重组质粒介导hTGF-β1基因在体转染模型动物椎间盘髓核细胞的表达

The expression of the recombinant plasmid mediated transfection of the human transforming growth factorβ1 encoding gene in the intervertebral disc of animal models

目的观察以质粒为载体的外源治疗基因pACCMV-hTGF-β1在体转染模型动物腰椎间盘髓核细胞后的表达。方法将扩增后的重组质粒基因注射于模型动物的腰椎间盘髓核细胞中,手术后不同时间取出椎间盘,用免疫组织化学方法对椎间盘髓核细胞进行基因表达产物的测定;并利用CMIAS图像分析系统进行半定量观察。结果免疫组织化学染色显示注射pACCMV-hTGF-β1椎间盘,4d组即呈现阳性颗粒,阳性表达可持续至少2周,实验对照组阳性表达仅维持3d,空白组呈阴性反应。结论椎间盘髓核作为靶细胞能够被外源基因转染,并且能在相当长时间内表达TGF-β1蛋白。

Objective To deliver an exogenous therapeutic gene-hTGF-β1 to rabbit lunbar intervertebral disc in vivo by plasmid vector and to observe the expression of the exogenous therapeutic gene. Methods The recombinant gene plasmid after amplified was injected into the nucleus pulposus cells of intervertebral disc of the animal models, and the intervertebral discs were taken out at different time points after operation, then immunohistochemical staining was used to determine the gene expression, and a half quantitative observation was performed by CMIAS image analysis system. Results The results of immunohistochemical staining showed that the intervertebral discs injected with pACCMV- hTGF-β1 exhibited positive granule at 4 days after injection, which lasted for at least 2 weeks, however, which in control group lasted 3 days only, furthermore, the blank group showed negative reaction. The OPIDM was significantly increased in the intervertebral discs injected with pACCMV- TGF-β1, as compared with that in blank group or protein group （ P 〈 0. 05 ）. Conclusion The nucleus pulposus of intervertebral disc of the rabbits can be transfected by the plasmid mediated vector carrying the human transforming growth factor β1 encoding gene, and can effectively express TGF-β1 protein for comparative long time.