CAJ | 학술논문

目的探讨复制缺损型腺病毒载体(Ad)介导人血管内皮细胞生长因子受体-2(hKDR)诱导抗小鼠肝癌血管免疫、打破免疫耐受的效果. 方法用RT-PCR方法从人脐静脉内皮细胞和小鼠胚胎细胞中分别克隆hKDR和小鼠KDR(mKDR),构建Ad hKDR和Ad mKDR.用Ad hKDR和Ad mKDR分别皮内免疫C57BL/6小鼠,7 d后取脾细胞作为效应细胞,Hepa 1-6/mKDR作为靶细胞,行乳酸脱氢酶释放试验,检测特异性细胞毒性T淋巴细胞(CTL)杀伤率;免疫小鼠接种Hepa 1-6肝癌细胞,观察荷瘤小鼠成活情况. 结果 Ad hKDR和Ad mKDR分别免疫小鼠1周后,在效应细胞:靶细胞为100:1、50:1和25:1时,Ad hKDR诱导的6 h CTL杀伤率分别为84.3%±6.7%、71.5%±5.2%和44.6%±4.7%;Ad mKDR诱导的6 h CTL杀伤率分别为65.2%±6.1%、46.7%±5.0%和22.6%±3.7%.Ad hKDR免疫小鼠后l周接种5×106个Hepal-6肝癌细胞,2个月后仍然有60%的小鼠无瘤生长;而接种2×106个Hepa 1-6细胞至Ad mKDR免疫小鼠,2个月后小鼠成活率为40%.上述CTL效应和肿瘤保护作用在清除CD8+和CD4+T淋巴细胞后消失. 结论 Ad介导异种KDR能有效地打破肿瘤的免疫耐受,诱发强烈的抗原特异性细胞免疫反应,这种特异性细胞免疫反应是CD8+和CD4+T淋巴细胞依赖性的.
목적 탐토복제결손형선병독재체(Ad)개도인혈관내피세포생장인자수체-2(hKDR)유도항소서간암혈관면역、타파면역내수적효과. 방법 용RT-PCR방법종인제정맥내피세포화소서배태세포중분별극륭hKDR화소서KDR(mKDR),구건Ad hKDR화Ad mKDR.용Ad hKDR화Ad mKDR분별피내면역C57BL/6소서,7 d후취비세포작위효응세포,Hepa 1-6/mKDR작위파세포,행유산탈경매석방시험,검측특이성세포독성T림파세포(CTL)살상솔;면역소서접충Hepa 1-6간암세포,관찰하류소서성활정황. 결과 Ad hKDR화Ad mKDR분별면역소서1주후,재효응세포:파세포위100:1、50:1화25:1시,Ad hKDR유도적6 h CTL살상솔분별위84.3%±6.7%、71.5%±5.2%화44.6%±4.7%;Ad mKDR유도적6 h CTL살상솔분별위65.2%±6.1%、46.7%±5.0%화22.6%±3.7%.Ad hKDR면역소서후l주접충5×106개Hepal-6간암세포,2개월후잉연유60%적소서무류생장;이접충2×106개Hepa 1-6세포지Ad mKDR면역소서,2개월후소서성활솔위40%.상술CTL효응화종류보호작용재청제CD8+화CD4+T림파세포후소실. 결론 Ad개도이충KDR능유효지타파종류적면역내수,유발강렬적항원특이성세포면역반응,저충특이성세포면역반응시CD8+화CD4+T림파세포의뢰성적.
Objective To investigate the effect of adenovirus vector encoding human vascular endothelial growth factor receptor-2(hVEGFR-2 or hKDR)on breaking the immune tolerance and inducing immunity against murine hepatocellular carcinomas.Methods Human and mouse KDR cDNA were cloned from human umbilical vein endothelial cells(HUVEC)and C57BL/6 mouse embryo cells respectively using RT-PCR,and then Ad hKDR and Ad mKDR were constructed.Seven days after immunization of the mice with Ad hKDR or Ad mKDR,an analysis of cytotoxic activity of antigen-specific cytotoxic T lymphocytes (CTL)was made by lactate dehydrogenase(LDH)release assay,in which splenocytes of the immunized mice acted as effectors and Hepa 1-6/mKDR cells as the targets.In addition,the survival of the mice immunized with Hepa 1-6 hepatoma cells was checked.Results Seven days after immunization,the 6 h killing activities of CTL elicited by the Ad hKDR were 84.3%±6.7%,71.5%±5.2%,and 44.6%±4.7%at the ratio of the effectors:targets(E:T)of100:1,50:1,and25:1,respectively.Correspondingly,the CTL activities by Ad mKDR were 65.2%±6.1%,46.7%±5.0%,and 22.6%±3.7%.Sixty percent of the Ad hKDR-immunized mice with 5×106Hepa 1-6 hepatoma cells were still alive two months after the inoculation,whereas just 40%of the Ad mKDR-immunized mice with 2×106Hepa 1-6 cells survived two months.When CD8+or CD4+T lymphocytes were deleted in the mice the above mentioned CTL activities and protection of the mice from tumors disappeared.Conclusion Adenovirus vector-mediated xenogeneic KDR can effectively break the immune tolerance to hepatocellular carcinomas in an animal model and induce a strong antigen-specific T cell response.which is dependent on CD8+and CD4+T cells.