Human immunodeficiency virus type-1 (HIV-1) chronic infection causes millions of deaths each year. We previously developed a novel HIV-1 Gag-spe cific exosome (EXO)-targeted T cell-based vaccine (Gag-Texo) using ConAs...Human immunodeficiency virus type-1 (HIV-1) chronic infection causes millions of deaths each year. We previously developed a novel HIV-1 Gag-spe cific exosome (EXO)-targeted T cell-based vaccine (Gag-Texo) using ConAstimulated polyclonal CD8+T (ConA-T) cells armed with Gag-specific dendritic cell (DC)-released EXOs, and showed that Gag-Texo stimulated more efficient cytotoxic T lymphocyte (CTL) responses than DCs. Tat HIV-1 early regulatory protein possesses immunomodulatory and adjuvant properties. To enhance Gag-Texo immunogenicity, we generated Tat-engineered OVA/Tat Texo and Gag/Tat-Texo vaccines using ConA-T cells armed with EXOs release by DCs infected with recombinant OVA/Tat- and Gag/Tat-expressing adenoviruses (AdVOVA/Tat and AdVGag/Tat). We then assessed vaccination-stimulated CTL responses in naive mice, and therapeutic immunity in transgenic HLA-A2 mice bearing Gag/HLA-A2-expressing BL6-10OVA/A2 melanoma lung metastases. We demonstrate that the OVA/Tat-Texo vaccine enhances functional OVA-specific CTL responses, compared to the OVA-Texo vaccine, and broadens CTL responses recognizing the cryptic OVA epitope in C57BL/6 mice. Furthermore, we determine that the Gag/Tat-Texo not only stimulates more efficient CTL responses than Gag-Texo, but also induces enhanced therapeutic immunity. We show that, 30% of Gag/Tat-Texo-immunized mice are free of tumor lung-metastases, compared to all Gag-Texo-immunized mice displaying lung-metastasis. In addition, the average number of tumor lung metastases colonies (32/lung) in the Gag/Tat-Texo-immunized mice was also significantly lower than that (78/lung) observed in Gag-Texo-immunized mice. Taken together, this indicates that HIV-1 Gag/Tat-Texo capable of stimulating enhanced Gag-specific CTL responses and therapeutic immunity may become a new immunotherapeutic vaccine candidate for controlling virus in HIV-1 patients.展开更多
目的探讨不同细胞因子鸡尾酒诱导的树突状细胞(DC)疫苗对裸鼠移植瘤的抗肿瘤作用。方法从38名外周血干细胞移植术供者的外周血中分离单个核细胞(PBMC),诱导成未成熟DC,将人肺腺癌细胞A549总RNA电转染入DC,分别用传统细胞因子鸡尾酒(IL-6...目的探讨不同细胞因子鸡尾酒诱导的树突状细胞(DC)疫苗对裸鼠移植瘤的抗肿瘤作用。方法从38名外周血干细胞移植术供者的外周血中分离单个核细胞(PBMC),诱导成未成熟DC,将人肺腺癌细胞A549总RNA电转染入DC,分别用传统细胞因子鸡尾酒(IL-6、IL-1β、TNF-α、PGE2)和改良细胞因子鸡尾酒(IL-1β、IL-6、TNF-α、PGE2、polyⅠ∶C、CpG ODN)刺激DC成熟。致敏DC与自体T细胞混合培养,诱导产生抗原特异性细胞毒T淋巴细胞(CTL),用流式细胞术鉴定T细胞表型,ELISA法检测IFN-γ、TNF-α、IL-4、IL-5和IL-10的分泌。建立肺腺癌A549荷瘤裸鼠模型,随机分为空白对照组、未成熟组、传统组和改良组(n=5),于肿瘤接种d15、d22、d29给予相应CTL治疗,测量肿瘤体积和重量,免疫组化法和Western blotting检测肿瘤组织COX-2、VEGF、Bcl-2和Bax蛋白表达水平。结果1)CD3+CD8+T细胞表达率(%):未成熟组、传统组和改良组分别为35.00±3.24 vs 57.10±2.69 vs 63.98±1.96(P<0.05);2)IFN-γ浓度(pg/mL):3组分别为160.12±42.01 vs 263.17±55.30 vs 1 034.30±253.07(P<0.05);TNF-α浓度(pg/mL):3组分别为72.72±9.13 vs 65.20±8.03 vs 295.89±123.80(P<0.05);IL-10浓度(pg/mL):3组分别为7.26±1.76 vs 31.06±4.19 vs 44.01±12.12(P<0.05);3组IL-4、IL-5含量均未检测出,可见改良组T细胞分泌Th1型细胞因子IFN-γ和TNF-α的能力较未成熟组和传统组明显增强;3)经CTL免疫治疗后,裸鼠肿瘤体积(mm3):3组分别为512±33 vs 345±63 vs203±52(P<0.05),且改良组抑瘤率最大,为69.62%;4)改良组DC诱导的CTL明显上调Bax的表达,下调COX-2、VEGF和Bcl-2的表达。结论改良细胞因子鸡尾酒诱导肺腺癌细胞总RNA转染的DC疫苗能诱导Th1型免疫应答及产生有效的抗原特异性CTL,对肺腺癌裸鼠移植瘤具有良好的抑制作用。展开更多
文摘Human immunodeficiency virus type-1 (HIV-1) chronic infection causes millions of deaths each year. We previously developed a novel HIV-1 Gag-spe cific exosome (EXO)-targeted T cell-based vaccine (Gag-Texo) using ConAstimulated polyclonal CD8+T (ConA-T) cells armed with Gag-specific dendritic cell (DC)-released EXOs, and showed that Gag-Texo stimulated more efficient cytotoxic T lymphocyte (CTL) responses than DCs. Tat HIV-1 early regulatory protein possesses immunomodulatory and adjuvant properties. To enhance Gag-Texo immunogenicity, we generated Tat-engineered OVA/Tat Texo and Gag/Tat-Texo vaccines using ConA-T cells armed with EXOs release by DCs infected with recombinant OVA/Tat- and Gag/Tat-expressing adenoviruses (AdVOVA/Tat and AdVGag/Tat). We then assessed vaccination-stimulated CTL responses in naive mice, and therapeutic immunity in transgenic HLA-A2 mice bearing Gag/HLA-A2-expressing BL6-10OVA/A2 melanoma lung metastases. We demonstrate that the OVA/Tat-Texo vaccine enhances functional OVA-specific CTL responses, compared to the OVA-Texo vaccine, and broadens CTL responses recognizing the cryptic OVA epitope in C57BL/6 mice. Furthermore, we determine that the Gag/Tat-Texo not only stimulates more efficient CTL responses than Gag-Texo, but also induces enhanced therapeutic immunity. We show that, 30% of Gag/Tat-Texo-immunized mice are free of tumor lung-metastases, compared to all Gag-Texo-immunized mice displaying lung-metastasis. In addition, the average number of tumor lung metastases colonies (32/lung) in the Gag/Tat-Texo-immunized mice was also significantly lower than that (78/lung) observed in Gag-Texo-immunized mice. Taken together, this indicates that HIV-1 Gag/Tat-Texo capable of stimulating enhanced Gag-specific CTL responses and therapeutic immunity may become a new immunotherapeutic vaccine candidate for controlling virus in HIV-1 patients.
文摘目的探讨不同细胞因子鸡尾酒诱导的树突状细胞(DC)疫苗对裸鼠移植瘤的抗肿瘤作用。方法从38名外周血干细胞移植术供者的外周血中分离单个核细胞(PBMC),诱导成未成熟DC,将人肺腺癌细胞A549总RNA电转染入DC,分别用传统细胞因子鸡尾酒(IL-6、IL-1β、TNF-α、PGE2)和改良细胞因子鸡尾酒(IL-1β、IL-6、TNF-α、PGE2、polyⅠ∶C、CpG ODN)刺激DC成熟。致敏DC与自体T细胞混合培养,诱导产生抗原特异性细胞毒T淋巴细胞(CTL),用流式细胞术鉴定T细胞表型,ELISA法检测IFN-γ、TNF-α、IL-4、IL-5和IL-10的分泌。建立肺腺癌A549荷瘤裸鼠模型,随机分为空白对照组、未成熟组、传统组和改良组(n=5),于肿瘤接种d15、d22、d29给予相应CTL治疗,测量肿瘤体积和重量,免疫组化法和Western blotting检测肿瘤组织COX-2、VEGF、Bcl-2和Bax蛋白表达水平。结果1)CD3+CD8+T细胞表达率(%):未成熟组、传统组和改良组分别为35.00±3.24 vs 57.10±2.69 vs 63.98±1.96(P<0.05);2)IFN-γ浓度(pg/mL):3组分别为160.12±42.01 vs 263.17±55.30 vs 1 034.30±253.07(P<0.05);TNF-α浓度(pg/mL):3组分别为72.72±9.13 vs 65.20±8.03 vs 295.89±123.80(P<0.05);IL-10浓度(pg/mL):3组分别为7.26±1.76 vs 31.06±4.19 vs 44.01±12.12(P<0.05);3组IL-4、IL-5含量均未检测出,可见改良组T细胞分泌Th1型细胞因子IFN-γ和TNF-α的能力较未成熟组和传统组明显增强;3)经CTL免疫治疗后,裸鼠肿瘤体积(mm3):3组分别为512±33 vs 345±63 vs203±52(P<0.05),且改良组抑瘤率最大,为69.62%;4)改良组DC诱导的CTL明显上调Bax的表达,下调COX-2、VEGF和Bcl-2的表达。结论改良细胞因子鸡尾酒诱导肺腺癌细胞总RNA转染的DC疫苗能诱导Th1型免疫应答及产生有效的抗原特异性CTL,对肺腺癌裸鼠移植瘤具有良好的抑制作用。
