Du De-Wei, Jia Zhan-Sheng, Li Guang-Yu, Zhou Yong-Ying
Department of Infectious Diseases, Tangdu Hospital, the Fourth Military Medical University, Xi'an 710038, Shaanxi Province, China.
World J Gastroenterol. 2003 Jan;9(1):108-11. doi: 10.3748/wjg.v9.i1.108.
To seek for an effective method to improve the immune responses induced by DNA vaccine expressing HBV surface antigen (pCR3.1-S) in Balb/c mice (H-2(d)).
The pCR3.1-S plasmid and the eukaryotic expression vectors expressing murine IL-2 (pDOR-IL-2) or IL-12 (pWRG3169) were injected into mice subcutaneously. The immune responses to pCR3.1-S and the adjuvant effect of the cytokines plasmid were studied. Meanwhile the effect of pCR3.1-S on anti-translated subcutaneous tumor of P815 mastocytoma cells stably expressing HBsAg (P815-HBV-S) was also studied. Anti-HBs in serum was detected by enzyme-linked immunoadsorbent assay (ELISA) and HBsAg specific cytotoxic T lymphocytes (CTLs) activity was measured by (51)Cr release assay. After three weeks of DNA immunization, the cells of P815-HBV-S were inoculated into mice subcutaneously and the tumor growth was measured every five days. The survival rate and living periods of mice were also calculated.
After 8 wk DNA immunization, the A 450 nm values of sera in mice immunized with pCR3.1, pCR3.1-S and pCR3.1-S codeliveried with IL-2 or IL-12 plasmids were 0.03+/-0.01, 1.24+/-0.10, 1.98+/-0.17 and 1.67+/-0.12 respectively. Data in mice codeliveried pCR3.1-S with IL-2 or IL-12 plasmids were significantly higher than that of mice injected pCR3.1 or pCR3.1-S only. The HBsAg specific CTL activities in mice coinjected with pCR3.1-S and IL-2 or IL-12 eukaryotic expression vectors were (61.9+/-7.1) % and (73.3+/-8.8) %, which were significantly higher than that of mice injected with pCR3.1 (10.1+/-2.1) % or pCR3.1-S (50.5 +/-6.4) %. The HBsAg specific CTL activities in mice injected with pCR3.1, pCR3.1-S, pCR3.1-S combined with IL-2 or IL-12 eukaryotic expression vectors decreased significantly to (3.2+/-0.8) %, (10.6+/-1.4) %, (13.6+/-1.3) % and (16.9+/-2.3) % respectively after the spleen cells were treated by anti-CD8(+) monoclonal antibody, but presented no significant change to anti-CD4(+) monoclonal antibody or unrelated to monoclonal antibody. The HBV-S DNA vaccine (pCR3.1-S) could evidently inhibit the tumor growth, prolong the survival period of mice and improve the survival rate of mice and these effects could be improved by IL-12 gene codeliveried.
HBV DNA vaccine has a strong antigenicity in humoral and cellular immunities, which can be promoted by plasmid expressing IL-2 or IL-12. CD8+ cells executed the CTL activities. DNA vaccine may be useful for both prophylaxis and treatment of HBV infection.
寻找一种有效的方法来增强在Balb/c小鼠（H-2(d)）中表达乙肝表面抗原的DNA疫苗（pCR3.1-S）所诱导的免疫反应。
将pCR3.1-S质粒以及表达小鼠白细胞介素-2（pDOR-IL-2）或白细胞介素-12（pWRG3169）的真核表达载体皮下注射到小鼠体内。研究对pCR3.1-S的免疫反应以及细胞因子质粒的佐剂效应。同时，也研究pCR3.1-S对稳定表达乙肝表面抗原的P815肥大细胞瘤细胞（P815-HBV-S）皮下移植瘤的作用。通过酶联免疫吸附测定（ELISA）检测血清中的抗-HBs，并通过（51）Cr释放试验测定乙肝表面抗原特异性细胞毒性T淋巴细胞（CTLs）活性。DNA免疫三周后，将P815-HBV-S细胞皮下接种到小鼠体内，每五天测量肿瘤生长情况。还计算小鼠的生存率和生存期。
DNA免疫8周后，用pCR3.1、pCR3.1-S以及与白细胞介素-2或白细胞介素-12质粒共递送的pCR3.1-S免疫的小鼠血清的A450nm值分别为0.03±0.01、1.24±0.10、1.98±0.17和1.67±0.12。与白细胞介素-2或白细胞介素-12质粒共递送pCR3.1-S的小鼠的数据显著高于仅注射pCR3.1或pCR3.1-S的小鼠。与pCR3.1-S和白细胞介素-2或白细胞介素-12真核表达载体共注射的小鼠中乙肝表面抗原特异性CTL活性分别为（61.9±7.1）%和（73.3±8.8）%，显著高于注射pCR3.1（10.1±2.1）%或pCR3.1-S（50.5±6.4）%的小鼠。用抗CD8(+)单克隆抗体处理脾细胞后，注射pCR3.1、pCR3.1-S、pCR3.1-S与白细胞介素-2或白细胞介素-12真核表达载体组合的小鼠中乙肝表面抗原特异性CTL活性分别显著降至（3.2±0.8）%、（10.6±1.4）%、（13.6±1.3）%和（16.9±2.3）%，但用抗CD4(+)单克隆抗体或无关单克隆抗体处理后无显著变化。乙肝表面抗原DNA疫苗（pCR3.1-S）可明显抑制肿瘤生长，延长小鼠生存期并提高小鼠生存率，且这些作用可通过共递送白细胞介素-12基因得到增强。
乙肝DNA疫苗在体液免疫和细胞免疫中具有较强的抗原性，可通过表达白细胞介素-2或白细胞介素-12的质粒来增强。CD8+细胞执行CTL活性。DNA疫苗可能对乙肝感染的预防和治疗均有用。
