Ag85A与Ag85B DNA疫苗对膀胱肿瘤免疫效应的实验研究

The experiment of immunological effect of Ag85A and Ag85B DNA vaccine on bladder tumor

目的研究单独及联合应用Ag85A与Ag85B DNA疫苗对膀胱肿瘤的免疫效应。方法选择6～8周龄雌性Wistar大鼠作为实验对象。以膀胱灌注MNU建立膀胱原位癌模型，左后肢皮下注射BTF739细胞株建立种植性膀胱肿瘤模型。于末次膀胱灌注后14d随机处死10只大鼠，以确定模型是否成功建立。每种模型均随机分为6组（每组8只）：生理盐水（NS）组、pcDNA3．1组（空白质粒组）、pcDNA3．1-Ag85A（Ag85A）组．pcDNA3．1-Ag85B（Ag85B）组、pcDNA3．1-Ag85A＋Ag85B组（联用组）、卡介苗（BCG）组。并予相应干预治疗。计算各组肿瘤生长抑制率、荷瘤鼠脾脏T细胞亚群数量变化，检测血清干扰素（IFN）-γ水平，剥离肿瘤进行病理组织学检查。结果肿瘤生长抑制率：BCG组最高达63．53％。其余依次为联用组53．56％、Ag85B组33．08％、Ag85A组11．50％、空白质粒组1．10％；干预治疗后空白质粒组T细胞亚群无明显变化，其余各组CD4^＋T细胞与CD8^＋T细胞亚群的数量及CD4^＋／CD8^＋比值均有增加（P〈0．05或P〈0．01）；空白质粒组血清IFN-γ水平无变化，BCG组升高最明显，其余依次为联用组、Ag85B组,Ag85A组。病理组织学观察显示BCG组、联用组、Ag85B组肿瘤坏死明显，而Ag85A组、NS组和空白质粒组肿瘤坏死不明显。结论Ag85A与Ag85B DNA疫苗对大鼠原位及种植性膀胱癌有明显的免疫效应，Ag85BDNA疫苗的免疫效力强于Ag85ADNA疫苗；两者联用效果更显著，但不及BCG的抗肿瘤效果。

Objective To explore the immunological effect of Ag85A, Ag85B and Ag85A ＋ Ag85B DNA vaccine on bladder cancer. Methods Female Wistar rats aged 6-8 weeks were served as experimental animals. Bladder tumor in situ model was established by bladder irrigation of MNU. Implanted bladder tumor model was established by subcutaneously injecting BTT739 cell strain in rear limb. Seleetod randomly 10 rats were killed at fourteenth day after end-irrigation to identify if successful for the models. The Wistar rats of each model group were randomly assigned to 6 subgroups （n = 8 each） ： normal saline group （NS group） ,peDNA3. 1 group （blank plasmid group）, peDNA3. 1-Ag85A greup（Ag85A group）, peDNA3. 1-Ag85B group（Ag85B group）,peDNA3. 1- Ag85A ＋ Ag85B group （combined group） and BCG vaccine group （ BCG group）. The vaccination was performed at seventh, fourteenth and twenty-first day in all experimental rats for observing immunolagical effect. T cell subgroups of spleen tissue were analyzed ; levels of serum IFN--/were deteetod; tumor inhibition rate of tumor-bearing rats was calculated ;histopathologlcal examination of stripped tumor was performed. Results The models of bladder tumor in situ and in implanted cancer were successfully established. The tumor inhibition rate were 63. 53% ,53.56% ,33. 08% ,11.50% and 1.10%, respectively, in BCG group, combined group, Ag85B group,Ag85A group and blank plasmid group. Besides the blank plasmid group, the number of CD4^＋ and CD8^＋ T eell subsets, the ratio of CD4^＋/CD8^＋ and IFN-γ level in caeh group were significantly higher than that in NS group （ P 〈0. 05 or P 〈 0. 01 ）. Pathological observation demonstrated that tumor necrosis was obvious in BCG group, combined group and Ag85B group , but was not in other three groups. The immunological effects of BCG group were the highest in all groups. Conclusions Ag85A and Ag85B DNA vaccine have stronger immunization effects on situ and implanted bladder tumor. Immunization effect of Ag85A DNA vaeeine is lower than Ag85B DNA vaccine, and synergetie effeet after joint application is more predominant, but no matter Ag85A DNA, Ag85B DNA or their joint application could not be compatible with BCG.