DNA免疫激发针对肾母细胞瘤CTL效应初步研究

目的 将鼠源肾母细胞瘤WT1基因一段序列(WT1y)构建于真核表达质粒pCDNA3.1( ),通过小鼠和体外细胞模型初步研究所激发的针对肾母细胞瘤CTL效应.方法 人工合成WT1基因一段序列,含有HLA-A*2402锚定残基的9个氨基酸.构建重组真核表达质粒pCDNA3.1( )/WT1y.免疫Balb/c小鼠,通过ELISA方法检测体液免疫反应;分离脾淋巴细胞,FCM检测脾淋巴细胞中CD4/CD8;将分离的脾淋巴细胞与小鼠肾母细胞瘤细胞共培养,检测其体外溶解组织相容性抗原型别一致的外源性靶细胞能力.结果 构建的重组质粒经测序鉴定,与GenBank中登录的序列完全一致;免疫组小鼠T淋巴细胞增殖及CTL活性均明显优于对照组(P＜0.01);体外具有较强溶解靶细胞的功能.结论 WT1基因的DNA疫苗初步研究具有很强的CTL效应,为小儿肾母细胞瘤的治疗提供了新的思路.

Human specific CTL responses to DNA vaccine of WT1y product

Objective To construct the eukaryotic expression vector pCDNA3.1( ) containing WT1y gene an assay the specific CTL responses to DNA vaccine of Wilms' tumor gene(WT1y) product.MethodsThe WT1y gene of 321 bp,including HLA-A*2402 anchoring residue,was synthetized,and then transfected into eukaryotic vector calledpCDNA3.1( ).The constructed eukaryotic vector calledpCDNA3.1( )/WT1y was used to immunize Balb/c mice.WT1-specific antibody was detected in serum of mice by ELISA method.T lymphocyte proliferation was detected by FCM.The CTLs response was detected by co-culture the murine spleen cells and the tumor cells.ResultsRecombinant gene was confirmed using DNA sequencing.WT1-specific antibodies were detected in serum of immunized mice by ELISA.T lymphocyte proliferation of immunized mice was superior to that of control group(P<0.01).The CTLs exerted specific lysis against WT1-expressing cells.ConclusionThe DNA vaccine of WT1y can induce specific CTLs,which offers a new way for treatment of Wilms' tumor.