Sjcb2 DNA疫苗在血吸虫感染小鼠中的保护性作用研究

目的 研究Sjcb2 DNA疫苗在日本血吸虫病小鼠模型中的保护性作用和机制,为血吸虫疫苗的研究提供有效的候选抗原分子。方法 构建pcDNA3.1(+)/Sjcb2 核酸疫苗,将6周龄雌性BALB/c小鼠随机分为pcDNA3.1(+)/Sjcb2 核酸疫苗组、pcDNA3.1(+)空质粒组及生理盐水组,每组35只,采用后腿股四头肌注射方法,每次免疫质粒DNA 100 μg,每2周免疫1次,共免疫3次,用日本血吸虫尾蚴攻击感染各组小鼠。PCR及免疫组化法检测Sjcb2基因在小鼠体内的稳定性及表达情况;MTT法检测小鼠脾淋巴细胞特异性增殖反应;ELISA法检测小鼠血清中Sjcb2抗体水平及攻击感染前后脾淋巴细胞培养上清中IFN-γ和IL-4的水平;计数小鼠荷成虫对数和肝脏荷虫卵数。结果 疫苗组小鼠均可在小鼠肌细胞中检测到Sjcb2基因及其抗原的表达;DNA疫苗组T细胞增殖显著增高(P <0.05);ELISA 结果显示疫苗组IFN-γ 水平显著增高(P <0.05),血吸虫尾蚴攻击感染后各组小鼠IL-4水平显著升高(P <0.05)。DNA疫苗组小鼠荷成虫对数及肝脏荷虫卵数与其它组比较显著性减少(P <0.05),其减虫率为36.32％,减卵率为60.61％。结论 Sjcb2 DNA疫苗接种小鼠后能在小鼠肌细胞中稳定存在和表达;Sjcb2可能通过提高IFN-γ 和降低IL-4水平调节Th1细胞亚群产生抗血吸虫感染的保护性作用。

Protective function of Sjcb2 DNA vaccine in mice infected with Schistosoma japonicum

The protective effect and mechanism of Schistosoma japonicum cathepsin B (Sjcb2) DNA vaccine in the mouse model of schistosomiasis were studied through construction pcDNA3.1 (+) / Sjcb2 DNA recombinant vector, which provided effective candidate antigen for anti-schistosome vaccine. The 6-week-old female BALB/c mice were randomly divided into pcDNA3.1(+)/Sjcb2 DNA vaccine group, pcDNA3.1(+) plasmid group and normal saline group, respectively. Each group was composed of 35 mice, and 100 μg of Sjcb2 plasmid DNA was injected in the hind leg quadriceps of mice once every two weeks. PCR and immunohistochemistry assay were used to detect the expression and stability of Sjcb2 gene in mice. MTT assay was used for testing the specific proliferation response of mice spleen lymphocytes. The level of Sjcb2 antibodies in mouse serum and the IFN-γ and IL-4 levels in mice spleen lymphocyte culture supernatant before and after schistosome infection were assayed by ELISA. At last, we counted load of Schistosome adult worms in mouse and eggs in liver of mouse. The results showed that the Sjcb2 gene was detected in all mice of the Sjcb2 DNA vaccine group, and Sjcb2 gene expression was positive in the muscle cells in Sjcb2 DNA immunized mice by IHC assay. MTT assay showed that T-cell proliferation rate was increased significantly in Sjcb2 DNA vaccinated group. ELISA results showed that the IFN-γ levels were increased significantly in the vaccinated group, while the IL-4 levels were significantly increased after Schistosoma japonicum infection in all mice of every group. The load of worms and eggs in Sjcb2 DNA vaccinated group was reduced significantly than that of control group (P<0.05), the reduction rates of adult worms and eggs were 36.32% and 60.61% respectively. In conclusion, the Sjcb2 gene was stably expressed in muscle cells of mice after injection of Sjcb2 recombinant plasmid, and Sjcb2 produced protective effects of anti-schistosoma infection in mice possibly by mean of regulating Th1 cell subgroups through increasing the IFN-γ level and decreasing IL-4 levels.