摘要
目的利用重组复制缺陷型腺病毒AdIL 10将外源IL 10基因导入肝星状细胞 (HSC) ,并观察其对HSC增殖的影响。方法将本实验室自行构建的重组腺病毒质粒pAdIL 10用Lipofectamin包装转染HEK 2 93细胞 ,即可获得重组腺病毒AdIL 10。应用胶原酶原位灌注和密度梯度离心法分离大鼠HSC ,当HSC生长至 80 %密度时 ,以 5× 10 4细胞 /孔接种于 96孔板 ;HSC培养 2 4h后 ,取病毒AdIL 10以感染复数 (MOI) 5 0感染HSC ;用MTT法检测HSC增殖率。结果pAdIL 10经 2 93细胞包装 ,3d后可观察到绿色荧光蛋白 (GFP)表达 ;AdIL 10感染HSC 3d后可观察到GFP表达 ,且HSC增殖受到抑制。结论重组腺病毒AdIL
Objective To investigate effects of IL-10 transgene into hepatic stellate cells (HSCs) on proliferation using recombinant replication-deficient adenoviruses carrying rat IL-10 cDNA(AdIL-10). Methods The pAdIL-10 constructed by our laboratory was transferred into HEK 293 cells by lipofectamin for packing, and the recombinant replication-deficient adenovirus, AdIL-10, was finally obtained. Rat hepatic setellate cells were prepared from male Sprague-Dawley rats by the pronase/collagenase method and a single-step density gradient centrifugation with Nycodenz. Primary cultured HSCs were trypsinized at 80% confluency and seeded on a 96-well plate at a density of 5×10 4 cells per well for 24 h. Then HSCs were infected with AdIL-10 by MOI 50 . HSCs proliferation was detected by MTT test. Results GFP expression could be observed on the third day after packing of the linearized pAdIL-10 in 293 cells. GFP expression could be observed on the third day after HSCs were infected with AdIL-10 and HSCs proliferation was inhibited. Conclusion AdIL-10 could infect HSCs and inhibit HSCs proliferation.
出处
《上海第二医科大学学报》
CSCD
北大核心
2005年第2期128-130,共3页
Acta Universitatis Medicinalis Secondae Shanghai