两种抗原致敏方式负载树突状细胞疫苗对LNCaP细胞株的体外杀伤作用

Effects of two different antigen-loading methods on activity of dendritic cell vaccine for LNCaP cell line inhibition in vitro

目的:比较验证重组腺相关病毒负载PSA基因转染方式与LNCaP细胞裂解物诱导方式对产生树突状细胞(DCs)疫苗的的影响.方法:抽取HLA表型为A2的健康志愿者外周血,分离单核细胞,体外培养.通过反复冻融LNCaP细胞、提取肿瘤细胞裂解物以及使用含PSA片段的重组腺相关病毒转染未成熟DCs,诱导特异性细胞毒性T细胞(CTL).检测体外培养的DCs和CTL活性,并使用MTT法检测两组CTL对LNCaP细胞的杀伤作用,同时使用腺相关病毒转染组诱导的CTL对LNCaP和DU145两种前列腺癌细胞进行杀伤实验检测.结果:两种方式均可培养成熟的DCs,诱导激活CTL并分泌IFN-γ,而腺相关病毒转染的DCs成熟度和CTL诱导率均高于细胞裂解物组;腺相关病毒转染组诱导的CTL对LNCaP细胞的杀伤率显著高于细胞裂解物组诱导的CTL;腺相关病毒转染组诱导的CTL对LNCaP细胞有特异性杀伤作用.结论:两种抗原递呈方式均可培养出成熟的DCs,并可诱导自体CTL增殖;而使用重组腺相关病毒转染PSA至DCs的方式明显优于肿瘤细胞裂解物的抗原负载方式,具有高效特异性.

AIM： To observe the changes in the activity of dendritic cells（DCs） vaccine after prostate specific antigen（PSA） gene transfeetion mediated by recombinant adeno-associated virus type2（rAAV） and tumor cell lysate. METHODS： Immature DCs were isolated from peripheral blood monocytes of healthy volunteers positive with HLA-A2, and were infected with the recombinant adeno-associated virus carrying PSA gene or loaded with tumor cell lysate. Then cytotoxic T cells （CTLs） were induced by these 2 kinds of DCs. The activity of DCs and CTLs were checked, meanwhile the specific killing activity of 2 kinds of CTLs to LNCaP cells and the former to LNCaP and DU145 cells were assessed by MTr assay. RESULTS： Mature DCs were obtained in both groups, but DCs in the virus transfection group were more mature and induced more CTLs. The killing effect of CTLs resulting from rAAV transfection of the DCs was more significant than that induced by tumor cell lysate loading, and CTLs transfected by rAAV/PSA specifically recognized LNCaP cells and killed them. CONCLUSION： Both methods of antigen loading can induce mature DCs from peripheral blood monocytes, but rAAV infection of the DCs is more effective than tumor cells lysate loading. DCs transfected with rAAV may have the potency to serve as an adjuvant immunotherapy for patients with prostate cancer.