纳米微粒包装肿瘤抗原诱导CTL抗肿瘤免疫

用纳米微粒包装肿瘤抗原,并探讨其诱导产生CTL的免疫学特性和杀瘤活性。采用复乳溶剂蒸发法制备包含有恶性黑色素瘤抗原肽Mart-1-27-35纳米微粒;扫描电镜观察所制得纳米颗粒的形态。用NP-Mart-1-27-35诱导抗原特异性CTL并用ELISPOT法和LDH法检测其特性和杀瘤活性。结果:电镜观察所制得的纳米颗粒为圆形,平均直径为(208.52±12.43)nm。HPLC分析纳米-抗原肽的抗原载入量为6.72%±0.28%,平均包封率为91.23%±3.56%;ELISPOT检测表明纳米微粒包装的抗原肽能引起较强的免疫反应,产生分泌IFN-γ的细胞斑点数分别为(476.37±29.43)/2×104,显著高于单纯使用抗原肽时所产生的斑点数;LDH检测显示与单纯使用抗原肽相比,纳米-抗原肽诱导的CTL对Mart-1+细胞的杀伤能力显著提高。纳米微粒包装的抗原肽能够显著增强并延长DC对抗原的提呈作用;其诱导产生的CTL能以MHC I限制的方式特异识别和杀伤表达相应抗原的肿瘤细胞。

Nanoparticles formulated with tumor antigen induce the anti-tumor immue responses of cytotoxic T lymphocytes

To investigate the immunological characteristics and the killing effect of cytotoxic T lymphocytes(CTLs) on tumors with nanoparticles formulated with tumor antigen,nanoparticles containing Mart-1-27-35 peptide of malignant melanoma were prepared by using double emulsion solvent evaporation method.The shape and size of NP-Mart-1-27-35 particle was analyzed using scanning electron microscopy,and this particle was used to induce the antigen-specific CTLs.Its properties and the killing effect on tumor cells were determined by ELIPOT and LDH assays.As demonstrated by scanning electron microscopy.this nanoparticle was spherical in shape with a mean diameter of(208.52±12.43) nm.The average loading and embedding ratio for antigens was 6.72%±0.28% and 91.23%±3.56% respectively as determined by using HPLC analysis.The result obtained from ELIPOT assay showed that the antigen presentation ability of human dendritic cells could be significantly enhanced due to uptake of nanoparticles formulated with this peptide.thereby to induce strong immune responses.The spot number of cells to secrete IFN-γ was(476.37±29.43)/2×104 which was higher than that simply used with antigenic peptide.As demonstrated by the LDH assay,the killing effect on Mart-1 cells of CTLs induced by nanoparticle-antigen was markedly enhanced in comparison with simple use of the antigenic peptides.It is concluded that the antigenic peptide formulated with nanoparticles can markedly enhance and prolong the antigen presentation action of dendritic cells and induce the generation of CTLs which can recognize and kill the target tumor cells in a MHC class I-restricted fashion.