CAJ | 학술논문

目的研究所构建的新型胰腺癌MUC1 DNA疫苗的免疫原性.方法采用三种优化策略共构建4个重组质粒,接种免疫C57BL/6J (H 2b)雌性小鼠.每2周加强免疫一次,共免疫3次.每次免疫前及处死小鼠前检测抗体和细胞因子.最后一次免疫结束后2周取脾,体外经VNTR合成肽刺激培养后进行杀伤性T淋巴细胞(cytotoxic T lymphocyte,CTL)杀伤功能分析.结果 pIRES2-EGFP3VNTR组、pIRES2-EGFP-3VNTR-CI-144组、pIRES2-EGFP-3VNTR-mIL-18组、pIRES2-EGFP-3VNTR-CI-144-mIL-18组的CTL特异杀伤率显著高于空载体对照组和生理盐水对照组,三重优化构建的质粒pIRES2-EGFP-3VNTR-mIL-18的CTL特异性杀伤率高于双重优化构建和单重优化构建的质粒.各优化构建质粒组的抗MUC1抗体OD值均显著高于对照组(P＜0.05),但组间差异无统计学意义.各优化构建质粒组的外周血IFN-γ均高于对照组(P＜0.05),且以含有IL-18优化策略的两组为高(P＜0.05).结论所构建的优化重组质粒免疫小鼠后均可诱发抗原特异的CTL应答和抗体应答.C1-144和IL-18可增强疫苗的免疫原性.
목적 연구소구건적신형이선암MUC1 DNA역묘적면역원성.방법 채용삼충우화책략공구건4개중조질립,접충면역C57BL/6J (H 2b)자성소서.매2주가강면역일차,공면역3차.매차면역전급처사소서전검측항체화세포인자.최후일차면역결속후2주취비,체외경VNTR합성태자격배양후진행살상성T림파세포(cytotoxic T lymphocyte,CTL)살상공능분석.결과 pIRES2-EGFP3VNTR조、pIRES2-EGFP-3VNTR-CI-144조、pIRES2-EGFP-3VNTR-mIL-18조、pIRES2-EGFP-3VNTR-CI-144-mIL-18조적CTL특이살상솔현저고우공재체대조조화생리염수대조조,삼중우화구건적질립pIRES2-EGFP-3VNTR-mIL-18적CTL특이성살상솔고우쌍중우화구건화단중우화구건적질립.각우화구건질립조적항MUC1항체OD치균현저고우대조조(P＜0.05),단조간차이무통계학의의.각우화구건질립조적외주혈IFN-γ균고우대조조(P＜0.05),차이함유IL-18우화책략적량조위고(P＜0.05).결론 소구건적우화중조질립면역소서후균가유발항원특이적CTL응답화항체응답.C1-144화IL-18가증강역묘적면역원성.
Objective The development of cancer vaccines deserves experimentation,specifically the immunogenicity of the new MUC1 DNA vaccine for pancreatic cancer.Methods Three strategies were combined to optimize the new MUC1 DNA vaccine.The female C57BL/6 mice were immunized,through tibial muscle injection,with 100 μg of plasmid DNA of the recombinant plasmids (pIRES2-EGFP-3VNTR group,pIRES2-EGFP-3VNTR-C1-144 group,pIRES2-EGFP 3VNTR-mIL-18 group,pIRES2-EGFP-3VNTR-C1-144-mIL-18 group,n=5) for a total volume of 100 μl.Mice inoculated with the empty vector pIRES2-EGFP (EV group,n=5) and normal saline (NS group,n=5) were used as vector and blank controls,respectively.All the mice were immunized again every two weeks.Two weeks after the third immunization,all the mice were euthanized and spleen cells were separated for CTL cytotoxic assay.Results The specific cytolysis percentages of the four groups (pIRES2-EG-FP-3VNTR,pIRES2-EGFP-3 VNTR-C1-144,pIRES2-EGFP-3VNTR-mIL-18,pIRES2-EGFP 3VNTR-C1-144-mIL-18) expressing VNTR were higher than the EV and NS group with the effector/target cells ratio (E/T) from 80:1 to 20:1 (P＜0.05).Therefore,it showed a difference among the four groups.After the primary immunization,the OD450 of the serum antibody level specific to MUC1 began to increase in the four groups which contained the gene of VNTR3 (P＜0.05).This suggests that the recombinant plasmids could induce a specific antibody response to MUC1,and showed no remarkable difference among the four groups.IFN-γ serum cytokine among the four groups were higher than that of the EV and NS groups (P＜0.05).There was a significant difference of OD450 between the groups containing mIL-18 pIRES2-EGFP-3VNTR-mIL-18,pIRES2-EGFP-3VNTR-C1-144-mIL-18) and those not (pIRES2-EGFP-3VNTR,pIRES2-EGFP-3VNTR-C1-144,)(P＜0.05).Conclusions In conclusion,all of the four recombinant plasmids could induce MUC1 specific CTL and antibodies responses,and C1-144 and IL-18 could enhance the immunogenicity of plasmids.