巨噬细胞肿瘤疫苗抗瘤作用的实验研究

Experimental study on the anti-tumor effects of the macrophage-tumor vaccine

目的:探究巨噬细胞肿瘤疫苗的细胞形态与表型特征,并观察其CTL反应的诱导效果。方法:分别采用透射与扫描电子显微镜、免疫荧光染色及流式细胞术等技术对巨噬细胞肿瘤疫苗的超微结构及CD14、CD68、CD80、CD86、MHCⅡ等分子进行测定。采用生长状态良好的H22肿瘤细胞移植于接种不同肿瘤疫苗的实验小鼠,分别采用直接测量法、MTT法及比色法测定瘤重量、瘤体积、肿瘤细胞杀伤率和培养上清液LDH活性。结果:巨噬细胞肿瘤疫苗细胞表面有许多的伪足皱褶、囊泡,胞浆内有大量大小不一、形态不规则的吞噬体;CD14、CD68、CD80、CD86及MHCⅡ的阳性细胞率分别为53.90%、98.60%、26.50%、90.20%和25.40%。小鼠体内实验结果显示,巨噬细胞肿瘤疫苗接种组的肿瘤形成率、瘤体积与瘤重量明显低于对照组和石蜡诱生的巨噬细胞接种组(P均<0.05);巨噬细胞肿瘤疫苗接种组的成瘤率与灭活肿瘤细胞接种组无明显差异(P>0.05),但瘤体积与瘤重量明显低于灭活肿瘤细胞接种组(P均<0.05)。另外,巨噬细胞肿瘤疫苗接种组的淋巴细胞自体肿瘤细胞杀伤率和培养上清液LDH活性分别高于对照组、灭活肿瘤细胞接种组和石蜡诱生的巨噬细胞接种组。结论:巨噬细胞肿瘤疫苗具备巨噬细胞的典型特征,该种细胞接种后可诱导机体产生特异性抗肿瘤免疫反应。

Objective： To study the morphologic and phenotypic characters of a macrophage-tumor vaccine,and to observe the effect of macmphage-tumor vaccine on inducing CTL respose. Methods： The super-structure and the expression of CD14, CD68, CD80, CD86, MHC Ⅱ molecules of macmphage-tumor cells were detected with electron microscope, immunofluorescence staining and flow cytometry respectively. Meanwhile, H22 tumor cells were transplanted to the mice that had been intmunized with different tumor vaccines. The weight and volume of tumors, the tumor cell injure rate and the level of LDH in culture supernatant were detected with direct measurement, MTT and selection methods. Results：The macrophage tumor vaccine cells were large cells with an irregular outline, and generally displayed pseudopodium, membrane folding, and vesicles on the cell surface. The predominant cytoplasmic organelles were lysosomes, secondary lysosomes and residual bodies. The percentage of CD14, CD68, CDS0, CD86 and MHC Ⅱ positive cells within the differentiated population were 53.90%, 98.60%, 26.50%, 90.20 % and 25.40 % respectively. The results of experiment in vivo revealed that the tumor forming rate, volume and weight of the group immunized with macrophage-tumor vaccine were much lower than that of control group and the group that were immunized with the macrophages that were induced by liquid paratfin（P〈 0.05）. Although there was no obvious difference in tumor forming rate between the group immunized with the macrophage-tumor vaccine and the group immunized with inactivated tumor cells（ P 〉 0.05）, the tumor weight and volume of the group immunized with the macmphage-tumor vaccine were lower than those of the group immunized with inactivated tumor cells（ P 〈 0.05）. In addition, the auto-tumor injury rate and the level of LDH in culture supematant of the group immunized with macrophages were higher than those of the other three groups. Conclusion： These results indicate that the macrophage-tumor vaccine possess classical characteristics of macrophages and this tumor vaccine could promote specific anti-tumor immunol responds.