Yuan Xiaolin, Li Weina, Cui Yifen, Zhan Qing, Zhang Chunlei, Yang Zhen, Li Xiaohuan, Li Shengfan, Guan Qinglin, Sun Xiuyan
Affiliated Hospital of Dalian University, Dalian 116001, PR China.
The Second Affiliated Hospital of Heilongjiang University of Traditional Chinese Medicine, Harbin 150001, PR China.
Int Immunopharmacol. 2015 Jul;27(1):171-6. doi: 10.1016/j.intimp.2015.04.056. Epub 2015 May 14.
To determine whether the necrotic tumor cell-stimulated macrophages (NTCSM) could elicit specific immune response.
Mice were immunized with the necrotic H22 tumor cell lysate-stimulated macrophages and the specific immune responses against the same tumor challenge were examined. The morphologic characteristics were observed with the transmission electron microscope and scanning electron microscopy. The expression of CD14, CD68, CD80 and CD86 were detected with the flow cytometer. The cytotoxicity and cytokine production of splenocytes were measured with the MTT assay and ELISA assay respectively.
Our research results reveal that NTCSMs are larger cells which generally generate spherical and elongated protrusions, folding membrane, and vesicles on their surface. Also, abundant lysosomes, secondary lysosomes, phagosomes, rough endoplasmic reticulum, and lipid bodies were found in their cytoplasm. The flow cytometry results show that the necrotic H22 tumor cell lysate could enhance the expression of CD14 and CD86 molecules and the NTCSM was characterized by the expression of CD14+/-CD68+CD80-CD86+. After the mice were vaccinated with NTCSMs, the tumor forming rate, tumor volume and weight of the NTCSM-vaccinated group were significantly lower than those of the sterile saline-injected group and untreated macrophage-vaccinated group (p<0.05). The cytotoxicity to H22 tumor cells of the splenocytes obtained from the NTCSM-immunized group was higher than that of the sterile saline-injected group and untreated macrophage-vaccinated group (p<0.05). Meanwhile, the levels of IL-2 and IFN-γ in the culture supernatant of the NTCSM-immunized group were higher significantly than those of the saline-injected group and untreated macrophage-vaccinated group. The level of IL-4 of the NTCSM-immunized group was significantly lower than those of the other two groups.
Our results indicated that NTCSMs could elicit specific cellular immune responses in vivo.
确定坏死肿瘤细胞刺激的巨噬细胞（NTCSM）是否能引发特异性免疫反应。
用坏死的H22肿瘤细胞裂解物刺激的巨噬细胞免疫小鼠，并检测针对相同肿瘤攻击的特异性免疫反应。用透射电子显微镜和扫描电子显微镜观察形态学特征。用流式细胞仪检测CD14、CD68、CD80和CD86的表达。分别用MTT法和ELISA法检测脾细胞的细胞毒性和细胞因子产生。
我们的研究结果显示，NTCSM是较大的细胞，其表面通常产生球形和细长的突起、折叠膜和囊泡。此外，在其细胞质中发现了丰富的溶酶体、次级溶酶体、吞噬体、粗面内质网和脂体。流式细胞术结果表明，坏死的H22肿瘤细胞裂解物可增强CD14和CD86分子的表达，NTCSM的特征为CD14+/-CD68+CD80-CD86+表达。用NTCSM免疫小鼠后，NTCSM免疫组的肿瘤形成率、肿瘤体积和重量均显著低于无菌盐水注射组和未处理巨噬细胞免疫组（p<0.05）。NTCSM免疫组获得的脾细胞对H22肿瘤细胞的细胞毒性高于无菌盐水注射组和未处理巨噬细胞免疫组（p<0.05）。同时，NTCSM免疫组培养上清中IL-2和IFN-γ水平显著高于盐水注射组和未处理巨噬细胞免疫组。NTCSM免疫组的IL-4水平显著低于其他两组。
我们的结果表明，NTCSM可在体内引发特异性细胞免疫反应。
