| Construction of genetically engineered macrophages expressing Smad6 andSmad7 genes with adeno-associated virus |
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| 引用本文： | 黄云剑 赵景宏 杨唐俊 范晓棠 张金海 蔡文琴. Construction of genetically engineered macrophages expressing Smad6 andSmad7 genes with adeno-associated virus[J]. 中国人民解放军军医大学学报, 2004, 19(2): 71-75,80 |
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| 作者姓名： | 黄云剑 赵景宏 杨唐俊 范晓棠 张金海 蔡文琴 |
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| 作者单位： | [1]DepartmentofNephrology,XinqiaoHospital,ThirdMilitaryMedicalUniversity,Chongqing400037,China [2]DepartmentofHistologyandEmbryology,ThirdMilitaryMedicalUniversity,Chongqing400038,China |
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| 基金项目： | theNationalNaturalScienceFoundationofChina(No .30 10 0 0 83) ,the“Tenthfive year plan”MedicalScienceFoun dationofPLA(No .0 1Q10 3) ,andthePostdoctorResearchFounda tionofChina(No .2 0 0 30 3336 4 ) |
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| 摘 要： | To construct the genetically engineered macrophages expressing Smad6 and Smad7 genes with adeno-associated virus (AAV). Methods: The plasmids containing peDNA3-Smad6/Flag and peDNA3-Smad7/Flag were digested with BamH I and Xho I , respectively. Then the Smad6/Flag and Smad7/Flag gene segments obtained were cloned into plasmid pAAV-MCS respectively to construct the recombinant pAAV-Smad6/Flag and pAAV-Smad7/Flag plasmids. The resulting recombinant plasmids (pAAV-Smad6/Flag or pAAV-Smad7/Flag) or pAAV-LacZ plasmid were co-transfected into the HEK 293cells with pHelper and pAAV-RC by calcium-phosphate precipitation method. Recombinant AAV-2 viral particles were prepared from infected HEK293 cells and then were used to infect mouse macrophages. The expressions of Smad6and Smad7 in macrophages were detected by immunocytochemical staining and expression of b-galactosidase was evaluated by X-gal staining. Results: The recombinant AAV vector containing Smad6 or Smad7 genes was successfully constructed. More than 95% macrophage cells expressed X-gal and Smad6 and Smad7 genes at 72 h after infection. Conclusion: These results indicate that the genetically engineered macrophages can express Smad6 and Smad7 proteins effectively, laying the foundation for the studies of TGF-β-induced diseases in vivo and highlighting the feasibility of macrophage-based gene therapy.
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| 关 键 词： | 腺相关病毒 巨噬细胞 基因疗法 Smad6基因 Smad7基因 肾脏疾病 |
Construction of genetically engineered macrophages expressing Smad6 and Smad7 genes with adeno-associated virus |
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| HUANG Yun-jian ,ZHAO Jing-hong ,YANG Tang-jun ,FAN Xiao-tang ,ZHANG Jin-hai ,CAI Wen-qin. Construction of genetically engineered macrophages expressing Smad6 and Smad7 genes with adeno-associated virus[J]. Journal of Medical Colleges of PLA(China), 2004, 19(2): 71-75,80 |
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| Authors: | HUANG Yun-jian ZHAO Jing-hong YANG Tang-jun FAN Xiao-tang ZHANG Jin-hai CAI Wen-qin |
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| Affiliation: | HUANG Yun-jian 1,ZHAO Jing-hong 1,YANG Tang-jun 1,FAN Xiao-tang 2,ZHANG Jin-hai 2,CAI Wen-qin 2 1Department of Nephrology,Xinqiao Hospital,Third Military Medical University,Chongqing 400037,China 2Department of Histology and Embryology,Third Military Medical University,Chongqing 400038,China |
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