的表达  

Role of human insulin-like growth factor-1 gene on collagen type Ⅱ expression of degenerative intervertebral disk

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,郑州450052 [2]中山大学附属第二医院骨科,广州510120 [3]中山大学附属第一医院骨科,广州510089

出　　处：《中国矫形外科杂志》2008年第5期369-372,共4页Orthopedic Journal of China

基　　金：国家自然科学基金资助项目(NO.30471746);广东省科委攻关项目(NO.C30702)

摘　　要：[目的]探讨人胰岛素样生长因子（human insulin-like growth factor-1，hIGF-1）基因在退变椎间盘中的表达及对椎间盘中Ⅱ型胶原表达的影响。[方法]参考文献制备出腰椎间盘退变（intervertebral disk degeneration，IVDD）动物模型24只，随机分为携带hIGF-1基因重组腺病毒（Ad／CMV-hIGF-1）、hIGF-1生长因子及PBS组。L4，5、L5,6椎间盘中分别注射25μl第2代Ad／CMV-hIGF-1（8×10^8PFU）、hIGF-1生长因子（100μg／L）、PBS。注射后1、2、4、8周，Western blot检测hIGF-1蛋白表达；半定量RT-PCR检测Ⅱ型胶原mRNA的表达。[结果]hIGF-1蛋白带出现在7540．00U。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。Ⅱ型胶原电泳条带出现在200～300bp；在注射后1～4周，Ad／CMV-hIGF-1组内Ⅱ型胶原mRNA相对表达量进行性增加，8周轻度下降，4个时期比较（F=12．18，P〈0．001）；注射后1～8周，hIGF-1注射组、PBS注射组Ⅱ型胶原mRNA相对表达量进行性下降（F=27．38、21．56，P〈0．001）。相同时间点3组比较，Ⅱ型胶原mRNA相对表达量Ad／CMV-hIGF-1组最高，PBS组最低（F=27．31～39．85，P〈0．001）。[结论]hIGF-1基因在退变椎间盘中的表达能够促进合成Ⅱ型胶原。[ Objective] To study the role of hIGF-1 gene on collagen type Ⅱ expression of degenerative intervertebral disk. [ Method] Twenty- four male New-Zealand rabbits intervertebral disk degenerontion （IVDD） models were done according to reference and randomly divided into Ad-CMV-hIGF-1, hIGF-1 growth factor and PBS group. Twenty five microlitre the second generation 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 under fluoroscopic guidance. One, two, four and eight weeks post-operation, rabbits were sacririced, intervertebral disk samples were harvested. Total proteins of equal mass intervertebral disks were extracted, isolated in SDS-Polyacrylamide gel electrophoresis （SDS-PAGE） and transferred to polyvinylidene difluoride （PVDF） Millipore. The hIGF-1 growth factor expression were indentified with Western blot. Collagen type Ⅱ gene fragments were amplified with RTPCR, and relative expression was done with GAPDH as intern control. [ Result] The hIGF-1 interest protein existed at 7.6 Kilo-Dalton. At one week after injection, its expression quantities 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. At four week after injection, it still expressed in Ad/CMV-hIGF-1 group. At eight week after injection, it did not express in theree groups. Collagen type Ⅱ mRNA relative expressions increased significantly from one to four weeks after injection, declined slightly at the end of eight weeks in Ad/CMV-hIGF-1 group. However, they appeared to decrease continuously in the other two groups with time. At the corresponding phases, those in PBS group were the lowest. [Conclusion] Ad/CMV-hIGF-1 could successfully infect degenerative intervertebral disk. The hIGF-lgene expression could last four weeks and could stimulate collagen type Ⅱ synthesis in Ad/CMV-hIGF-1 group.

关 键 词：椎间盘 退变 表达 HIGF-1 

分 类 号：R681.5[医药卫生—骨科学]