rAAV2-CTGF和TIMP1双基因联合转染恒河猴退变腰椎间盘的生物学效应  

Biological effects of CTGF combined with TIMP1 mediated by adeno associated virus to degenerated lumbar intervertebral discs in rhesus monkey

作　　者：孔杰 胡有谷[1] 刘勇[1] 齐宗华[1] 

机构地区：[1]青岛大学医学院附属医院脊柱外科,266003 [2]青岛市骨伤科医院

出　　处：《中华骨科杂志》2010年第3期287-292,共6页Chinese Journal of Orthopaedics

基　　金：基金项目：国家自然科学基金（30471750）

摘　　要：目的研究腺相关病毒（adeno—associated virus2，AAV2）介导的结缔组织生长冈子（connective tissue growth factor，CTGF）和基质金属蛋白酶组织抑制因子1（tissue inhibitor of metalloproteinasesl，TIMP1）双基因体内转染退变的恒河猴腰椎间盘的生物学效应。方法恒河猴9只，雌性4只，雄性5只；年龄4-7岁，体重4．5-7kg。CT引导下经皮穿刺纤维环诱导恒河猴腰椎间盘退变，微创经皮注入携带CTGF和TlMPI双基因腺相关病毒颗粒。应用Rrr_PCR测定细胞因子的表达，RT—PCR、35S整合法观察双基因联合转染对退变椎间盘的生物学作用。结果双基因转染后8、16、24周时CTGFmRNA表达量分别为PBS对照组的10．02、2．39、0．91倍；TIMPImRNA表达量分别为PBS对照组的6．08、3．8l、2．67倍；Ⅱ型胶mRNA表达量分别为PBS对照组的145．51％、174．72％、113．73％；蛋白多糖mRNA表达量分别为PBS对照组的461．19％、191．46％、301．39％；蛋白多糖合成效率为分别PBS对照组的455．06％、285．97％、165．58％。结论CTGF和TIMPI双基因联合转染后可以在体内较长期表达，并能促进体内蛋白多糖和Ⅱ型胶原的合成，延缓椎间盘的退变。Objective To study the biological effect of connective tissue growth factor （CTGF） and tissue inhibitor of metalloproteinases 1 （TIMPI） mediated by adeno-associated virus 2 （AAV2） to rhesus monkey lumbar intervertebral discs in vivo. Methods To establish a novel model of lumbar disc degenera- tion in the rhesus monkey using percutaneous needle puncture guided by CT. The rAAV2-CTGF-IRES- TIMPI was injected percutaneously with minimal invasive technique. The mRNA expression of CTGF and TIMP1 was determined through RT-PCR. The biological effect of rAAV2-CTGF-IRES-TIMPI was examined by RT-PCR, ~5S incorporation assay. Results At the 8th0 16th, 24th week ＇after gene transfection, the ex- pression of CTGF mRNA were 10.02, 2.39, 0.91 times respectively compared with the PBS control group; TIMPI mRNA expression were 6.08, 3.81, 2.67 times respectively compared with the PBS control group; col- lagen type II mRNA expression for the PBS control group were 145.51%, 174.72%, 113.73% respectively; proteoglycan mRNA expression were 461.19%, 191.46%, 301.39% respectively compared with the PBS con- trol group; the efficiency of proteoglycan synthesis were 455.06%, 285.97%, 165.58% respectively compared with PBS control group. Conclusion The longer-term expression of CTGF and TIMPI genes mediated by adeno-associated virus can achieve in vivo after transferring into the intervertebral disc, and ＇also can in- crease the proteoglycan and collagen type II synthesis, which delay disc, degeneration.

关 键 词：腺病毒 猴 基质金属蛋白酶1 恒河猴 椎间盘 

分 类 号：R681.53[医药卫生—骨科学]