结核病Mtb8.4/hIL-12嵌合基因疫苗免疫保护效果研究  

Study on the protective effecacy of Mtb8.4/hIL-12 chimeric gene vaccine

作　　者：李晖[1] 李榕 钟森[3] 李强[1] 任红[4] 黄永茂[1] 陈宣世[5] 龙汉安[5] 

机构地区：[1]泸州医学院附属医院感染病科 [2]江西省南昌市血站,南昌330008 [3]成都中医药大学附属医院,成都641000 [4]重庆医科大学病毒性肝炎研究所,重庆400010 [5]泸州医学院病理学教研室,四川泸州646000

出　　处：《免疫学杂志》2007年第2期139-143,共5页Immunological Journal

基　　金：四川省青年科技基金资助([2002]1号)

摘　　要：目的研究Mtb8.4/hIL-12嵌合基因疫苗的免疫原性及对小鼠结核杆菌感染的免疫保护效果。方法将雌性C57BL/6N小鼠40只,随机分为5组,即Mtb8.4/hIL-12嵌合基因、Mtb8.4基因疫苗组、BCG组、空载体组和PBS组。小鼠脾细胞培养上清液检测细胞因子水平;并按效靶比例分别为100∶1、50∶1、10∶1进行细胞毒性T细胞(CTL)杀伤检测。用结核杆菌H37Rv强毒株静脉攻击小鼠,计数肺和脾组织中的结核杆菌菌落数,对小鼠部分肺和脾组织作病理切片,HE染色观察组织病变程度,Z-N染色查抗酸杆菌,观察该疫苗对小鼠结核杆菌感染的免疫保护效果。结果Mtb8.4/hIL-12嵌合基因疫苗对小鼠结核杆菌感染有一定的免疫保护效果,能够诱导较强的抗原特异性Th1型细胞免疫应答,细胞因子IFN-γ和IL-2分泌增加,IL-4分泌减少,特异性CTL活性增加;使小鼠肺和脾组织中的结核杆菌菌落数较空载体组显著减少,组织病变明显减轻,其效果优于卡介苗(BCG)组和Mtb8.4基因疫苗组。结论hIL-12与Mtb8.4构建成嵌合基因疫苗后,使Mtb8.4基因疫苗的免疫效力得到很大提高。Objective To study the immunogenicity and protective effecacy of Mtb8.4/hIL-12 chimeric gene vaccine. Methods 40 C57BL/6N mice were divided into 5 groups： Mtb8.4/hlL-12 chimeric gene vaccine, Mtb8.4 gene vaccine, BCG, empty vector alone and PBS. Mice were vaccinated intramuscularly in both hind limbs three times at the intervals of three weeks or once subcutaneously with 1× 10^6 of viable M, bovis BCG Pasteur at the time of the first DNA immunization, 4 weeks after the final inoculation, three mice were sacrificed to assess cytokine response and CTL induction and the other five mice were challenged intravenously in a lateral tail vein with 1 × 10^6 CFU of virulent M, tuberculosis H37Rv. Spleen and the left lung were harvested from each mouse at 4 weeks after infection and honlogenized in sterile saline. Serial dilutions of organ homogenates were plated on L-J agar and incubated 37℃ until colonies were visible 4 weeks later. Protective efficacies in each experiment were expressed as reduced CFU and were compared with the negative control group. The right lung was obtained from each mouse and inflated with and stored in 10% formahn saline immediately. Tissues were embedded in paraffin, sectioned and stained with hematoxylin and eosin. Results Mtb8.4/hIL-12 chime-ric gene vaccine induced the secretion of more ofThl cytokines, but not IL-4 and enhanced CTL activity while BCG induced the secretion of both types of cytokines （IFN-γ, IL-2 and IL-4）. Mice immunized with Mtb8.4/hIL-12 chimeric gene vaccine had fewer and smaller tubercles than control groups. As expected, control mice had the highest bacterial loads in both lung and spleen. Immuization with Mtb8.4/hIL-12 chimeric gene vaccine could remarkably reduced CFU counts in organs. Conclusion When it was used to construct the chimeric gene vaccine, hIL-12 could improve the immune efficacy of Mtb8.4 gene vaccine. Protective efficacy

关 键 词：结核病 Mtb8.4/hIL-12 嵌合基因 基因疫苗 免疫保护 

分 类 号：R392[医药卫生—免疫学]