CAJ | 학술논문

背景与目的：研究发现核糖体蛋白L8(ribosomal protein L8，RPL8)在黑色素瘤中表达能激活黑色素瘤患者外周血单个核细胞增殖并产生白细胞介素2，提示RPL8可能参与抗肿瘤免疫应答，有望成为抗肿瘤治疗的靶点。本研究通过RPL8蛋白负载树突状细胞(dentritic cell，DC)，探讨负载RPL8蛋白的DC对黑色素瘤的免疫效应。方法：原核表达RPL8蛋白，纯化后致敏小鼠骨髓来源DC，流式细胞仪检测DC表面标志，MTT法检测细胞毒性T淋巴细胞对小鼠黑色素瘤细胞的杀伤作用；负载RPL8蛋白DC免疫治疗小鼠后，观察肿瘤体积变化及小鼠生存时间。结果：纯化蛋白经蛋白质印迹法(Western blot)分析见约28×103大小的特异性条带；DC经RPL8及细菌脂多糖(Lipoplysaccharide，LPS)诱导成熟后细胞表面CD11c、CD80、MHC-Ⅰ类、MHC-Ⅱ类分子表达增高，能激活T淋巴细胞，对B16细胞有抑制作用，RPL8-DC组的抑制率在效靶比为30∶1时高达70%，较PBS组和DC组明显增高；负载RPL8蛋白DC免疫治疗小鼠后，肿瘤体积缩小，小鼠的生存期明显延长。结论：负载RPL8的DC对黑色素瘤有生长抑制作用。
배경여목적：연구발현핵당체단백L8(ribosomal protein L8，RPL8)재흑색소류중표체능격활흑색소류환자외주혈단개핵세포증식병산생백세포개소2，제시RPL8가능삼여항종류면역응답，유망성위항종류치료적파점。본연구통과RPL8단백부재수돌상세포(dentritic cell，DC)，탐토부재RPL8단백적DC대흑색소류적면역효응。방법：원핵표체RPL8단백，순화후치민소서골수래원DC，류식세포의검측DC표면표지，MTT법검측세포독성T림파세포대소서흑색소류세포적살상작용；부재RPL8단백DC면역치료소서후，관찰종류체적변화급소서생존시간。결과：순화단백경단백질인적법(Western blot)분석견약28×103대소적특이성조대；DC경RPL8급세균지다당(Lipoplysaccharide，LPS)유도성숙후세포표면CD11c、CD80、MHC-Ⅰ류、MHC-Ⅱ류분자표체증고，능격활T림파세포，대B16세포유억제작용，RPL8-DC조적억제솔재효파비위30∶1시고체70%，교PBS조화DC조명현증고；부재RPL8단백DC면역치료소서후，종류체적축소，소서적생존기명현연장。결론：부재RPL8적DC대흑색소류유생장억제작용。
Background and purpose:Studies have shown that ribosomal protein L8 (RPL8) is shared by melanomas, gliomas and ovarian carcinomas. A peptide of RPL8 signiifcantly stimulated proliferation and cytokine expression of the hepler T cell clone and lymphocytes in melanoma patients. RPL8 may stimulate anti-tumor immunity, making RPL8 an attractive candidate for therapeutic intervention. In this study, we prepared DC pulsed by RPL8 (RPL8-DC) and investigate the anti-tumor effect of RPL8-DC on melanoma in mice.Methods: The recombinant protein was achieved through IPTG induction in E. coli and identiifed with Western blot. Bone marrow-derived DC was loaded with RPL8 protein. RPL8 and CD11c, CD80, MHC-Ⅰ, MHC-Ⅱmolecules on dentritic cells were monitored by lfuorescence microscope and FACS analysis, respectively. The anti-tumor effect of T cells in vitro was detected by MTT assay. Subcutaneous tumors were induced in C57BL/6 mice using B16 cells. The tumor volumes were measured after injection with RPL8-DC. Results:The puriifed protein was combined with speciifc antibodies. DCs pulsed by RPL8 were visualized under lfuorescent microscopy. CD11c, CD80, MHC-Ⅰ, MHC-Ⅱmolecules on DCs were up-regulated after stimulation with RPL8 and LPS. B16 cells were inhibited by T cells stimulated with RPL8-DC. The inhibition rate of tumor cells was 70%in RPL8-DC group when effector-to-target ratio was 30∶1, which was higher than PBS and DC groups. Inhibition of growth could be observed more signiifcantly in mice after the treatment with RPL8-DC. The mice receiving the therapy of RPL8-DC were able to survive much longer than the mice receiving control therapy. Conclusion:The DC pulsed by RPL8 protein can inhibit the growth of melanoma.