人食管癌Eca-109细胞M_r小于3000膜蛋白肿瘤抗原肽的筛选  

Screening of tumor antigenic peptides in membrane antigen with M_r less than 3000 of human esophageal carcinoma Eca-109 cell line

作　　者：李付广[1] 金戈[2] 董子明[3] 

机构地区：[1]郑州大学基础医学院微生物学与免疫学教研室,郑州450052 [2]郑州大学基础医学院生物化学教研室,郑州450052 [3]郑州大学基础医学院病理生理学教研室,郑州450052

出　　处：《郑州大学学报（医学版）》2006年第4期614-616,共3页Journal of Zhengzhou University(Medical Sciences)

基　　金：河南省科技攻关基金资助项目324410035

摘　　要：目的:寻找被T细胞识别的食管癌细胞抗原肽,为食管癌免疫治疗奠定基础。方法:应用pH3.3的枸橼酸磷酸盐缓冲液间隔24h多次酸洗贴壁生长的Eca109细胞,获得人主要组织相容性复合体Ⅰ类分子(HLAⅠ)类分子结合的多肽,经超过滤,反相HPLC色谱层析得到不同组分多肽,DC递呈抗原肽刺激特异性细胞毒T淋巴细胞(CTL)反应,51Cr杀伤实验检测CTL对肿瘤细胞杀伤作用。结果:Mr小于3000的混合多肽经过反相HPLC可获得50多个组分,混合肽、P17、P29和P41组分体外诱导实验表明可以诱导出较强的CTL反应。结论:酸洗法能有效获得HLAⅠ类分子结合的多肽抗原,Mr小于3000的多肽成分中3个组分存在能引起特异性的抗肿瘤CTL免疫反应的肿瘤抗原,具有潜在的免疫治疗作用。Aim： To seek the antigenic peptides from the esophageal carcinoma cell, which could be identified by T lymphocytes and lie a foundation of immune theraphy for esophageal carcinoma. Methods： Eca-109 cell line was treated by citrate-phosphate buffer at pH 3.3, to denature HLA class Ⅰ molecules and release HLA classⅠ molecules-associated peptides into the supernatant. The acid solution containing peptides was desahed, vacuum concentrated, and centrifuged on Microcon YM-3. Peptides were subsequently fractioned by reverse-phase high performance liquid chromatography （ RP- HPLC）. PBMCs were stimulated by peptide-pulsed dendritic cells （DCs）. The specific cytotoxicity against the Eca-109 cell line was determined by ^51Cr release assay. Results： RP-HPLC analysis showed that about fifty different fractions were obtained from Mr 〈 3 000 HLA- Ⅰ molecules-associated peptides. P17, P29 and P41 fractions excited cytotoxic activity of CTLs in vitro. Conclusion： Mild acid wash method can efficiently elute the HLA-Ⅰ molecules-associated peptides from Eca- 109 cell line. There are three fractions （P17, P29, P41 ） that can stimulate specific CTLs reaction against Eca-109 cell line.

关 键 词：食管癌 肿瘤抗原 HLA 抗原肽 树突状细胞 

分 类 号：R730.51[医药卫生—肿瘤]