中华微生物学和免疫学杂志
中華微生物學和免疫學雜誌
중화미생물학화면역학잡지
CHINESE JOURNAL OF MICROBIOLOGY AND IMMUNOLOGY
2012年
8期
696-701
,共6页
沈晗%黄昭亮%陈晓华%林燕梅%王腾%邵红伟%黄树林
瀋晗%黃昭亮%陳曉華%林燕梅%王騰%邵紅偉%黃樹林
침함%황소량%진효화%림연매%왕등%소홍위%황수림
Survivin%抗肝癌%点突变肽%HLA-A2限制性
Survivin%抗肝癌%點突變肽%HLA-A2限製性
Survivin%항간암%점돌변태%HLA-A2한제성
Survivin%Anti-hepatocarcinoma%Point-mutated peptide%HLA-A2-restricted
目的 探讨HLA-A2限制性Survivin点突变抗原肽体外诱导CTLs的抗肝癌作用.方法 生物信息学软件BIMAS和SYFPEITHI用来鉴定点突变的HLA-A2限制性Survivin抗原九肽；流式细胞术检测突变肽与HLA-A2分子结合效率；肽体外刺激肝癌患者腹水来源的肿瘤浸润淋巴细胞(TILs)诱导反应性CTLs,用流式细胞术及ELISA检测反应性CTLs释放IFN-γ情况；肽诱导的CTLs与肝癌细胞系HepG2及BEL-7402共孵育,CytoTox 96(R)非放射性细胞毒性检测法检测对肿瘤细胞的裂解情况,倒置相差显微镜观察肝癌细胞形态学变化.结果 生物信息学分析筛选出与MHC分子结合效率评分显著提高的点突变Survivin抗原九肽Sur79M2 (KMSSGCAFL),流式细胞术检测证实负载Sur79M2的T2细胞,HLA-A2表达率显著提高,经Sur79M2体外刺激诱导的CTLs与靶细胞共孵育后能释放较高水平的IFN-γ,Sur79M2诱导的CTLs能通过HLA-A2限制性机制有效杀伤肝癌细胞系HepG2,对HLA-A2阴性的BEL-7402细胞无明显杀伤作用.结论 点突变肽Sur79M2能在体外诱导反应性CTLs产生,该CTLs能以HLA-A2限制性方式有效杀伤肝癌细胞系.
目的 探討HLA-A2限製性Survivin點突變抗原肽體外誘導CTLs的抗肝癌作用.方法 生物信息學軟件BIMAS和SYFPEITHI用來鑒定點突變的HLA-A2限製性Survivin抗原九肽；流式細胞術檢測突變肽與HLA-A2分子結閤效率；肽體外刺激肝癌患者腹水來源的腫瘤浸潤淋巴細胞(TILs)誘導反應性CTLs,用流式細胞術及ELISA檢測反應性CTLs釋放IFN-γ情況；肽誘導的CTLs與肝癌細胞繫HepG2及BEL-7402共孵育,CytoTox 96(R)非放射性細胞毒性檢測法檢測對腫瘤細胞的裂解情況,倒置相差顯微鏡觀察肝癌細胞形態學變化.結果 生物信息學分析篩選齣與MHC分子結閤效率評分顯著提高的點突變Survivin抗原九肽Sur79M2 (KMSSGCAFL),流式細胞術檢測證實負載Sur79M2的T2細胞,HLA-A2錶達率顯著提高,經Sur79M2體外刺激誘導的CTLs與靶細胞共孵育後能釋放較高水平的IFN-γ,Sur79M2誘導的CTLs能通過HLA-A2限製性機製有效殺傷肝癌細胞繫HepG2,對HLA-A2陰性的BEL-7402細胞無明顯殺傷作用.結論 點突變肽Sur79M2能在體外誘導反應性CTLs產生,該CTLs能以HLA-A2限製性方式有效殺傷肝癌細胞繫.
목적 탐토HLA-A2한제성Survivin점돌변항원태체외유도CTLs적항간암작용.방법 생물신식학연건BIMAS화SYFPEITHI용래감정점돌변적HLA-A2한제성Survivin항원구태；류식세포술검측돌변태여HLA-A2분자결합효솔；태체외자격간암환자복수래원적종류침윤림파세포(TILs)유도반응성CTLs,용류식세포술급ELISA검측반응성CTLs석방IFN-γ정황；태유도적CTLs여간암세포계HepG2급BEL-7402공부육,CytoTox 96(R)비방사성세포독성검측법검측대종류세포적렬해정황,도치상차현미경관찰간암세포형태학변화.결과 생물신식학분석사선출여MHC분자결합효솔평분현저제고적점돌변Survivin항원구태Sur79M2 (KMSSGCAFL),류식세포술검측증실부재Sur79M2적T2세포,HLA-A2표체솔현저제고,경Sur79M2체외자격유도적CTLs여파세포공부육후능석방교고수평적IFN-γ,Sur79M2유도적CTLs능통과HLA-A2한제성궤제유효살상간암세포계HepG2,대HLA-A2음성적BEL-7402세포무명현살상작용.결론 점돌변태Sur79M2능재체외유도반응성CTLs산생,해CTLs능이HLA-A2한제성방식유효살상간암세포계.
Objective To investigate the anti-hepatocarcinoma(anti-HCC) function of HLA-A2-restricted point-mutated Survivin peptide induced CTLs.Methods The HLA-A2-restricted Survivin nonapeptides were evaluated using bioinformatics software.The binding affinity of Survivin peptide to HLA-A2 molecular was determined with flow cytometry analysis.After peptide-induced CTLs were generated in vitro,flow cytometry and ELISA were performed to detect the levels of IFN-γ,which were secreted by reactive CTLs.Peptide-induced CTLs were co-cultured with hepatoma cell lines HepG2 and BEL-7402.The rates of tumor cells lysis were assayed using CytoTox 96(R) and the morphological changes of tumor cells were observed with inverted phase contrast microscope.Results Point-mutated Survivin nonapeptide Sur79M2 (KMSSGCAFL) was filtered out,which was shown higher scores compared with the wild-type peptide Sur79.Consistent with the results of software analysis,Sur79M2 showed higher binding ability in T2 binding assays.At the same time,Sur79M2-induced CTLs could release a large number of IFN-γ after incubated with target cells rather than Sur79.When co-cultured with HCC cell lines HepG2 and BEL-7402,Sur79M2-induced CTLs effectively lysis HepG2 on HLA-A2-restricted manner without killing effect on BEL-7402 that do not express HLA-A2 molecules.Conclusion Sur79M2 could elicited specific cytotoxic T lymphocytes in vitro,which were able to specifically kill HCC cell lines on HLA-A2-restricted manner.
