shRNA慢病毒转染串珠素对大鼠硬膜外瘢痕中成纤维细胞增殖能力的影响  

Transfection of perlecan by shRNA lentivirai particles on proliferation of fibroblasts in epidural scar

作　　者：马青跃[1] 褚言琛[1] 王进[1] 候庆先[1] 袁万青[1] 杨慧英[1] 邹云雯[1] 

机构地区：[1]青岛大学医学院附属医院骨外科,266003

出　　处：《中华创伤骨科杂志》2013年第3期252-255,共4页Chinese Journal of Orthopaedic Trauma

基　　金：国家自然科学基金（81071482）

摘　　要：目的通过研究经shRNA慢病毒转染串珠索的大鼠硬膜外瘢痕组织成纤维细胞的增殖能力．探讨其对硬膜外瘢痕组织形成的影像。方法30只Wistar大鼠行椎板切除术，建立模型，于培养4周后提取硬膜外瘢痕组织。通过组织块贴壁法培养硬膜外瘢痕组织中的成纤维细胞井将细胞分为3组：未转染组（空白对照组）、荧光蛋白（GFP）慢病毒转染组（GFP组）和shRNA慢病毒转染组（shRNA组．内含GFP）。成纤维细胞转染成功后，分别通过Western—blot、荧光定城PCR及MTT法榆测3组的成纤维细胞的RNA、蛋白质表达及细胞增殖的差异。结果Weste11-blot检测显示shRNA纰串珠素蛋白明显低f空白对照组和GFP组。荧光定量PCR检测显示shRNA组串珠素RNA表达低下空白对照组和GFP组。MTT检测显示0h各绀吸光度（A）值差异均无统计学意义（P〉0．05），24～96hshRNA组吸光度值（A）低于空白对照组和GFP组，差异均有统计学意义（P〈0．05）。结论经shRNA慢病毒转染串珠素能抑制大鼠硬膜外瘢痕组织成纤维细胞的增殖。Objective To research the proliferation of fibroblasts in epidural sear transfected by perlecan shRNA lentiviral particles. Methods Experimental models were established in 30 Wistar mice which undewent lamioectomv. The epidural scar tissue was harvested after culture tor 4 days. The fibrobiasts in the epidural scar tissue were cultured by tissue explant adherence before they were randomly assigned into no transfection （contrnl group）, green fluorescent protein （GFP） lentiviral transfectioo （GFP group） and shRNA lenliviral transfection （shRNA group） . After the fibroblasts were successfully transtected, their ex- pressions of perlecan mRNA and protein and proliferation were detected by Western-blot. RT-PCR and TT and compared as well among the 3 groups. Results Western-blot and RT-PCR showed the expressions of perleean mRNA and protein were significantly lower in the shRNA group than in the control and GFP groups （ P 〈 0.05）. MTT showed no significant difference among the 3 groups in the optical density at 0 hour （ P 〉 0.05） but significantly lower optical density in the shRNA group than in the control and GFP groups at 24 to 96 hours （ P 〈 0. 05）. Conclusion Growth of fibroblasts in epidural sear can be inhibited significantly by transfeetion with perlecan shRNA lentiviral particles.

关 键 词：RNA 转染 成纤维细胞 瘢痕 慢病毒属 

分 类 号：R651[医药卫生—外科学]