中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2011年
29期
2077-2080
,共4页
肝肿瘤%树突细胞%甲胎蛋白类%重组腺相关病毒
肝腫瘤%樹突細胞%甲胎蛋白類%重組腺相關病毒
간종류%수돌세포%갑태단백류%중조선상관병독
Liver neoplasms%Dendritic cells%Alpha-fetoproteins%Recombinant adeno-associated virus
目的 探讨携带甲胎蛋白基因的重组腺相关病毒(rAAV-AFP)转导人树突状细胞(DC)体外诱导抗肝癌免疫应答.方法 分离健康志愿者外周血单核细胞,贴壁细胞转导rAAV-AFP后,在粒细胞巨噬细胞集落刺激因子(GMCSF)和白细胞介素4(IL-4)的联合作用下,诱导分化为DC.用四唑盐(MTS)比色法检测AAV-AFP转导的DC(AAV-AFP+DC)刺激自体T细胞增殖的能力;流式细胞仪检测DC表型、AAV-AFP转导DC的效率以及AAV-AFP+DC激活的T细胞干扰素γ(IFN-γ)和IL-4的分泌;乳酸脱氢酶(LDH)释放细胞毒试验检测AAV-AFP+DC刺激的T细胞对AFP阳性肝癌细胞系的杀伤活性.结果 AAV-AFP转导的DC高表达人类主要组织相容性复合体Ⅰ类分子(HLA Ⅰ,97.12%)、HLAⅡ(97.32%)、CD80(38.94%)、CD83(60.84%)、CD86(98.14%)等DC的典型表面标记;AAV-AFP转导后,81.2%的DC表达AFP;AAV-AFP+DC能够有效地刺激自体T细胞增殖、活化,激活后19.84%的CD4+T细胞和18.65%的CD8+T细胞能够分泌IFN-γ而不分泌IL-4,对AFP阳性的肝癌细胞系HepG2、BEL7402杀伤率分别为56.45%和78.94%.结论 AAV-AFP+DC体外能够诱导出有效地抗AFP阳性肝癌免疫应答,为AFP表达阳性的肝癌患者的主动性免疫治疗提供了实验依据.
目的 探討攜帶甲胎蛋白基因的重組腺相關病毒(rAAV-AFP)轉導人樹突狀細胞(DC)體外誘導抗肝癌免疫應答.方法 分離健康誌願者外週血單覈細胞,貼壁細胞轉導rAAV-AFP後,在粒細胞巨噬細胞集落刺激因子(GMCSF)和白細胞介素4(IL-4)的聯閤作用下,誘導分化為DC.用四唑鹽(MTS)比色法檢測AAV-AFP轉導的DC(AAV-AFP+DC)刺激自體T細胞增殖的能力;流式細胞儀檢測DC錶型、AAV-AFP轉導DC的效率以及AAV-AFP+DC激活的T細胞榦擾素γ(IFN-γ)和IL-4的分泌;乳痠脫氫酶(LDH)釋放細胞毒試驗檢測AAV-AFP+DC刺激的T細胞對AFP暘性肝癌細胞繫的殺傷活性.結果 AAV-AFP轉導的DC高錶達人類主要組織相容性複閤體Ⅰ類分子(HLA Ⅰ,97.12%)、HLAⅡ(97.32%)、CD80(38.94%)、CD83(60.84%)、CD86(98.14%)等DC的典型錶麵標記;AAV-AFP轉導後,81.2%的DC錶達AFP;AAV-AFP+DC能夠有效地刺激自體T細胞增殖、活化,激活後19.84%的CD4+T細胞和18.65%的CD8+T細胞能夠分泌IFN-γ而不分泌IL-4,對AFP暘性的肝癌細胞繫HepG2、BEL7402殺傷率分彆為56.45%和78.94%.結論 AAV-AFP+DC體外能夠誘導齣有效地抗AFP暘性肝癌免疫應答,為AFP錶達暘性的肝癌患者的主動性免疫治療提供瞭實驗依據.
목적 탐토휴대갑태단백기인적중조선상관병독(rAAV-AFP)전도인수돌상세포(DC)체외유도항간암면역응답.방법 분리건강지원자외주혈단핵세포,첩벽세포전도rAAV-AFP후,재립세포거서세포집락자격인자(GMCSF)화백세포개소4(IL-4)적연합작용하,유도분화위DC.용사서염(MTS)비색법검측AAV-AFP전도적DC(AAV-AFP+DC)자격자체T세포증식적능력;류식세포의검측DC표형、AAV-AFP전도DC적효솔이급AAV-AFP+DC격활적T세포간우소γ(IFN-γ)화IL-4적분비;유산탈경매(LDH)석방세포독시험검측AAV-AFP+DC자격적T세포대AFP양성간암세포계적살상활성.결과 AAV-AFP전도적DC고표체인류주요조직상용성복합체Ⅰ류분자(HLA Ⅰ,97.12%)、HLAⅡ(97.32%)、CD80(38.94%)、CD83(60.84%)、CD86(98.14%)등DC적전형표면표기;AAV-AFP전도후,81.2%적DC표체AFP;AAV-AFP+DC능구유효지자격자체T세포증식、활화,격활후19.84%적CD4+T세포화18.65%적CD8+T세포능구분비IFN-γ이불분비IL-4,대AFP양성적간암세포계HepG2、BEL7402살상솔분별위56.45%화78.94%.결론 AAV-AFP+DC체외능구유도출유효지항AFP양성간암면역응답,위AFP표체양성적간암환자적주동성면역치료제공료실험의거.
Objective To investigate the generation of antitumor response against hepatocellular carcinoma by in vitro transduction of dendritic cells (DC)with recombinant adeno-associated virus expressing α-fetoprotein (rAAV-AFP). Methods Peripheral blood mononuclear cells were isolated from healthy volunteers. Adherent peripheral blood mononuclear cells were transduced with AAV-AFP and cultured in the presence of granulocyte macrophage colony stimulating factor and interleukin-4 to generate dendritic cells.MTS assay was used to measure the ability of DC transduced with AAV-AFP ( AAV-AFP + DC) to stimulate the proliferation of T cell. The phenotype and AFP protein expression of DC and the secretion of IFN (interferon)-γ and IL (interleukin)-4 by T cells were detected by flow cytometry. The killing efficacy of cytotoxic T lymphocytes (CTL) activated by AAV-AFP + DC against AFP positive hepatocellular carcinoma cell lines was detected by lactate dehydrogenase (LDH) release assay. Results AAV-AFP + DC expressed HLA Ⅰ (97. 12%), HLAⅡ (97.32%), CD80(38.94%), CD83(60.84%)and CD86(98. 14%). AFP was secreted by 81.2% of AAV-AFP + DC. And it could stimulate effectively the proliferation of T cell.19. 84% of CD4 + T cells and 18.65% of CD8 + T cells activated by AAV-AFP + DC produced IFN-γbut not IL-4 and showed distinct killing activities against AFP positive hepatocellular carcinoma cell lines HepG2 (56. 45% ) and BEL7402 (78. 84% ). Conclusion AAV-AFP + DC can elicit distinct antitumor responses against AFP positive hepatocellular carcinoma cell lines so as to provide a basis for further researches on the clinical application of AAV-AFP + DC in the treatment of hepatocellular carcinoma.
