负载人AFP肽段的树突细胞在体内外对小鼠肝癌的抑制作用

Inhibitory Effects of Human AFP-derived Peptide-pulsed Dendritic Cells on Mouse Hepatocellular Carcinoma

背景与目的:甲胎蛋白(alpha-fetoprotein,AFP)是原发性肝细胞癌(hepatocellular carcinoma,HCC)免疫治疗的一个良好的靶分子,如何克服对自身抗原的免疫耐受状态是诱导有效抗肿瘤免疫反应的关键。本研究探讨异种同源蛋白疫苗负载人AFP肽段的树突细胞(human AFP-derived peptide-pulse ddendritic cells,hAFP-DCs)对小鼠肝癌的体外杀伤作用和体内抑瘤效应。方法:传统方法制备骨髓来源的DCs。MTT法检测hAFP-DCs诱导的CTL对小鼠肝癌细胞Hepa1-6的体外杀伤活性。建立Hepa1-6细胞C57BL/6小鼠移植瘤模型,分别瘤内注射hAFP-DCs、DCs和PBS(每周两次),观察小鼠肿瘤体积和荷瘤存活时间。结果:成功制备小鼠骨髓来源的DCs。体外杀伤实验显示,hAFP-DCs刺激组和单纯DCs刺激组CTL对Hepa1-6细胞的杀伤作用强于PBS组,但组间差异无统计学意义(P>0.05)。体内实验表明,每个C57BL/6小鼠接种7×106个Hepa1-6细胞31d后,hAFP-DCs﹑DCs和PBS组小鼠平均移植瘤体积分别为(195.04±155.22)mm3﹑(360.65±209.02)mm3和(756.19±503.24)mm3,组间比较差异有显著性(P<0.001)。在40d的观察期内,小鼠的累积存活率分别为100%、90%和50%(P=0.008)。结论:负载人AFP抗原肽的DCs疫苗在体外和体内均能有效抑制小鼠肝癌的生长。

BACKGROUND ＆ OBJECTIVE= Alpha-fetoprotein （AFP） is a good candidate antigen for the immunotherapy of hepatocellular carcinoma （HCC）. How to overcome the immune tolerance induced by autologous antigen is one of key points for inducing effective antitumor immune reaction. This research was to invesigate the effect of human AFP-derived peptidepulsed dendritic cells （hAFP-DCs） on immunity against mouse HCC. METHODS： Bone marrow-originated DCs were prepared routinely. The activity of hAFP-DC-stimulated cytotoxic T lymphocyte （CTL） against Hepal-6 cells was examined by MIT assay. C57BL/6 mice were inoculated subcutaneously with 7×10^6 Hepal-6 cells to develop hepatoma, and received intratumor injection of hAFP-DCs, DCs and PBS, respectively, twice a week. Tumor volume was evaluated and the survival of mice after inoculation was observed. RESULTS： We successfully prepared DCs from bone marrow of mice. The cytotoxic activity of CTLs stimulated by hAFP-DCs and DCs showed stronger tendency than that of control, but without significance. The mean tumor volume at 31 days after inoculation with Hepal-6 cells was （195,04± 155.22） mm^3 in hAFP-DCs group, （360.65±209.02） mm^3 in DCs group and （756.19±503.24） mma in PBS group. The differences among these three groups were significant （P〈0.001）. The survival rate of mice at 40 days after inoculation was 100% in hAFP-DCs group, 90% in DCs group and 50% in PBS group （P=0.008）. CONCLUSION. Human AFP-derived peptide-pulsed DCs can efficiently enhance immunity against HCC in mice.