CAJ | 학술논문

目的：研究 S jcb2 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（＋）/S jcb2核酸疫苗，将6周龄雌性BALB/c小鼠随机分为pcDNA3．1（＋）/S jcb2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 100μg ，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 S jcb2基因在小鼠体内的稳定性及表达情况；M T T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 S jcb2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 S jcb2 DNA疫苗接种小鼠后能在小鼠肌细胞中稳定存在和表达；S jcb2可能通过提高IFN-γ和降低IL-4水平调节 T h1细胞亚群产生抗血吸虫感染的保护性作用。
목적：연구 S jcb2 DNA역묘재일본혈흡충병소서모형중적보호성작용화궤제，위혈흡충역묘적연구제공유효적후선항원분자。방법구건pcDNA3．1（＋）/S jcb2핵산역묘，장6주령자성BALB/c소서수궤분위pcDNA3．1（＋）/S jcb2핵산역묘조、pcDNA3．1（＋）공질립조급생리염수조，매조35지，채용후퇴고사두기주사방법，매차면역질립DNA 100μg ，매2주면역1차，공면역3차，용일본혈흡충미유공격감염각조소서。PCR급면역조화법검측 S jcb2기인재소서체내적은정성급표체정황；M T T법검측소서비림파세포특이성증식반응；ELISA법검측소서혈청중 S jcb2항체수평급공격감염전후비림파세포배양상청중IFN-γ화IL-4적수평；계수소서하성충대수화간장하충란수。결과역묘조소서균가재소서기세포중검측도 Sjcb2기인급기항원적표체；DNA역묘조T세포증식현저증고（P ＜0．05）；ELISA 결과현시역묘조IFN-γ수평현저증고（P ＜0．05），혈흡충미유공격감염후각조소서IL-4수평현저승고（P ＜0．05）。DNA역묘조소서하성충대수급간장하충란수여기타조비교현저성감소（P ＜0．05），기감충솔위36．32％，감란솔위60．61％。결론 S jcb2 DNA역묘접충소서후능재소서기세포중은정존재화표체；S jcb2가능통과제고IFN-γ화강저IL-4수평조절 T h1세포아군산생항혈흡충감염적보호성작용。
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 S jcb2 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-γand 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 in-creased significantly in S jcb2 DNA vaccinated group .ELISA results showed that the IFN-γlevels 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 S jcb2 recombinant plasmid ,and S jcb2 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 .