VP22增强人巨细胞病毒pp65核酸疫苗在小鼠体内免疫活性的实验研究  

VP22 enhances the immunological activity of human cytomegalovirus pp65 DNA vaccine: experimental study with mice

作　　者：马道新[1] 于修平[2] 张晓梅[2] 周亚滨[2] 李勇[1] 贾继辉[2] 赵蔚明[2] 

机构地区：[1]山东大学齐鲁医院,济南250012 [2]山东大学医学院

出　　处：《中华医学杂志》2005年第15期1049-1052,共4页National Medical Journal of China

基　　金：国家自然科学基金资助项目(30070321)

摘　　要：目的构建人巨细胞病毒(HCMV)主要被膜磷蛋白pp65(HCMVpp65)基因高表达载体以及VP22.pp65联合表达载体,并评价VP22增强pp65核酸疫苗的体内免疫效果。方法8周龄正常BALB/C雌性小鼠分为pcDNA3.pp65组、pVP22.pp65组、实验对照组,每组7只。利用分子克隆技术构建HCMVpp65表达载体pcDNA3.pp65、VP22与pp65融合基因载体pVP22.pp65,免疫小鼠,四甲基偶氮唑蓝法测定T淋巴细胞增殖反应、白细胞介素(IL)2生物学活性,乳酸脱氢酶释放法测定自然杀伤细胞(NK)活性,酶联免疫吸附实验测定血清HCMVIgM、IgG抗体含量、血清及脾细胞培养液中的IL2、IL4含量。结果小鼠的一般状况始终良好,成功构建了HCMV核酸疫苗载体pcDNA3.pp65、pVP22.pp65;免疫小鼠后,两种疫苗T淋巴细胞增殖反应(8周时刺激指数分别为5.11和5.55)和NK细胞活性(12周分别为8.74%和12.08%)及HCMVIgM(6周时A值分别为1.20和1.58)、IgG抗体(6周时A值分别为1.09和1.78)均高于对照组,且pVP22.pp65组高于pcDNA3.pp65组(P<0.05)。小鼠血清中IL2和IL4含量及IL2的生物学活性较低,各组间差异无统计学意义(均P>0.05);小鼠脾细胞上清中IL2和IL4含量以pVP22.pp65组(411.11pg/ml、76.10pg/ml)最高,IL2生物学活性也以pVP22.pp65组(A值为0.22)最高。结论HCMV核酸疫苗无毒性。Objective To construct the human cytomegalovirus(HCMV)pp65 DNA vaccine vector and VP22 and pp65 coexpressing vector. To evaluate and compare immunological effects in mice. Methods^Twenty-one BALB/C mice were divided into 3 equal groups: pcDNA3.pp65 group undergoing injection of pcDNA3.pp65 as DNA vaccine, pVP22.pp65 group undergoing injection of pVP22.pp65, and control group undergoing injection of normal saline. HCMV pp65 expression vector pcDNA3.pp65, VP22 and pp65 coexpressing vector pVP22.pp65 were constructed by molecular biology methods. The vectors were immunized into BALB/C mice as DNA vaccines. The T cell proliferation activity and IL-2 biological activity were determined using MTT method. NK activity was detected using LDH release test. The serum IgM and IgG levels of HCMV, the concentrations of IL-2 and IL-4 in serum and supernatant of spleen cells were detected using ELISA method. Results The HCMV DNA vaccine expression vectors were successfully constructed. Some indexes of the two vaccine groups ( pcDNA3.pp65 and pVP22.pp65), that is, T cell proliferation activity (5.11 and 5.55 for SI at 8~th week respectively) , NK activity (8.74% and 12.08% at 12~th week respectively), IgM level (1.20 and 1.58 for A value at 6~th week respectively) and IgG level ^(1.09 and 1.78 for A value at 6^th week respectively) were higher than those of negative control, and pVP22.pp65 group was higher than pcDNA3.pp65 group (P<0.05). The concentrations of IL-2 and IL-4 and the IL-2 biological activity were very low in sera of three groups which showed no significant difference between them (P>0.05), but higher in the spleen supernatant and the pVP22.pp65 group was highest (411.11 pg/ml, 76.10 pg/ml for the concentrations of IL-2 and IL-4 and 0.22 for IL-2 biological activity). Conclusion The HCMV pp65 could induce certain immunological activity, and VP22 could significantly enhance pp65 in vivo immunological activity.

关 键 词：人巨细胞病毒PP65 核酸疫苗 VP22 实验研究 免疫活性 体内 T淋巴细胞增殖反应 pcDNA3 四甲基偶氮唑蓝法 乳酸脱氢酶释放法 酶联免疫吸附实验 生物学活性 IL-2 IgG抗体 表达载体 BALB/C 分子克隆技术 融合基因载体 自然杀伤细胞 

分 类 号：R392.1[医药卫生—免疫学]