Liu Xia, Zhang Leilei, Zhang Xingqian, Yu Hai, Zhao Xiaoping, Lu Jian, Qian Guanxiang, Ge Shengfang
Research Center for Human Gene Therapy, Department of Biochemistry and Molecular Biology, School of Medicine, Shanghai Jiao Tong University, Shanghai, PR China.
Int J Oncol. 2007 Jun;30(6):1389-96.
CD80, a costimulatory molecule, plays an important role in eliciting antitumor immunity. Without costimulation, recognition of antigens by T cells may not cause a response, even if tumor cells express MHC class I molecules and specific antigens. On the basis of the recombinant GFP-tagged Kb molecule, we constructed a co-expression vector of CD80 and GFP-tagged Kb molecules. The recombinant fusion was transfected into mouse melanoma B16 cells by electroporation; positive cells were obtained by G418 screening. Highly expressing monoclonal cells, irradiated by 137Cs, were used to immunize mice to obtain specific T cells, which were then cultivated with tumor cells in vitro and examined with a laser confocal microscope. The evident and intense uptake of the antigen peptide-MHC class I-GFP complex by specific T cells was visualized from the culture of B16/CD80-Kb-GFP and T cells. However, little uptake was observed from the culture of B16/Kb-GFP and T cells. These results show that co-expression of CD80 molecules with Kb, an MHC class I molecule, on the surface of B16 tumor cells can enhance the response of specific T cells and thus increase the uptake of the antigen peptide-MHC class I-GFP complex. The absorbed green fluorescence was concentrated mainly on the T cell surface, and this result might pave the way to eluting specific antigen peptides directly from T cells to find and isolate novel tumor-specific antigen peptides.
共刺激分子CD80在引发抗肿瘤免疫中发挥重要作用。没有共刺激的情况下，即使肿瘤细胞表达MHC I类分子和特异性抗原，T细胞对抗原的识别也可能不会引发反应。基于重组绿色荧光蛋白（GFP）标记的Kb分子，我们构建了CD80与GFP标记的Kb分子的共表达载体。通过电穿孔将重组融合体转染到小鼠黑色素瘤B16细胞中；通过G418筛选获得阳性细胞。用137Cs照射高表达单克隆细胞以免疫小鼠获得特异性T细胞，然后将其与肿瘤细胞在体外培养并用激光共聚焦显微镜检查。从B16/CD80-Kb-GFP和T细胞的培养物中可以看到特异性T细胞对抗原肽-MHC I类-GFP复合物有明显且强烈的摄取。然而，从B16/Kb-GFP和T细胞的培养物中观察到的摄取很少。这些结果表明，在B16肿瘤细胞表面将CD80分子与MHC I类分子Kb共表达可以增强特异性T细胞的反应，从而增加对抗原肽-MHC I类-GFP复合物的摄取。吸收的绿色荧光主要集中在T细胞表面，这一结果可能为直接从T细胞洗脱特异性抗原肽以寻找和分离新型肿瘤特异性抗原肽铺平道路。
