摘要
目的 :观察已构建好的腺病毒介导的骨形态蛋白 7基因感染大鼠髓腔内细胞的情况 ,为腺病毒介导BMP 7的进一步基因治疗提供可靠的实验基础。材料和方法 :从人骨肉瘤细胞株U 2OS中获得hBMP 7的cDNA片段 ,插入穿梭质粒后在大肠杆菌内与腺病毒实现同源重组 ,阳性克隆转染 2 93细胞 ,扩增纯化得到大量病毒后注入大鼠椎体髓腔中 ,用冲洗髓腔的方法和流式细胞仪分析来观察注射椎体和上位椎体髓腔内细胞感染腺病毒BMP 7的情况 ,并观察病毒感染的时限。结果 :经体外检测能高效表达活性BMF 7的构建好的AdBMP 7注入椎体髓腔后 ,髓腔冲洗液的荧光显微镜观察 ,术后 1 2d和 2 4d都有感染了带有荧光蛋白标记的髓腔内细胞 ,而且在注射椎体的上位椎体中也出现了感染病毒的细胞。结论 :腺病毒介导的BMP -7基因能够在体内感染髓腔细胞并分泌活性的BMP -7,可以成为基因治疗的有效手段。
Objective: To evaluate the Ad-BMP-7 infecting the bone marrow cells in rats and set a gene therapy basis with Ad-BMP-7.Method:Transgenes BMP-7 and green fluorescence protein (GFP) was cloned into the AdEasy ^(TM) vector system and injected into the vertebral bodies of rats,fluorescence microscopy and flow cytometer were used to detect the bone marrow cells infected by Ad-BMP-7.Result:Rinsing solution of bone marrow observed by fluorescence microscope and flow cytometer showed that the Ad-BMP-7 infected the cells in the bone marrow.And Luciferase activity persisted within the vertebral bodies for at least 3 weeks.Conclusion:Ad-BMP-7 injected into the vertebral bodies of rats may infect the bone marrow cells for three weeks.It is a potential gene therapy.
出处
《中国矫形外科杂志》
CAS
CSCD
2004年第12期927-929,共3页
Orthopedic Journal of China
