熊果酸增强小鼠肝癌细胞疫苗免疫原性的作用

Immunoadjuvant Effect of Ursolic Acid for Murine H22 Hepatocarcinoma Cell Vaccine

目的:探讨熊果酸(UA)作为免疫佐剂对小鼠肝癌肿瘤疫苗免疫原性的增强作用。方法:UA和H22细胞共培养,制备肝癌细胞疫苗,免疫小鼠后建立肝癌模型,观察肿瘤生长曲线,记录小鼠存活情况,MTT法测定脾淋巴细胞增殖能力,ELISA法测定血清IL-2、IL-4细胞因子含量,流式细胞术测定CD4+、CD8+T细胞亚群百分率,免疫荧光法测定血清中抗肿瘤特异性抗体的产生,ELISA法测定特异性抗体的水平,Western blot检测血清特异性抗体结合能力。结果:免疫小鼠后,与模型组相比,荷瘤小鼠肿瘤的生长明显受到抑制(P<0.05),小鼠的生命延长率明显高于模型组(P<0.01);与模型组相比,T淋巴细胞和B淋巴细胞增殖能力显著升高(P<0.01),CD4+/CD8+T细胞含量比例明显升高(P<0.01),血清中细胞因子IL-2、IL-4含量明显升高(P<0.01)。免疫鼠血清可检测到较高水平特异性抗体,抗体亚型IgG2a/IgG1比值明显升高(P<0.01);Western blot出现4条大小约为95kDa、60kDa、50kDa和34kDa杂交条带。结论:UA作为佐剂能够提高小鼠H22肝癌细胞疫苗的免疫原性,增强小鼠抗肿瘤细胞免疫和体液免疫能力。

To study the immunoadjuvant effect of ursolic acid （UA） co-immunized with murine H22 hepatocarcinoma cell vaccine. The mice were injected lysate of H22 hepatocarcinoma cell and UA co-culture as vaccine three times. After one week of the last immunization the H22 cell model were set up. The tumor growth curve and survival rate of the mice were observed. The proliferations of T lymphocytes were measured by MTT and the levels of serum cytokines IL-2 and IL-4 were determined by ELISA. CD4＋, CD8＋ lymphocyte populations were determined by flow cytometry. The serum antitumor specific antibody were determined by immunofluorescence, ELISA and Western blot. It showed that after the mice were immunized with the tumor vaccines treated with UA as immunoadjuvant. Compared with model group, the tumor growth was inhibited obviously （P〈0.05）, the life span of vaccine group was significantly prolonged （P〈0.01）. The T and B lymphocyte proliferations were significantly promoted （P〈0.01）, the ratio of CD4＋/CD8＋ was promoted obviously （P〈0.01） and the levels of serum cytokines IL-2 and IL-4 significantly higher （P〈0.01）.The specific binding of antiserum and antigen was found and the higher levels of serum antibody was detected. It is concluded that UA as adjuvant could remarkably improves the immunogenicity of H22 tumor vaccines and the mice cellular immunity and humoral immunity could enhance on antitumor.