氨肽酶N启动子调控的MnSOD基因对骨髓细胞的特异性辐射防护作用  

MnSOD Gene Regulated by Aminopeptid ase N Promoter Specifically Protects Bone Marrow from Radiation

作　　者：梁利波[1] 马业伟[1] 赵清正1 周小山[1] 杨军[2] 李艳春[1] 刘玉英[1] 王争[1] 章扬培[2] 

机构地区：[1]中国协和医科大学中国医学科学院肿瘤研究所,北京100021 [2]军事医学科学院野战输血研究所,北京100850

出　　处：《癌症》2002年第9期939-943,共5页Chinese Journal of Cancer

基　　金：国家自然科学基金(No.39970818)

摘　　要：背景与目的:骨髓造血细胞中导入耐辐射基因是克服放疗对造血系统抑制作用的有效手段,但这也增加了肿瘤细胞的辐射耐受性。本实验旨在探索不明显增加肿瘤细胞对辐射抗拒性的同时,能特异性保护骨髓细胞免受辐射导致的损伤的方法。方法:构建以氨肽酶N(aminopeptidaseN,APN)基因骨髓特异性启动子调控的锰超氧化物歧化酶(manganesesuperoxidedismutase,MnSOD)基因逆转录病毒载体,并导入成骨髓细胞KG1a和肝癌细胞BEL7402中。用RT-PCR分析MnSODmRNA水平,测定细胞中MnSOD活性,以细胞存活实验检测骨髓细胞和肝癌细胞对X射线的敏感性,并用流式细胞术和断裂DNA电泳方法对辐射诱导的细胞凋亡进行分析。结果:导入基因的KG1a细胞中MnSODmRNA水平和酶活性明显升高。APN骨髓特异性启动子调控的MnSOD基因表达能有效抑制辐射引起的KG1a细胞凋亡。导入基因的KG1a细胞对辐射的耐受性提高,在用10Gy剂量照射时,KG1a细胞存活率比原来增加3.7倍。而导入MnSOD基因并未使BEL7402细胞中MnSODmRNA水平和酶活性提高,其放射敏感性未发生明显变化。结论:APN骨髓特异性启动子能控制MnSOD基因在骨髓细胞中高表达,而在癌细胞中低表达。在用X射线杀伤癌细胞的过程中,APN骨髓特异性启动子调控的MnSOD基因能特异性保护骨髓细胞。Background &Objective:It is an effective way to induce radio-tolerant gene into hematopoietic c ells in bone marrow for overcoming the suppr ession of radiotherapy on hematopoietic system.However,this also increases the radiation tolerance of tumor cells.This study was designed to investigate a method to specifically protect b one marrow cell from being damaged by radiation,along without increasing resistence of tumor cell to radiation.Methods:The retrovirus vector of manganese superoxide dismutase(MnSOD)gene regulated by aminopeptidase N(APN)bone marrow-specific gene promoter was constructed and induced into myeloblastic KG1a and c ancer cell BEL7402.MnSOD mRNA level was analyzed by PT-P CR;MnSOD activity in the cells was determined;the sensitivity of bone marrow cell and hepatic carcinoma cell to x-ray was detected by cell survival test;the cell apoptosis was analyze d with flow cytometry and fractural DNA electrophoresis.Results:The MnSOD mRNA level and enzyme activ ity in KG1a cells transferred with th e gene was obviously increased.Expre ssion of MnSOD mRNA drove by APN myelo-specific promoter effectively inh ibited apoptosis of KG1a cells induced by ra diation and endowed KG1a cell line wi th the enhancement of tolerance to ra diation,which increased by 3.7folds compare d to parental cells at the dose of 10Gy .In contrast,the level of MnSOD mRNA,the enzyme activity of MnSOD and the radi osensitivity had no significant change in BEL 7402cells transduced with MnSOD gene.Conclusion:APN bone marrow-specific promoter c ould control MnSOD gene expression h ighly in myeloid cell and lower in cancer cell.In the process o f killing of cancer cell by x-ray,MnSOD gene regulated by APN bone marrow-specific promoter could specifically protec t myeloid cell.This study provides a new clue to solve the bone marrow supp ression in high dose radiotherapy.

关 键 词：氨肽酶N 骨髓特异性启动子 锰超氧化物歧化酶基因 逆转录病毒载体 骨髓细胞 肝癌细胞 

分 类 号：R730.55[医药卫生—肿瘤]